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Anselme, Marc Joseph. "Immobilized yeast reactor for ethanol production." Thesis, Georgia Institute of Technology, 1985. http://hdl.handle.net/1853/11706.
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Namthabad, Sainath, and Ramesh Chinta. "Robust Encapsulation of Yeast for Bioethanol Production." Thesis, Högskolan i Borås, Institutionen Ingenjörshögskolan, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:hb:diva-17499.
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In the future the demand for ethanol is expected to increase greatly due to the rising energy requirements in the world. Lignocellulosic materials are a suitable and potentially cheap feedstock for sustainable production of fuel ethanol, since vast quantities of agricultural and forest residues are available in many countries. However, there are several problems involved in the utilization of lignocellulosic raw materials as sugar source. The most common way of releasing the simple sugars in the material is by dilute acid hydrolysis. This procedure is relatively simple and cheap, but in addition to the sugars it creates inhibitory compounds. These inhibitors make it very hard for the yeast to ferment the hydrolyzate and detoxification is often necessary. One way to overcome this problem is to encapsulate the yeast. Encapsulation is an attractive method since it improves the cells stability and inhibitor tolerance, increases the biomass amount inside the reactor, and decreases the cost of cell recovery, recycling and downstream processing. However, the method does not yet permit long-term cultivation since the capsules used so far are not robust enough. Therefore more studies have to be conducted in order to find methods which produce mechanically robust capsules. The main goal of this paper is to find a suitable method to produce robust capsules using different concentration of the chemicals at different pH and also implementing some modifications such as addition of cross-linkers in preparation procedure. In this paper comparison of three different encapsulation techniques were studied based on the mechanical robustness of the capsules. The three different techniques were calcium mineralized alginate-chitosan capsules, alginate capsules coated with 2% chitosan (2% AC) and genipin crosslinked alginate-chitosan (GCAC) capsules. The results indicate that GCAC capsules are most robust and were good enough for prolonged use since most of the capsules were not deformed in mechanical strength test. There were slight differences in the diameter and membrane thickness before and after swelling. No negative influence was observed on the yeast growth when applying the cross-linker. The results of this study will hopefully add valuable information and helps in further studies using other cross-linkers to prepare robust capsules.Program: Industrial Biotechnology
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Armstrong, Gareth Owen. "The production of resveratrol by wine yeast." Thesis, Stellenbosch : Stellenbosch University, 2001. http://hdl.handle.net/10019.1/52557.
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Thesis (MSc)--Stellenbosch University, 2001.ENGLISH ABSTRACT: Grapevine is constantly under attack from a wide variety of pathogens including viruses, bacteria and fungi. In order to ensure survival, the grapevine has developed a vast array of defense mechanisms to combat invading organisms. A key element of this disease resistance is the production of phytoalexins, of which resveratrol is the primary component. The synthesis of resveratrol, together with other structural and biochemical defense mechanisms equips the plant to combat a number of pathogens resulting in the production of healthy grapes for the vinification of top quality wine. As part of the active disease response resveratrol is synthesised de novo in the berry skin at the site of infection, on recognition of the pathogen. Here it is able to limit the damage caused by the pathogen as well as preventing it from spreading. This gives the plant the opportunity to initiate its systemic acquired resistance thereby protecting the rest of the plant and preventing secondary infections. The fermentation of red wine on the grape skins allows for the extraction of resveratrol from the skin into the wine. Red wines therefore have a significantly higher concentration of resveratrol than white varieties, which contain little or no resveratrol at all. It is for this reason that the moderate consumption of wine, in particular red wine, is synonymous with a healthy lifestyle. The antioxidant and anti-inflammatory activities of resveratrol are important contributors to the cardiovascular benefits derived from the consumption of red wine. It now seems, however, that significant cardiovascular protection is derived from the synergistic action of resveratrol, the polyphenols and the alcohol in wine. With the wholesomeness of any food or beverage being of extreme importance, the aim of this project was to manipulate wine yeast to produce resveratrol during fermentation. This required the introduction of an entire metabolic pathway, by integrating plant genes into the yeast. Resveratrol synthase utilises three malonyl-CoA and one pcoumaroyl- CoA molecules to produce one molecule of resveratrol, Saccharomyces cerevisiae produces malonyl-CoA but no p-coumaroyl-CoA. Therefore, the following genes were obtained to enable yeast to produce p-coumaroyl-CoA: PAL, encoding phenylalanine ammonia-lyase to convert phenylalanine into cinnamic acid; C4H, encoding cinnamate-4- hydroxlyase to convert cinnamic acid into p-coumaric acid; and 4CL9 or 4CL216 encoding CoA-ligases to convert the p-coumaric acid into p-coumaroyl-CoA. To attain high-level expression, the genes were subcloned under the control of the phosphoglycerate kinase gene (PGK1) promoter and terminator. Due to integration problems with these expression cassettes and the fact that the yeast was able to consume p-coumaric acid, the 4CL9, 4CL216 and Vst1 (encoding resveratrol synthase) genes were subcloned under the control of the alcohol dehydrogenase (ADH2) and PGK1 promoters into episomal plasmids, respectively. A laboratory yeast strain containing both the Vst1 and 4CL9, or the Vst1 and 4CL216 genes was evaluated for its ability to utilise p-coumaric acid and produce resveratrol. Northem analysis confirmed that the Vst1, 4CL9 and 4CL216 genes were transcribed and over-expressed compared to the control strain. The transformants expressing the CoA-ligase genes utilised the p-coumaric acid faster than the control, although it was not possible to determine whether p-coumaroyl-CoA was produced. No resveratrol was produced under the assay conditions used. The results indicated that the yeast is unable to produce active resveratrol synthase, which is required to catalyse the final reaction in the production of resveratrol. Posttranslational modification, such as overglycosylation and disulphide formation, of the heterologous protein in yeast has been indicated as the possible reason for the lack of enzyme activity. This introduces an exciting area of research for the development of biotechnological tools with the ability to increase the production of active heterologous proteins in yeast.
AFRIKAANSE OPSOMMING: Wingerde word voortdurend deur 'n groot verskeidenheid patogene, insluitende virusse, bakteriee en swamme, aangeval. Ten einde oorlewing te verseker, het die wingerdstok In wye reeks verdedigingsmeganismes ontwikkel om weerstand te bied teen indringerorganismes. 'n Belangrike faktor in hierdie weerstand teen siektes is die produksie van fitoaleksiene, waarvan resveratrol die hoofkomponent is. Oeur die sintese van resveratrol, asook ander strukturele en biochemiese verdedigingsmeganismes, word die plant toegerus om weerstand te kan bied teen In hele aantal patogene ten einde gesonde druiwe te produseer wat gebruik kan word vir die vinifikasie van topgehalte wyn. As deel van die aktiewe reaksie teen siektes, word resveratrol de novo in die dop van die korrel by die plek van infeksie gesintetiseer sodra 'n patogeen herken word. Hier kan dit die skade deur die patogeen veroorsaak, beperk en verhoed dat dit versprei. Oit gee aan die plant die geleentheid om sy sistemies-verworwe weerstand te inisieer, en daardeur die res van die plant te beskerm, sowel as sekondere infeksies te verhoed. Die fermentasie van rooiwyn op die druifdoppe maak voorsiening vir die ekstraksie van resveratrol uit die dop na die wyn. Die konsentrasie van resveratrol in rooiwyn is dus beduidend hoer as in die wit varietelte, wat geen of baie min resveratrol bevat. Oit is dan juis die rede waarom die matige inname van wyn, veral rooi wyn, gesien word as In integrale deel van 'n gesonde leefwyse. Resveratrol se aktiwiteit as antioksidant en antiinflammatoriese middel lewer In belangrike bydrae tot die kardiovaskulere voordele wat verkry word uit die inname van rooiwyn. Oit blyk egter nou dat die beduidende kardiovaskulere beskerming gesetel is in die sinergistiese werking van resve ratro I, die polifenole en die alkohol in wyn. Aangesien die heilsaamheid van enige voedsel of drank van die uiterste belang is, was dit die doel van hierdie projek om wyngis te manipuleer ten einde tydens die fermentasieproses resveratrol te produseer. Hiervoor moes 'n volledige metaboliese pad daargestel word deur plantgene in die gis te inkorporeer. Resveratrol-sintase maak gebruik van drie maloniel-KoA-molekules en een p-kumarotel-Kos-molekule om een molekule resveratrol te produseer. Saccharomyces cerevisiae produseer maloniel-KoA, maar nie p-kumaroiel-Kcs, nie. Oie volgende gene is dus aangewend om die gis in staat te stel om p-kumarolel-Koe, te produseer: PAL, wat fenielalanien-ammoniak-liase enkodeer om fenielalanien om te sit na kaneelsuur; C4H, wat sinnamaat-4-hidroksliase enkodeer om kaneelsuur om te sit na p-kumaarsuur; en 4CL9 of 4CL216 wat KoA-ligases enkodeer om p-kumaarsuur om te sit na p-kumarolel-Kos, Om hoevlak-uitdrukking te verkry, is die gene gesubkloneer onder beheer van die fosfogliseraat-kinase-geen(PGK1)- promotor en -terminator. As gevolg van integrasieprobleme met hierdie uitdrukkingskassette en die feit dat die gis die p-kumaarsuur kon verteer, is die 4CL9-, 4CL216- en Vst1- (wat resveratrol-sintase enkodeer) gene na episomale plasmiede gesubkloneer onder beheer van die alkohol-dehidrogenase(ADH2)- en PGK1-promotors onderskeidelik. 'n Laboratorium-gisstam wat 6f beide die Vst1-geen en die 4CL9-geen, 6f die Vst1-geen en die 4CL216-geen bevat het, is geevalueer vir die verrnoe om pkumaarsuur te benut en resveratrol te produseer. Noordelike klad analises het bevestig dat die Vst1-, 4CL9- en 4CL216-gene getranskribeer en ooruitgedruk was in vergelyking met die kontrole-stam. Die transformante wat die KoA-ligases uitgedruk het, het die pkumaarsuur vinniger benut as wat die kontrole dit gedoen het, alhoewel dit nie moontlik was om vas te stel of o-kurnarotel-Kos, geproduseer is nie. Met die essai-kondisies wat gebruik is, is geen resveratroI geproduseer nie. Die resultate het daarop gedui dat die gis nie daartoe in staat is om aktiewe resveratrol-sintase, wat nodig is vir die katalise van die finale reaksie in die produksie van resveratrol, te produseer nie. Naomsettingsmodifikasies van die heteroloe protelen in die gis, soos oor-glikosilasie en disulfiedvorming, is aangewys as die moontlike rede vir die gebrek aan ensiemaktiwiteit. Dit stel In opwindende veld vir verdere navorsing voor, naamlik die ontwikkeling van biotegnologiese middele met die vermoe om die produksie van aktiewe heteroloe protelene in gis te verhoog.
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Thipayarat, Aluck. "Production of human serum albumin by immobilized yeast." Related electronic resource: Current Research at SU : database of SU dissertations, recent titles available full text, 2002. http://wwwlib.umi.com/cr/syr/main.
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Grant, Stephanie Mary. "Production of astaxanthin by the yeast Phaffia rhodozyma." Thesis, Queen's University Belfast, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.324833.
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Fairbairn, Samantha. "Stress, fermentation performance and aroma production by yeast." Thesis, Stellenbosch : Stellenbosch University, 2012. http://hdl.handle.net/10019.1/20336.
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Thesis (MSc)--Stellenbosch University, 2012.ENGLISH ABSTRACT: Yeast strains contend with numerous stresses during winemaking. An inability to perceive and initiate the physiological changes needed to adapt to stress, has been linked to slow or incomplete (residual sugar > 4 g/L) fermentations. Wine yeast strains differ in genotype; this is manifested as differences in their stress tolerance, and fermentation performance. The first goal of this study was to evaluate how the initial sugar (200 or 240 g/L) and nitrogen (50, 100, 250, or 400 mg/L) content, and the fermentation temperature (15°C or 20°C) affected the fermentation performance of 17 commercial wine yeast strains. Fermentation performance was evaluated based on the fermentation kinetics (lag phase, maximum fermentation rate and total weight loss by CO2 evolution), residual sugar content and yeast dry weight. The results demonstrate that the fermentation performances of commercial yeast cultures are significantly and differently affected by initial nitrogen and sugar levels, as well as the fermentation temperature. Additionally, excess nitrogen had a negative impact on the fermentation kinetics and sugar consumption. Nitrogen deficiency is a common cause of slow and incomplete fermentations, as it affects yeast growth and thus fermentation rates. Nitrogen supplements are routinely added at the onset of fermentation, reducing the risk of problematic fermentations. Therefore characterising the fermentative ability of a strain over a range of oenologically relevant conditions, could aid winemakers in selecting a yeast strain capable of fermenting a grape must (of known sugar and nitrogen levels) to completion at the desired fermentation temperature. Investigations on fermentation related stress generally focus on its influence on fermentation rate and sugar consumption. However, from a winemaking perspective, the strain’s ability to produce the desired volatile aroma compounds is equally important. Yet, literature provides little insight into the influence stress has on the volatile aroma profile; this is surprising as wine aroma is closely linked to wine quality and consumer liking. The final goal of this study was to evaluate changes to the volatile aroma profiles produced by five commercial yeast strains, in response to hyperosmotic and temperature stress. The concentrations of the aroma compounds were quantified using a gas chromatograph coupled to a flame ionization detector. The results show that hyperosmotic and temperature stress caused significant changes in the levels of a number of aroma compounds. Furthermore, the changes observed differed among the evaluated strains, as well as for the fermentation stress treatments studied. Future aims should be directed towards the potential application of yeast strain selection as a means to avoid problematic fermentations in grape must; in addition to the further characterisation of the relationship between stress and the resultant volatile aroma profile in wine.
AFRIKAANSE OPSOMMING: Gisrasse moet verskeie stresfaktore afweer tydens die wynmaak proses. Die onvermoë van ‘n wyngis om stres waar te neem en die nodige fisiologiese veranderinge te inisieer om aan te pas by die strestoestande word met slepende of onvolledige fermentasies (met ‘n residuele suiker van meer as 4 g/L) geassosieer. Wyngisrasse verkil in genotipe; wat as groot verskille in die graad van strestoleransie, en dus ook fermentasie sukses geopenbaar word. Die eerste doelwit van hierdie studie was om te evalueer hoe die suiker (200 of 240 g/L) en stikstof (50, 100, 250, of 400 mg/L), asook die fermentasie temperatuur (15°C of 20°C) die fermentasie prestasie van 17 kommersiële wyngiskulture beïnvloed. Die sukses van fermentasie is geëvalueer op grond van fermentasie kinetika (sloerfase, maksimum fermentasiespoed en totale gewigsverlies as CO2 verlies), die residuele suiker inhoud en die gis droë massa. Die resultate demonstreer dat die fermentasie sukses van kommersiële giskulture beduidend en verskillend beïnvloed word deur die aanvangsstikstof en – suikerkonsentrasies, asook die fermentasie temperatuur. Daarbenewens, wanneer stikstof in oormaat teenwoordig is kan dit ‘n negatiewe impak op fermentasietempo en suiker metabolisme hê. Beperkende vlakke van stikstof ‘n algemene oorsaak van slepende of onvolledige fermentasies, aangesien stikstof die groei en gevolglik ook die fermentasiespoed van gis beïnvloed. Stikstofaanvullings word dikwels tot druiwemos toegevoeg aan die begin van gisting, wat die risiko van probleemfermentasies verlaag. Dus kan die karakterisering van die fermentasievermoë van ‘n gisras vir ‘n reeks wynkundig relevante kondisies die wynmaker help om ‘n gisras te selekteer wat in staat is om ‘n druiwemos (waarvan die suiker en stikstofvlakke bekend is) droog te gis by die gewenste temperatuur. Meeste studies wat fermentasieverwante stress ondersoek, fokus op die die invloed daarvan op fermentasietempo en suikerverbruik. Van ‘n wynmaakperspektief is die gis se vermoë om die gewensde vlugtige aroma komponente te produseer egter ewe belangrik as die vermoë om fermentasie te voltooi. Tog verskaf die literatuur min insig tot die invloed van stres op die vlugtige aromaprofiel; wat verbasend is aangesien die aromaprofiel ‘n belangrike faktor is van die waargenome wynkwaliteit en daarom ook verbruikersvoorkeur. Die finale doelwit van hierdie projek was om die veranderinge tot die vlugtige aromaprofiel geproduseer deur vyf kommersiële gisrasse in reaksie op hiperosmotiese stres en temperatuur stres te evalueer. Die konsentrasies van die aromakomponente is gekwantifiseer deur gas chromatografie gekoppel aan vlam‐ioniserende deteksie. Die resultate wys dat hiperosmotiese‐ en temperatuur stres beduidende veranderinge meebring in die vlakke van ‘n aantal aromakomponente. Verder is die waargenome veranderinge ook verskillend vir die geëvalueerde gisrasse, asook vir die verskille stresbehandelings wat ondersoek is. Toekomstige studies behoort gerig te wees op die toepassing van gis seleksie om potensiële probleemfermentasies in druiwemos te voorkom; asook die verdere karakterisering van die verhouding tussen omgewingstresfaktore en die gevolglike vlugtige aromaprofiel in wyn.
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Hemmati, Naghmeh. "Engineering yeast strains to enhance bioethanol production efficiency /." Available to subscribers only, 2008. http://proquest.umi.com/pqdweb?did=1674956301&sid=4&Fmt=2&clientId=1509&RQT=309&VName=PQD.
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Tai-Wong, Sue Mei. "Origin and genetic manipulation of brewing lager yeast." Thesis, University of Oxford, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.249282.
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McCormack, P. J. "The ecological significance of antibiotic production to yeasts and yeast-like organisms on the phylloplane." Thesis, University of Kent, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.304835.
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Altabet, Altaher Ibrahim. "Siderophore and pigment production by Candida albicans." Thesis, University of Glasgow, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.360169.
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Borrull, Riera Anna. "Yeast stress responses to acclimation for sparkling wine production." Doctoral thesis, Universitat Rovira i Virgili, 2016. http://hdl.handle.net/10803/396076.
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La present tesi doctoral pretén profunditzar en el coneixement sobre la resposta estrès dels llevats durant el procés d’aclimatació per realitzar la segona fermentació per la producció de vins escumosos. El principal estrès, l’etanol, té un efecte sinèrgic amb molts altres estrès. Tot i que aquest procés dels llevats, que té lloc abans de ser inoculats a la segona fermentació, ha estat realitzat a les bodegues durant anys, la fase de latència després de la inoculació i la petita proliferació cel•lular indican que els llevats no estan completament adaptats al medi. Un escenari tant complex com aquest hauria de ser estudiat per evitar cèl•lules susceptibles a morir i que puguin donar lloc a una parada de fermentació abans d’acabar-la. Es va fer un estudi citològic, metabòlic i lipídic de les cèl•lules aclimatades per destacar les condicions (com la disponibilitat de sucre i oxigen) per obtenir un millor diagnòstic cel•lular, així com els factors clau (com l’activitat i morfologia vacuolar, les reserves de glúcids, les espècies reactives d’oxígen o la rigidesa de membrana) que podrien confirmar que aquells llevats estan preparats per realitzar al segona fermentació.La presente tesis doctoral pretende profundizar en el conocimiento sobre la respuesta estrés de las levaduras durante el proceso de aclimatación para la realización de la segunda fermentación para producir vinos escumosos. El principal estrés, el etanol, tiene un efecto sinérgico con muchos otros estreses. A pesar de que este proceso, que tiene lugar antes de la inoculación a la segunda fermentación, ha sido realizado en bodegas durante muchos años, la fase de latencia después de la inoculación, así como la pequeña proliferación celular indican que las levaduras no están completamente adaptadas al medio. Un escenario tan complejo como este debería ser estudiado para evitar células susceptibles a morir y que puedan dar lugar a una parada en la fermentación antes de acabarla. Se hizo un estudio citológico, metabólico y lipídico de las células aclimatadas para poder destacar las condiciones (como la disponibilidad de azúcar y oxígeno) para obtener un mejor diagnostico celular, así como los factores clave (como la actividad y morfología vacuolar, las reservas de carbohidratos, las especies reactivas de oxígeno o la rigidez de membrana) que podrían confirmar que esas levaduras están preparadas para realizar la segunda fermentación.
he present doctoral thesis deepends knowledge of the stress responses of yeast during acclimation process to perform secondary fermentation for sparkling wine production. The main stress factor, ethanol has as synergic effect with many others. Even acclimation process of yeast cells before being inoculated into secondary fermentation have been done by winemakers for ages, the lag phase after inoculation and the short cell proliferation indicate cells yeast are not completely adapted. Such a complex scenario should be studied to avoid cells susceptible to death which would stop fermentation before finishing. A cytological, metabolomic and lipidomic study has been done of acclimated cells; pointing out the conditions (as sugar and oxygen availability) to get the best cellular diagnostic as possible and the key factors (as vacuolar activity and morphology, carbohydrate reserves, reactive oxygen spices accumulation or membrane rigidity) which could confirm yeasts are ready to perform secondary fermentation.
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Zhang, Zhigen. "Bioreactor studies of heterologous protein production by recombinant yeast." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq21404.pdf.
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Jakkamsetty, Chamundeshwari, and Chaitanya Medapudi. "Stress tolerance of encapsulated yeast used for bioethanol production." Thesis, Högskolan i Borås, Akademin för textil, teknik och ekonomi, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:hb:diva-9745.
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Environmental and economic issues have drawn the world’s attention to produce and utilize energy from renewable sources for sustainable development. One of the attempts includes the production of ethanol from various substrates. Many researchers have focused on utilizing lignocelluloses biomass as substrate for the production of ethanol, which mainly contains cellulose and is a cheap and abundantly available material in the world. One of the major problems faced by researchers during production of ethanol from the lignocellulosic biomass is the stress tolerance of yeast cells, due to the nature of the hydrolysed substrate (lignocellulosic material treated with Nitro methyl cellulose (NMC)). One of the solutions for this problem is to encapsulate the yeast cells. Encapsulation is an attractive method, which can enhance the stress tolerance of the yeast cells in the reactor, and also aid in maintaining a high yeast concentration inside the bioreactor and thereby increase the volumetric productivity of ethanol. This report includes a major study on the sodium chloride and ethanol stress tolerance of alginate chitosan alginate (ACA), alginate chitosan (AC) and APTES treated ACA encapsulated yeast biomass in medium containing different concentrations of glucose under anaerobic conditions. AC capsules shows significant results towards osmotic stress and ethanol stress compared with that of freely suspended cells in stress conditions.AC capsule encapsulated yeast tolerated osmotic stress better than ACA capsules in 2M of NaCl where as freely suspended yeast cells unable to tolerate 2M of NaCl . At 100th hour in AC capsules glucose consumption was 12 g/l where as in ACA capsules glucose consumption at same 100th hour was 2 g/l. At 10% ethanol concentration yeast inside ACA capsules showed 5 g/l of glucose consumption but in freely suspended yeast cells there is no glucose consumption as they cannot tolerate higher stress levels.
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Bawa, Zharain. "Improving recombinant human adenosine A2A receptor production in yeast." Thesis, Aston University, 2014. http://publications.aston.ac.uk/23176/.
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Over 50% of clinically-marketed drugs target membrane proteins; in particular G protein-coupled receptors (GPCRs). GPCRs are vital to living cells, performing an active role in many processes, making them integral to drug development. In nature, GPCRs are not sufficiently abundant for research and their structural integrity is often lost during extraction from cell membranes. The objectives of this thesis were to increase recombinant yield of the GPCR, human adenosine A2A receptor (hA2AR) by investigating bioprocess conditions in large-scale Pichia pastoris and small-scale Saccharomyces cerevisiae cultivations. Extraction of hA2AR from membranes using novel polymers was also investigated. An increased yield of hA2AR from P. pastoris was achieved by investigating the methanol feeding regime. Slow, exponential feed during induction (μlow) was compared to a faster, exponential feed (μhigh) in 35 L pilot-scale bioreactors. Overall hA2AR yields were increased for the μlow cultivation (536.4pmol g-1) compared to the μhigh148.1 pmol g-1. hA2AR levels were maintained in cytotoxic methanol conditions and unexpectedly, pre-induction levels of hA2AR were detected. Small-scale bioreactor work showed that Design of Experiments (DoE) could be applied to screen for bioprocess conditions to give optimal hA2AR yields. Optimal conditions were retrieved for S. cerevisiae using a d-optimal screen and response surface methodology. The conditions were 22°C, pH 6.0, 30% DO without dimethyl sulphoxide. A polynomial equation was generated to predict hA2AR yields if conditions varied. Regarding the extraction, poly (maleic anhydride-styrene) or PMAS was successful in solubilising hA2AR from P. pastoris membranes compared with dodcecyl-β-D-maltoside (DDM) detergent. Variants of PMAS worked well as solubilising agents with either 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) or cholesteryl hemisuccinate (CHS). Moreover, esterification of PMAS improved solubilisation, suggesting that increased hydrophobicity stabilises hA2AR during extraction. Overall, hA2AR yields were improved in both, P. pastoris and S. cerevisiae and the use of novel polymers for efficient extraction was achieved.
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Rizzi, John. "Production of emulsifier by Torulopsis petrophilum." Thesis, McGill University, 1987. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=64014.
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Ogunlabi, Olugbenga. "Metabolic engineering of yeast (Saccharomyces cerevisiae) with a view to optimising butanol production." Thesis, University of Manchester, 2018. https://www.research.manchester.ac.uk/portal/en/theses/metabolic-engineering-of-yeast-saccharomyces-cerevisiae-with-a-view-to-optimising-butanol-production(6820196d-30e9-43fa-a3ee-8983c82374a5).html.
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Global energetic and environmental concerns have generated interest in the biological systems for the production liquid biofuels. Butanol is one such biofuel, which can be naturally produced by some Clostridia species. However, possible limitations in Clostridial engineering and large-scale fermentation have led to an examination of other potential organisms that might house this pathway for butanol production. As a robust industrial host and key model organism in the study of fundamental biological processes, the yeast Saccharomyces cerevisiae has been used to house the Clostridial ABE-butanol pathway. However, butanol yields and titres in this yeast are relatively low. Therefore, in this thesis, three distinct strategies were carried out with the goal of optimising butanol production in the strain of yeast (previously constructed in the Ashe lab) bearing the ABE-butanol pathway: 1. Mutation of genes involved in the regulation of carbon source usage. 2. Deletion of genes where the product is involved in the consumption of cytosolic acetyl-CoA (the starting precursor for the butanol synthetic pathway). 3. Targeted mutagenesis to improve the efficiency of the thiolase enzyme, which catalyses the condensation of 2x acetyl-CoA to initiate the ABE-butanol synthesis pathway. The results showed the first two strategies did not lead to improvements in butanol yields. However, increases of intracellular acetyl-CoA were observed in some mutant strains, even though butanol production did not increase in these strains. In order to make maximum use of the accumulating cytosolic acetyl-CoA, thiolase engineering in the butanol production yeast strain was pursued. The introduced changes caused an increase in butanol (about two fold). Overall, this project has used a minimal engineering approach by modulation of associated pathways or optimisation of the heterologous enzyme with a view to improve butanol production in yeast. To achieve high and scalable butanol production in yeast, a robust approach involving whole synthetic biology - Design, Build Test, and Learn will need to be adopted to create a more efficient yeast-butanol system.
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Trollope, Kim. "Investigation of resveratrol production by genetically engineered Saccharomyces cervisiae strains /." Link to the online version, 2006. http://hdl.handle.net/10019/1247.
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Wardrop, Forbes Robert. "The physiology of industrial yeast in continuous culture." Thesis, Abertay University, 1999. https://rke.abertay.ac.uk/en/studentTheses/61c9f775-b287-4b25-b92c-409c78252572.
Full textAbstract:
The growth and physiology of Saccharomyces cerevisiae GB4918 (baker’s yeast) was studied under glucose-limitation in chemostat culture. Levels of lg/1 (0.1% w /v) glucose allowed cell growth while preventing fermentation in a defined medium (QEMM3). Metabolism of glucose by respiration or fermentation was shown to affect the mean cell volume, with fermentative use of glucose causing an increase in cell size. This was also a major physiological difference between S. cerevisiae GB4918 (a Crabtree positive yeast) and Kliiyveromyces marxianus DBVPG 6165 (a Crabtree negative yeast). The ability of the Crabtree positive yeast to substantially increase its mean cell volume was also reflected in a 5-fold greater consumption of glucose, reduced biomass yield and increased ethanol yield, compared with the Crabtree negative K . marxianus. Growth of both these yeasts was seen in 50g/l glucose in the presence of the respiratory inhibitor, antimycin A. This was evident by the switching to fermentation in K . marxianus, and the complete fermentation of glucose by S. cerevisiae. The growth and physiology of S. cerevisiae GB4918 was also established in glucose-limited chemostat cultures, with special regard to the intracellular macromolecular compounds that are relevant to industrial yeast biomass production. This showed that in respiring cultures of S. cerevisiae, increasing growth rate resulted in decrease in both trehalose and glycogen content, while increasing protein and RNA. This is true until μmax (in this context the growth rate at which respiro-fermentativemetabolism occurs) when accumulation of trehalose and glycogen is apparent. Once μerit (growth rate at which washout of the culture begins) was reached then biomass significantly reduced. In describing the steady-state condition of baker’s yeast it was then possible to describe changes occurring in yeast when subjected to a variety of nutrient perturbations. With a lactic acid (2% v/v) perturbation there were dramatic effects on both growth and metabolism at a growth rate of 0.12h_1, but significant decreases in biomass and protein, and significant increases in trehalose and glycogen. At a higher growth rate (0.22h_1) the effect was much severer on protein content, and on reduced levels of trehalose and glycogen. The effect of perturbing the cultures with elevated levels of calcium was also most significant on reducing yeast trehalose and glycogen levels, probably due to inhibition of the biosynthesis of these compounds. Zinc additions to chemostat cultures acted to increase the levels of protein in the cells,while having little effect on any of the other cellular macromolecules. This suggests that increasing calcium levels during the latter stages of yeast propagations may produce a yeast with reduced stress responses. Increased zinc may also encourage a greater protein content, which would, in turn, provide a better nutritive content for both protein and amino acids in yeasts destined for use as a food additive.
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ZHUANG, XUN. "ENGINEERING NOVEL TERPENE PRODUCTION PLATFORMS IN THE YEAST SACCHAROMYCES CEREVISIAE." UKnowledge, 2013. http://uknowledge.uky.edu/pss_etds/17.
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The chemical diversity and biological activities of terpene and terpenoids have served in the development of new flavors, fragrances, medicines and pesticides. While terpenes are made predominantly by plants and microbes in small amounts and as components of complex mixtures, chemical synthesis of terpenes remains technically challenging, costly and inefficient. In this dissertation, methods to create new yeast lines possessing a dispensable mevalonate biosynthetic pathway wherein carbon flux can be diverted to build any chemical class of terpene product are described. The ability of this line to generate diterpenes was next investigated. Using a 5.5 L fed bath fermentation system, about 569 mg/L kaurene and approximately 207 mg/L abietadiene plus 136 mg/L additional isomers were achieved. To engineer more highly modified diterpenes might have greater industrial, agricultural or medicinal applications, kaurenoic acid production reached 514 mg/L with byproduct kaurene and kaurenal at 71.7mg/L and 20.1mg/L, respectively, in fed batch fermentation conditions. Furthermore, ZXM lines for engineer monoterpene and ZXB lines for engineer triterpene were generated by additional specific genomic modification, 84.76 ±13.2 mg/L linalool, 20.54±3.8 mg/L nerolidol and 297.7mg/L squalene were accumulate in ZXM144 line ana ZXB line, respectively, in shake flask conditions.
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Chamberlain, H. "Production of oraganoleptic compounds by distillers' yeast in continuous culture." Thesis, University of Glasgow, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.381361.
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Tai, Mitchell. "Metabolic Engineering of oleaginous yeast for the production of biofuels." Thesis, Massachusetts Institute of Technology, 2012. http://hdl.handle.net/1721.1/76486.
Full textAbstract:
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemical Engineering, 2012.Cataloged from PDF version of thesis.
Includes bibliographical references.
The past few years have introduced a flurry of interest over renewable energy sources. Biofuels have gained attention as renewable alternatives to liquid transportation fuels. Microbial platforms for biofuel production have become an attractive option for this purpose, mitigating numerous challenges found in crop-based production. Towards this end, metabolic engineering has established itself as an enabling technology for biofuels development. In this work we investigate the strategies of metabolic engineering for developing a biodiesel production platform, utilizing the oleaginous yeast Yarrowia lipolytica as the host organism. We establish new genetic tools for engineering Y. lipolytica beginning with an expression vector utilizing the genetic features from translation elongation factor 1-a (TEF). Additionally, a complementary plasmid was developed allowing for multiple plasmid integration. Bioinformatics analysis of intronic genes in hemiascomycetous yeast also identified relationships between functional pathways and intron enrichment, chronicling the evolutionary journey of yeast species. Next gene targets were examined within the lipid synthesis pathway: acetyl-coA carboxylase (ACC), delta9-desaturase (D9), ATP citrate lyase (ACL), and diacylglycerol acyltransferase (DGA). A combinatorial investigation revealed the order of contribution to lipid overproduction (from strongest to weakest): DGA, ACC, D9, ACL. Scale-up batch fermentation of selected strains revealed exceptionally high lipid accumulation and yield. These results demonstrate the balance between cellular growth and lipid production which is being modified through these genetic manipulations. We next explored utilization of alternative substrates to expand the capabilities and utility of Y. lipolytica. For xylose, a prevalent substrate in cellulosic feedstocks, expression of the redox pathway from Scheffersomyces stipitis and adaptation led to successful substrate utilization. Through the use of cofermentation, growth and productivity on xylose was improved dramatically with xylose-to-lipids conversion successfully demonstrated. For acetate, a potentially useful substrate for electrofuel production, lipid production using our strongest performing strain resulted in high lipid accumulation and yield. From this study, metabolic engineering of Y. lipolytica was successfully used to achieve exceptional lipid overproduction from a variety of substrates. Our genetic tools and recombinant strains establish a strong platform for the study and development of microbial processes for the production of biofuels.
by Mitchell Tai.
Ph.D.
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Silverman, Andrew Michael. "Metabolic engineering strategies for increasing lipid production in oleaginous yeast." Thesis, Massachusetts Institute of Technology, 2015. http://hdl.handle.net/1721.1/103274.
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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemical Engineering, 2015.Page 11 out of sequence; inserted between page 4 and page 5. Page 209 blank. Cataloged from PDF version of thesis.
Includes bibliographical references.
Although petroleum and other fossil fuels have traditionally been used to fulfill our energy needs, rising concerns over energy security and the climate-changing effects of our continual greenhouse gas emissions have led to great interest in developing a domestic source of renewable fuel with low net carbon emissions. Biodiesel is an attractive option for replacing petroleum-based fuels used in the transportation sector due to its compatibility with existing infrastructure. Single cell oils from heterotrophic oleaginous microorganisms as a source of bio diesel allow for high productivity from a wide array of potential feedstocks, including agroindustrial and municipal waste streams. The goal of this work is to use the tools of rational metabolic engineering to improve lipid production in the non-conventional oleaginous yeast Yarrowia lipoytica on two representative carbon sources, glucose and acetate. Previous work in this area achieved considerable success with the simultaneous overexpression of the native acetyl-CoA carboxylase (ACC 1) and diacylglycerol acyltransferase (DGA2) genes; the resulting strain was used as a benchmark to evaluate our own efforts. We began with the compilation of a set of 44 genes and evaluated the effects of the individual overexpression of each gene on the ability of the resulting strain to produce lipids in fermentations of glucose and acetate. The genes tested here represent many different functions potentially important to lipid production, including the Kennedy pathway, fatty acid synthesis, central carbon metabolism, NADPH generation, regulation, and metabolite transport. Our results demonstrate that a diverse subset of gene overexpressions led to significant improvements in lipid production on at least one substrate. The largest improvements unsurprisingly came from overexpressing genes directly related to triacylglycerol synthesis, such as diacylglycerol acyltransferase DGAI, which on glucose increased the lipid titer, content and yield by 236%. 165%, and 246%, respectively, over our wild-type control strain, and the acylglycerolphosphate acyltransferase SLC1 gene, which increased titer/content/yield on glucose by 86%/73%/87% and on acetate by 99%/91%/151%. Significant improvements were also detected from genes that more indirectly effect lipogenesis, such as glycerol-3-phosphate dehydrogenase GPD (which produces head groups for triacylglycerol molecules) and the 6-phosphogluconolactoase SOL3 (catalyzing the middle step of the NADPH-producing oxidative pentose phosphate pathway). We next chose the aforementioned SLCl, GPD, and SOL3 genes for use in continued rational engineering of our benchmark strain due to the significance of their effects and the lack of redundancy in their likely mechanism of improving lipogenesis when overexpressed along with ACC I and DGA2. The results of this investigation indicate that the strain overexpressing ACC 1, DGA2,'and SLC 1 may be superior to our benchmark strain, increasing lipid content and yield by 24% and 20%, respectively, with a statistically equivalent titer on acetate. This strain produces the highest reported overall lipid yield of an oleaginous yeast on acetate, at 0.207 g lipids/g acetate.
by Andrew Michael Silverman.
Ph. D.
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Ndlovu, Thulile. "Mannoprotein production and wine haze reduction by wine yeast strains." Thesis, Stellenbosch : Stellenbosch University, 2012. http://hdl.handle.net/10019.1/71938.
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Thesis (PhD)--Stellenbosch University, 2012.ENGLISH ABSTRACT: Wine protein haze formation is a major challenge for wine makers, and several wine clarifying agents such as bentonite are used in the industry to protect wine from this occurrence. However, clarifying agents may have an undesirable impact on wine quality. Yeast mannoproteins have been shown to possess haze-protective properties, while also positively impacting on the sensorial properties of the product. However, while such mannoproteins are released into the wine during the wine making process, the amounts are low and therefore of limited oenological significance. However, and although commercial wine yeast strains display significant genotypic and phenotypic diversity, no broader assessment of haze protective activity and of mannoproteins release by different wine yeast strains has been undertaken. In this study, several yeast strains were screened for their impact on wine haze formation in Chardonnay must and in a grape juice model system. The data show that strains of the species Saccharomyces paradoxus possess better haze protective properties than the common Saccharomyces cerevisiae wine yeast strains. Differences in the nature of the proteins released by these two species were investigated, and indicated that several mannoproteins were released at significantly higher levels by S. paradoxus, and that some of these proteins might indeed contribute to the haze-protective activity. A further exploration of yeast cell wall properties indicated that the cell walls of haze-protective S. paradoxus strains contained higher levels of chitin than non-haze protective strains. Grape chitinases are likely to be primarily responsible for wine haze formation, and the data clearly demonstrate that these enzymes are able to bind to the yeast cell walls, and that strains with higher amounts of chitin in the cell wall will bind more chitinases. This finding suggests that the haze-protective nature of the strains is at least in part linked to the chitin levels of the strains. Furthermore, the impact of some genetic modifications in two wine strains (namely S. cerevisiae VIN13 and S. paradoxus RO88) suggests that several proteins contribute to wine haze protection. However, none of the mannoprotein-encoding flocculation genes, FLO1, FLO5, and FLO11 showed any impact on this property. Further studies are required to assess the full impact of the S. paradoxus strains on haze protection. In particular, the possible use of such strains as starter cultures or the use of S. paradoxus yeast hulls as clarifying agent needs to be further explored.
AFRIKAANSE OPSOMMING: Wyn proteïen-waas vorming is 'n groot uitdaging vir wynmakers en verskeie wyn verhelderings agente soos bentoniet word in die wynbedryf gebruik om wyn te beskerm teen die vorming van waas. Hierdie verheldering agente het egter 'n ongewenste impak op wynkwaliteit. Gis mannoproteïene is uitgewys as proteïene met moontlike waas-beskermende eienskappe wat ook 'n positiewe uitwerking op die sensoriese eienskappe van die produk het. Al word hierdie mannoproteïene egter vrygestel in die wyn tydens die wynmaak proses, is die hoeveelhede oor die algemeen te laag om van wynkundige belang te wees. Verder, ten spyte van die beduidende genotipiese en fenotipiese diversiteit van kommersiële wyngisrasse is daar nog geen breër assessering van die waas beskermende aktiwiteit van mannoproteïene, vrygestel deur verskillende rasse, tot dusver onderneem nie. In hierdie studie is verskeie gisrasse gekeur vir hul impak op wyn waas-vorming in Chardonnay mos en ook in 'n model druiwesap. Die data wys dat rasse van die spesie Saccharomyces paradoxus besit beter waas beskermende eienskappe as die algemene Saccharomyces cerevisiae wyngisrasse. Verskille in die aard van die proteïene wat vrygestel is deur hierdie twee spesies is ondersoek, en dit is aangedui aangedui dat verskeie mannoproteins vrygestel aan aansienlik hoër vlakke deur S. Paradoxus. Dit is ook aangedui dat sommige van hierdie proteïene wel bydra tot die waas-beskermende aktiwiteit. 'n Verdere verkenning van gis selwand eienskappe het aangedui dat die selwande van waas-beskermende rasse van S. paradoxus hoër vlakke chitien as nie-waas beskermende stamme bevat. Druiwe chitinases is waarskynlik hoofsaaklik verantwoordelik vir wyn waas vorming, en die data toon duidelik dat hierdie ensieme in staat is om te bind aan die gis selwande, en dat die stamme met hoër vlakke chitien in die selwand meer chitinases sal bind. Hierdie bevinding dui daarop dat die waas-beskermende aard van die stamme ten minste gedeeltelik gekoppel is aan die chitien vlakke van die stamme. Die impak van sekere genetiese modifikasies in twee verskillende gisrasse, naamlik die S. cerevisiae ras VIN13 en die S. paradoxus ras RO88, dui verder daarop dat verskeie proteïene dra by tot die beskerming teen wyn waas. Geeneen van die mannoprotein-koderende flokkulasie gene, FLO1, FLO5 en FLO11 het egter 'n impak op hierdie eienskap nie. Verdere studies is nodig om die volle impak van die S. paradoxus rasse op waas beskerming te assesseer. In die besonder, die moontlike gebruik van sulke rasse as 'n inkolasie kultuur of die gebruik van S. paradoxus gis doppe as verheldering agent moet verder ondersoek word.
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Gorgens, Johann Ferdinand. "Quantitative yeast physiology and nitrogen metabolism during heterologous protein production." Thesis, Stellenbosch : University of Stellenbosch, 2003. http://hdl.handle.net/10019.1/16051.
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Dissertation (PhD)--University of Stellenbosch, 2003.ENGLISH ABSTRACT: QUANTITATIVE YEAST PHYSIOLOGY AND NITROGEN METABOLISM DURING HETEROLOGOUS PROTEIN PRODUCTION By Johann F. Görgens The physiology and nitrogen metabolism of the yeast, Saccharomyces cerevisiae, during heterologous xylanase production in a defined medium was quantified by the comparison of isogenic yeast strains, whereby several potential limitations in the production of the heterologous xylanase could be identified. The presence of global sensing and regulatory mechanisms, by which the yeast is able to actively regulate both heterologous gene expression and the physiological response to the process, was also investigated. The deleterious effects of heterologous xylanase production on the physiology of the recombinant host were disproportionately large with respect to the amount of foreign protein produced. The cellular processes involved in this response were identified by the transcriptional profiling of isogenic recombinant strains, in a novel analytical approach to investigating foreign protein production by S. cerevisiae. Heterologous gene expression affected a combination of cellular processes and induced the yeast stringent stress response. The corresponding loss of metabolic functionality resulted in the disproportionate physiological effects of foreign protein production, similar to previous observations in recombinant Escherichia coli, and a possible reduction in attainable production levels. Reducing the propensity of recombinant gene expression to introduce metabolic stress may therefore increase production levels of foreign proteins by yeast. The metabolic vitality of transformed strains was also reduced by the presence of multiple copies of active, plasmid-based PGK1-promoters in the cell without expression of the heterologous gene. The negative effect was caused by an increase in the biosynthetic and glycolytic capacity of the strain at the expense of other processes. Production levels of heterologous xylanase were influenced by expression vector selection and the presence of auxotrophic mutations in transformed strains of S. cerevisiae. The increased transcription levels obtained with the multicopy plasmidbased YEp-type expression system, compared to the integrative YIp-type expression system, resulted in higher levels of xylanase production. Heterologous xylanase production thus did not saturate the secretory capacity of the host strain. The genetic stability of the autoselective YEp-type expression system in long-term chemostat culture was also demonstrated. High levels of heterologous xylanase production by transformed S. cerevisiae strains containing auxotrophic markers required the stabilisation of nitrogen metabolism via saturation of yeast cells with an excess of imported amino acids. By the removal of excessive auxotrophic markers, high levels of xylanase production by a prototrophic transformant in defined medium without amino acid addition could be obtained. Heterologous xylanase production by the prototrophic transformant was further enhanced by increasing the availability of preferred amino acids or succinate in the defined medium, indicating an additional requirement for metabolic precursors and building blocks for foreign protein synthesis. Comparable levels of heterologous xylanase production were obtained in high cell density cultures of the alternative yeast, Pichia stipitis, by the proper induction of the native ADH2-promoter, the control of oxygenation, and addition of an amino acid mixture to the defined medium, indicating the presence of generic limitations in transcription, nutrient availability and the yeast biosynthetic capacity for foreign protein production by various yeasts. The presence of global sensing and regulatory mechanisms was confirmed by the physiological response of S. cerevisiae to heterologous protein production, which included the downregulation of biosynthesis and growth, and the induction of various processes involved in the stringent stress response. Additionally, heterologous xylanase production was actively regulated on a posttranscriptional level by the auxotrophic transformants in response to the level of amino acid availability. The biosynthetic capacity for foreign protein production by both recombinant S. cerevisiae and P. stiptis was also regulated in response to the physiological state of the yeast and the availability of nutrients. The presence of these regulatory mechanisms complicated the manipulation of cellular biosynthesis at will.
AFRIKAANSE OPSOMMING: KWANTITATIEWE GIS-FISIOLOGIE EN -STIKSTOF METABOLISME GEDURENDE HETEROLOË PROTEÏEN PRODUKSIE Deur Johann Ferdinand Görgens Die fisiologie en stikstof-metabolisme van die gis, Saccharomyces cerevisiae, gedurende heteroloë xilanase produksie in ‘n gedefiniëerde medium is gekarakteriseer deur isogeniese gis-rasse te vergelyk, waardeur verskeie moontlike beperkings in die produksie van die heteroloë xilanase uitgewys kon word. Die teenwoordigheid van globale sensoriese- en beheer-meganismes, wat die gis in staat stel om beide heteroloë geen uitdrukking en die fisiologiese respons op die proses aktief te reguleer, is ook ondersoek. Die nadelige effekte van heteroloë xilanase produksie op die fisiologie van die rekombinante gasheer-organisme was uitermatig groot in vergelyking met die hoeveelheid vreemde proteïen wat geproduseer is. Die sellulêre prosesse verantwoordelik vir hierdie respons is identifiseer deur die transkripsionele profiele van isogeniese rekombinante rasse te vergelyk, in ‘n nuwe analitiese benadering tot die bestudering van vreemde proteïen produksie deur S. cerevisiae. Heteroloë geen uitdrukking het ‘n kombinasie van sellulêre prosesse geaffekteer en die gis se algemene voedingstres-respons geaktiveer. Die gepaardgaande verlies aan metaboliese funksie het die uitermatige fisiologiese effek van vreemde proteïen produksie veroorsaak, soortgelyk aan vorige waarnemings met rekombinante Escherichia coli. Die haalbare produksie-vlakke is moontlik ook verlaag deur hierdie respons. ‘n Verlaging van die geneigdheid van rekombinante geen uitdrukking om metaboliese stres te veroorsaak, mag dus die produksievlakke van vreemde proteïene in gis verbeter. Die metaboliese groei-potensiaal van die getransformeerde rasse is ook verlaag deur die teenwoordigheid van etlike aktiewe kopieë van plasmied-gebaseerde PGK1-promotors in die sel, sonder uitdrukking van die heteroloë geen, deur ‘n toename in die biosintetiese en glikolitiese kapasiteit ten koste van die ander sellulêre prosesse. Die produksievlakke van heteroloë xilanase is deur die keuse van uitdrukkings-sisteem en die teenwoordigheid van autotrofiese mutasies in die getransformeerde rasse van S.cerevisiae beïnvloed. Die verhoogde transkripsie vlakke wat met die multi-kopie, plasmied-gebaseerde YEp-tipe uitdrukkingsisteem, eerder as die geïntegreerde YIp-tipe sisteem, verkry is, het tot verhoogde xilanase produksie gelei. Heteroloë xilanase produksie het dus nie die uitskeidingskapasiteit van die gasheer versadig nie. Die genetiese stabiliteit van die autoselektiewe, YEp-tipe uitdrukkingsisteem in langtermyn chemostaat-kulture is ook gedemonstreer. Hoë vlakke van xilanase produksie deur getransformeerde S. cerevisiae rasse met autotrofiese merkers het die stabilisering van die stikstof metabolisme, deur die versadiging van die sel met ingevoerde aminosure, vereis. Die verwydering van oormatige autotrofiese merkers het tot hoë vlakke van xilanase produksie deur die prototrofiese transformant in gedefinieerde medium sonder aminosuur byvoeging gelei. Heteroloë xilanase produksie deur die prototrofiese transformant kon verder verbeter word deur die byvoeging van voorkeur-aminosure of suksinaat tot die gedefinieerde medium, en ‘n addisionele behoefte aan metaboliese voorloper-molekules en bou-blokke vir vreemde proteïensintese het dus bestaan. Vergelykbare vlakke van heteroloë xilanase produksie is in kulture met hoë sel-digthede van die alternatiewe gis, Pichia stipitis, verkry deur die doeltreffende induksie van die eiesoortige ADH2-promotor en die byvoeging van ‘n aminosuur-mengsel tot die gedefinieerde medium, wat die teenwoordigheid van generiese beperkinge in transkripsie, voedingstof-beskikbaarheid en biosintetiese kapasiteit van die gis vir vreemde proteïen produksie deur verskeie giste uitgewys het. Die teenwoordigheid van globale sensoriese- en beheer-meganismes is bevestig deur die fisiologiese respons van S. cerevisiae tot heteroloë proteïen produksie, wat die afwaartse regulering van biosintese en groei, en die induksie van verskeie prosesse betrokke by die algemene voedingstres-respons, ingesluit het. Heteroloë xilanase produksie is ook op ‘n na-transkripsionele vlak aktief gereguleer deur die autotrofiese transformante in reaksie tot die vlak van aminosuur beskikbaarheid. Die biosintetiese kapasiteit vir vreemde proteïen-produksie van beide rekombinante S. cerevisiae en P. stipitis is ook in reaksie tot die fisiologiese toestand van die gis en die beskikbaarheid van voedingstowwe gereguleer. Die teenwoordigheid van hierdie regulatoriese meganismes het die willekeurige manipulasie van sellulêre proteïen-biosintese bemoeilik.
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Pandit, Shalmalee(Shalmalee Dhananjay). "Environmental remediation and biofuel production through nanoparticle stimulation of yeast." Thesis, Massachusetts Institute of Technology, 2019. https://hdl.handle.net/1721.1/128313.
Full textAbstract:
Thesis: S.M., Massachusetts Institute of Technology, Department of Biological Engineering, 2019Cataloged from PDF version of thesis.
Includes bibliographical references (pages 43-47).
Artificially photosynthetic systems aim to store solar energy and chemically reduce carbon dioxide. These systems have been developed in order to use light to drive processes for carbon fixation into biomass and/or liquid fuels. We have developed a hybrid-biological system that manages both genetically controlled generation of products along with the photoactivability of a semiconductor system. We show an increase in the production of ethanol, a common biofuel, through the electron transfer stimulated by biologically produced cadmium sulfide nanoparticles and light. This work provides a basis on which to improve the production of many metabolites and products through endogenously produced nanoparticles.
by Shalmalee Pandit.
S.M.
S.M. Massachusetts Institute of Technology, Department of Biological Engineering
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Dilworth, Marvin. "Novel strategies to improve recombinant membrane protein production in yeast." Thesis, Aston University, 2014. http://publications.aston.ac.uk/24556/.
Full textAbstract:
Approximately 60% of pharmaceuticals target membrane proteins; 30% of the human genome codes for membrane proteins yet they represent less than 1% of known unique crystal structures deposited in the Protein Data Bank (PDB), with 50% of structures derived from recombinant membrane proteins having been synthesized in yeasts. G protein-coupled receptors (GPCRs) are an important class of membrane proteins that are not naturally abundant in their native membranes. Unfortunately their recombinant synthesis often suffers from low yields; moreover, function may be lost during extraction and purification from cell membranes, impeding research aimed at structural and functional determination. We therefore devised two novel strategies to improve functional yields of recombinant membrane proteins in the yeast Saccharomyces cerevisiae. We used human adenosine A2A receptor (hA2AR) as a model GPRC since it is functionally and structurally well characterised. In the first strategy, we investigated whether it is possible to provide yeast cells with a selective advantage (SA) in producing the fusion protein hA2AR-Ura3p when grown in medium lacking uracil; Ura3p is a decarboxylase that catalyzes the sixth enzymatic step in the de novo biosynthesis of pyrimidines, generating uridine monophosphate. The first transformant (H1) selected using the SA strategy gave high total yields of hA2AR-Ura3p, but low functional yields as determined by radio-ligand binding, leading to the discovery that the majority of the hA2AR-Ura3p had been internalized to the vacuole. The yeast deletion strain spt3Δ is thought to have slower translation rates and improved folding capabilities compared to wild-type cells and was therefore utilised for the SA strategy to generate a second transformant, SU1, which gave higher functional yields than H1. Subsequently hA2AR-Ura3p from H1 was solubilised with n-dodecyl-β-D-maltoside and cholesteryl hemisuccinate, which yielded functional hA2AR-Ura3p at the highest yield of all approaches used. The second strategy involved using knowledge of translational processes to improve recombinant protein synthesis to increase functional yield. Modification of existing expression vectors with an internal ribosome entry site (IRES) inserted into the 5ˊ untranslated region (UTR) of the gene encoding hA2AR was employed to circumvent regulatory controls on recombinant synthesis in the yeast host cell. The mechanisms involved were investigated through the use of yeast deletion strains and drugs that cause translation inhibition, which is known to improve protein folding and yield. The data highlight the potential to use deletion strains to increase IRES-mediated expression of recombinant hA2AR. Overall, the data presented in this thesis provide mechanistic insights into two novel strategies that can increase functional membrane protein yields in the eukaryotic microbe, S. cerevisiae.
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Routledge, Sarah. "The effect of antifoams upon recombinant protein production in yeast." Thesis, Aston University, 2012. http://publications.aston.ac.uk/16713/.
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Foaming during fermentation reduces the efficiency of the process leading to increased costs and reduced productivity. Foaming can be overcome by the use of chemical antifoaming agents, however their influence upon the growth of organisms and protein yield is poorly understood. The objective of this work was to evaluate the effects of different antifoams on recombinant protein production. Antifoam A, Antifoam C, J673A, P2000 and SB2121 were tested at different concentrations for their effect on the growth characteristics of Pichia pastoris producing GFP, EPO and A2aR and the yield of protein in shake flasks over 48 h. All antifoams tested increased the total GFP in the shake flasks compared to controls, at higher concentrations than would normally be used for defoaming purposes. The highest yield was achieved by adding 1 % P2000 which nearly doubled the total yield followed by 1 % SB2121, 1 % J673A, 0.6 % Antifoam A and lastly 0.8 % Antifoam C. The antifoams had a detrimental effect upon the production of EPO and A2aR in shake flasks, suggesting that their effects may be protein specific. The mechanisms of action of the antifoams was investigated and suggested that although the volumetric mass oxygen transfer coefficient (kLa) was influenced by the agents, their effect upon the concentration of dissolved oxygen did not contribute to the changes in growth or recombinant protein yield. Findings in small scale also suggested that antifoams of different compositions such as silicone polymers and alcoxylated fatty acid esters may influence growth characteristics of host organisms and the ability of the cells to secrete recombinant protein, indirectly affecting the protein yield. Upon scale-up, the concentration effects of the antifoams upon GFP yield in bioreactors was reversed, with lower concentrations producing a higher yield. These data suggest that antifoam can affect cells in a multifactorial manner and highlights the importance of screening for optimum antifoam types and concentrations for each bioprocesses.
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Üzelyalçın, Berna Harsa Şebnem. "Effects Of Trace Elements On The Production Of Baker's Yeast/." [s.l.]: [s.n.], 2002. http://library.iyte.edu.tr/tezler/master/biyoteknoloji/T000146.rar.
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Canonico, Laura. "Mixed fermentations, yeast interactions and metabolites production in industrial fermentations." Doctoral thesis, Università Politecnica delle Marche, 2015. http://hdl.handle.net/11566/243078.
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Saccharomyces cerevisiae è il lievito principalmente utilizzato nella produzione di bevande fermentate come vino e birra. È in questo contesto che si sono sviluppate colture starter selezionate di ceppi di S. cerevisiae da impiegare in questi settori.
Negli ultimi anni però la ricerca, soprattutto in campo enologico, si è concentrata sull’utilizzo di fermentazioni multistarter controllate di lieviti non-Saccharomyces e ceppi di S.cerevisiae, con l’obiettivo di ottenere prodotti caratterizzati da maggiore complessità e caratteristiche sensoriali peculiari. Per tale motivo, la presente ricerca si è incentrata sullo studio di fermentazioni miste e di interazioni tra diverse specie di lievito non-Saccharomyces e S. cerevisiae in due diverse bevande alcoliche: birra artigianale e vino.
Nella prima parte della tesi è stata effettuata una caratterizzazione molecolare di 28 ceppi di Torulaspora delbrueckii, isolati da matrici naturali provenienti da diverse aree geografiche (Italia e Camerun), utilizzando differenti metodi molecolari: RAPD-PCR (Random Amplification of Polymorphic DNA), la PCR-fingerprinting con i minisatellites SED1, AGA1, DAN4 e nuovi primers per T. delbrueckii (Td)PIR disegnati in questo laboratorio, e i microsatelliti (GAC)5 and (GTG)5. L’obiettivo del lavoro è quello di utilizzare una nuova coppia di primers (Td)PIR disegnati su sequenze geniche codificanti per proteine di parete (CWPs) per effettuare una tipizzazione di ceppi di T. delbrueckii. L’utilizzo della PCR fingerprinting con il primer TdPIR3 proposto in questo lavoro ha mostrato un’alta efficienza e un alto potere discriminatorio se confrontati con il metodo RAPD e i microsatelliti, il che li rende particolarmente utili per la tipizzazione molecolare di T. delbrueckii.
La seconda parte della ricerca è volta a migliorare la qualità della birra artigianale. In particolar modo si sono studiate la dominanza e l’influenza di ceppi selezionati di S. cerevisiae sul profilo analitico di birra artigianale dopo rifermentazione in bottiglia e l’impiego di ceppi di T. delbrueckii in fermentazioni miste per conferire un particolare “bioflavour” al prodotto finito. I risultati ottenuti hanno mostrato come l’impiego di ceppi selezionati di S. cerevisiae nel processo di rifermentazione e le fermentazioni miste (S. cerevisiae / non- Saccharomyces) abbiano effetto sul profilo aromatico della birra e possano quindi essere impiegati per produrre caratteristici bioflavours.
Nella terza parte sono stati valutati il comportamento fermentativo di diversi ceppi non-Saccharomyces appartenenti ai generi (Candida, Metschnikowia, Torulaspora, Starmerella), e ceppi di S. cerevisiae in condizioni aerobiche e anaerobiche e differenti modalità fermentative con lo scopo di ridurre il contenuto alcolico nei vini. Fermentazioni condotte con lieviti non-Saccharomyces in condizioni aerobiche ed anaerobiche hanno mostrato differenze nella cinetica fermentativa, produzione di biomassa ed etanolo e rendimento. Inoltre, sono state condotte fermentazioni sequenziali, utilizzando cellule di lievito non-Saccharomyces appartenenti al genere Starmerella, Hanseniaspora, Candida, Metschnikowia in forma immobilizzata e un ceppo starter di S. cerevisiae su mosto sintetico e naturale. I risultati hanno mostrato che l’obiettivo di ridurre il contenuto di etanolo di circa 2% è raggiungibile, e che differenti condizioni fermentative hanno influenza sul comportamento di differenti lieviti.
La quarta e ultima parte è stata rivolta allo studio delle fermentazioni multistarter vinarie, andando a studiare la cinetica di crescita, il profilo analitico e aromatico dei vini. Sono state quindi selezionate diverse coppie di ceppi di S.cerevisiae e non-Saccharomyces e si è studiata la loro interazione sia su mosto Verdicchio che su Montepulciano. I risultati ottenuti hanno mostrato prodotti con differenti caratteristiche analitiche e sensoriali da quelli ottenuti in fermentazioni pure con l’utilizzo di ceppi di S. cereviaiae del commercio.The Saccharomyces cerevisiae yeast is widely used in the production of fermented beverages such as wine and beer. In this context, selected cultures of S. cerevisiae have developed to be used in these fermentation industries. However, in recent years, the research in winemaking was focused on use of controlled multistarter fermentations by selected cultures of non-Saccharomyces and S. cerevisiae strains, with the aim to obtain products characterized by a more complexity and peculiar sensory characteristics. In this context the present research focused on mixed fermentation and yeast interactions of different non-Saccharomyces yeasts species and S. cerevisiae strains in two different alcoholic beverages: craft beer and wine. In the first part of the thesis has been carried out a molecular characterization of 28 Torulaspora delbrueckii strains, isolated from various natural matrices coming from different environments (Italy and Cameroon), using different PCR-fingerprinting molecular methods: random amplified polymorphic DNA with polymerase chain reaction (RAPD-PCR), minisatellites SED1, AGA1, DAN4 and the newly designed T. delbrueckii (Td)PIR, and microsatellites (GAC)5 and (GTG)5. In this work we found new specific primers pairs (Td)PIR, designed on gene sequence of PIR coding for cell-wall proteins (CWPs), for T. delbrueckii typing. TdPIR3 minisatellite fingerprinting approach proposed in this work, showed high efficacy and discrimination power when compared with RAPD and microsatellites methods, making it particularly useful to T. delbrueckii molecular typing. The second part of the research was focused on the improvement of quality of craft beer. In particular, we have investigated on the dominance and influence of selected S. cerevisiae strains on the analytical profile of craft beer after bottle refermentation and the use of T. delbrueckii selected strains in mixed fermentations to give a peculiar "bioflavour" to the final product. The results showed that selected S. cerevisiae in the refermention process and mixed fermentation (S. cerevisiae / non-Saccharomyces) affected the aromatic composition of beer and could be used to generate distinctive bioflavour. In the third part, we evaluated the behavior of different non-Saccharomyces genera (Candida, Metschnikowia, Torulaspora, Starmerella), and several strains of S. cerevisiae in aerobic and anaerobic condition and different fermentation modalities with the aim to reduce the alcohol content in wine. Non-Saccharomyces yeasts fermentation trials carried out in aerobic and anaerobic condition showed differences in fermentation kinetic, biomass and ethanol production and ethanol yield. Moreover, sequential fermentations using immobilized cells of some non-Saccharomyces yeast strains belonging to Starmerella, Hanseniaspora, Candida, Metschnikowia genera and S. cerevisiae starter strain on synthetic and natural grape juice were carried out. Results showed that the aim to reduce the ethanol concentration of about 2% is attainable, and the different fermentation conditions have effected on the behavior of different yeasts. In the fourth and final part of the study we focused on multistarter fermentations in wine, with the aim to study the kinetics growth, the analytical and aromatic profile of wines. Then were selected different pairs of S. cerevisiae and non-Saccharomyces strains and we studied their interactions on Verdicchio and Montepulciano grape juice. The results showed wines with different analytical and sensorial profiles in comparison with wines obtained with pure fermentation using S. cerevisae commercial starter strains.
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Smerdon, Gary Randall. "Production of human gastric lipase in the fission yeast Schizosaccharomyces pombe." Thesis, University of Exeter, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.261145.
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McKay, Ian Donald. "Aspects of growth and lipid production in the oleaginous yeast Trichosporon cutaneum." Thesis, University of Reading, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.329311.
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Skogsberg, Zara. "Investigation of the Impact on Yeast Fermentation Performance in Production of Pale Lager Beer through Management Control." Thesis, Linköpings universitet, Teknisk biologi, 2013. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-100150.
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Through a full factorial design experiment, the effects of time between worts, wort aeration and yeast dosage in production of a pale lager beer were examined in the beer process at Spendrups Bryggeri AB. The aim was to learn how different parameters may affect the yeast fermentation performance during beer production. Response variables used were the concentrations of ethyl acetate and isoamyl acetate, free amino nitrogen (FAN) degradation and change in extract. A statistical analysis showed that the concentration of ethyl acetate is dependent on yeast dosage and the interaction between time between worts and aeration while the isoamyl acetate concentration is dependent on yeast dosage and time between worts. No parameters are statistically significant for FAN degradation while the change in extract is dependent on the yeast dosage. Due to botched runs, mostly because of aeration problems, it was not possible to verify theoretical parameter values and responses. Since the aeration was not properly performed, the management of the aeration control should be further investigated. Ester analysis and analysis of FAN were performed as worts entered and exited horizontal fermentation tanks. An additional analysis of ester content was also performed as the early stage beer was transferred into lagering tanks. Cell viability as well as extract, pH and tank temperature was measured daily to verify the state of fermentation. Statistical calculations showed that when using NucleoCounter YC-100, there is no significant difference between analysis made of samples homogenized by a magnetic stirrer and samples shaken by hand.
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Harrison, Joanna Shan. "Production of recombinant antibody fragments in microorgansms." Thesis, University College London (University of London), 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.244739.
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PASSOLUNGHI, SIMONE. "Matching biotech needs and yeast physiology." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2009. http://hdl.handle.net/10281/8206.
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The research in the field of industrial biotechnology, especially regards bioproducts and bioprocesses, are aimed at developing innovative technologies that lead to obtaining compounds with the use of microorganisms, or seeking to enhance existing processes to increase yield, production and productivity, trying to ensure a higher degree of sustainability and reducing environmental impact.
To pursue these goals is possible to intervene by adopting a “technologic” approach that includes the development of systems capable of ensuring a more effective control of the parameters that govern the production processes or, with a “molecular / metabolic” approach, acting directly on the host system, namely by working on production capacity of the microorganism itself. To achieve this goal the pathways responsible for the processes of synthesis or secretion of products of interest must be identified and, if necessary, modified, or the environmental conditions in which the organism is operating during the process must be considered, to study how to improve its production capacity even in non-optimal physiological condition.
The process conditions that characterize industrial production processes often put the cells through a series of stress that inevitably act negatively on yields. It therefore becomes necessary to identify the limiting factors in relation to the host organism and, on this basis, to act in an appropriate manner. To do this it is possible to adopt different strategies, complementary and not mutually exclusive.
The most immediate is the exploration of biodiversity in order to choose a host that is intrinsically and naturally more resistant to the type of stress imposed by the process. This way is not always easy to follow, given the extent of the possible solutions and the lack of resources that allow the exploration and characterization in a reasonable time for the development of a biotechnological process. An alternative strategy is focused on the characterization of the cellular response to these stress conditions to identify the key factors involved in the mechanism. In this way, for example, through genetic manipulation on these factors it could be possible to improve the resistance of the cell itself, or by the transfer of these specific genetic traits to improve the resistance of other micro-organisms selected as host system.
The yeast Saccharomyces cerevisiae is one of the most widely used microorganisms for the production of compounds of biotechnological interest because of the available knowledge on the physiology, genetics, biochemistry, and on the existence of technological and molecular tools suitable for its manipulation in order to optimize the production by fermentation. It is important to underline that S. cerevisiae is recognized as a GRAS organism (generally regarded as safe) by the Food & Drug Administration that has allowed the use for the production of pharmaceutical compounds for human use. While it is now increasingly clear the potential of S. cerevisiae as a platform for metabolic engineering, for some heterologous proteins production on a large scale this yeast is not the ideal host system. Very often the expressed proteins are hyper-glycosilated or, if retained in the periplasmic space, they suffer significant degradation. For other industrial productions, especially where the product of interest has to be cheap, the fermentation technologies needed are too complex and sophisticated (and expensive) to be implemented on a large scale. With these assumptions in recent years it has been explored the opportunity of adopting other yeasts, called "unconventional yeast" by developing new systems of expression. In this thesis project an "alternative" yeast, Zygosaccharomyces bailii, is considered. Although it is not well characterized from molecular and genetic point of view, it presents interesting features in view of potential biotechnological production: it allows high yields of biomass, it has a high specific growth rate and a higher resistance compared to S. cerevisiae, to certain types of stress, and in particular to stresses generated by an acid medium.
The cell surface, as an area of communication and exchange between the extracellular environment and the cell itself is one of the targets of this study.
In S. cerevisiae, thanks to the availability of molecular tools and the knowledge of the entire genome sequence, it is possible the design of in-depth studies and the engineering of the cellular metabolic network. Given the difficulties in its genetic manipulation, in the case of Z. bailii was first necessary to address another preliminary issue. In this diploid yeast the deletion of an essential gene was never been made. It has been developed a reproducible protocol for gene deletion by a gene-targeting approach and it was obtained the deletion of the gene ZbLEU2, who gave birth to the first auxotroph strain of Z. bailii (leuˉ). This represents an important step for a possible use of Z. bailii as host system. Thanks to this protocol a mutant strain of Z. bailii of potential interest for heterologous protein production was also obtained, in analogy with what reported in the literature for S. cerevisiae. The deletion concerns the homologue of ScGAS1, coding for the enzyme β-1 ,3-glucanosyltransferase which catalyzes the crosslinking of cell wall glucans. ZbGAS1. The gene was cloned by PCR and sequenced. The deleted mutant of Z. bailii has morphological and phenotypic characteristics very similar to the correspondent in S. cerevisiae, showing an alteration of the cell wall structure, and enhanced secretive capacity than the wild type strain for some heterologous proteins that have been considered.
In parallel to these studies, populations of Z. bailii growing on different carbon sources were analyzed by flow cytometry. The analysis of DNA and protein content was performed to better characterize this yeast not only from molecular point of view, but also to explore its cellular features.
The characterization of this unconventional yeast confirmed once again one of the most appreciated features for yeasts used as cell factories: versatility.
This property is so strong that yeasts has been exploited for natural abilities, such as production of ethanol, and also for processes where a targeted manipulation was introduced, for example in lactic acid production, just to cite a pair of biotechnological production of industrial relevance. To make this process competitive on the market, in terms of yield, production and productivity, yeasts were pushed to their physiological limits. These limits are given by the accumulation of vast amounts of product that, in the case of ethanol, can cause damage to the lipid component of the plasma membrane, in the case of lactic acid can result in a loss of proper cellular homeostasis with the fall of the intracellular pH.
It is therefore necessary to assess whether these limitations can be overcome by acting in particular on the plasma membrane, whose role in controlling the transport and the cellular homeostasis makes it a target of interest to improve the robustness of cells in response to stress generated by the process, such as oxidative stress or generated by low pH.
The optimization of transport through the membrane plays a key role in the mechanisms of adaptation to these stresses. In particular, improving the flux of nutrients entering the cell could allow an optimal uptake of nutrient in the cytoplasm (as in the case of bioethanol production), an improvement of outflows from the cells might instead allow effective removal of compounds that may be deleterious for cell viability when present beyond a threshold value (eg organic acids).
By focusing on the protein fraction that characterizes the cytoplasmic membrane, we have studied the effects of modulation of the expression of H +-ATPase pump of the plasma membrane (Pma1p), involved in the intracellular pH homeostasis. In particular, the gene ScPMA1 was overexpressed in S. cerevisiae and this overexpression was able to confer a greater resistance to acid stress, evidenced by growth kinetics in the presence of lactic acid. The increased cell viability under restrictive conditions in respect to the wild type strain was also checked by flow cytometry. The use of this tool has enabled the development of a system able to assess quantitatively the degree of robustness of the cells in stressful conditions.
With this instrument able to assess the robustness of the cells as a function of various types of stress (oxidative, pH, ...) it is possible to design new interventions of metabolic engineering in order to provide greater resistance to yeast in restrictive process conditions, similar to conditions prevailing in the processes of production of compounds of biotech interest. It will be possible to evaluate the effectiveness of these interventions with the flow cytometer by assessing the response of the engineered cells under restrictive conditions by measuring the ability of cells to increase their robustness. The robustness remains one of the key features of yeasts as microbial cell factories, in particular to reach one of the main goal of White Biotechnology: to provide value products from renewable resources through sustainable processes with low environmental impact.
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D'Anjou, Marc Charles. "Production of recombinant antifreeze proteins in the methylotrophic yeast Pichia pastoris." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape3/PQDD_0021/MQ54447.pdf.
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Ferhane, Akila. "A novel method for the production of a selenium-enriched yeast /." Thesis, McGill University, 2001. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=37890.
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Selenium (Se) is an essential element. Supplementation of Se as yeast-Se in animal and human diets has been proven to have beneficial health effects. The goal of this study was to add a maximum amount of Se in yeast metabolism in order to optimize its incorporation in amino acids. A bakery yeast strain of Saccharomyces cerevisiae (S. cerevisiae) was studied for its tolerance to Se when the latter was incorporated at different levels. Fermentations were run at 27°C for 8 h and 24 h. The maximum Se incorporation was achieved when 12.6 mmol of Se, as sodium selenite salt, was added to the culture medium and fermented for 24 h. A final Se concentration of 1550 +/- 35 mug/g yeast was obtained by this treatment.Different yeast strains of S. cerevisiae were also studied for their capacity to incorporate Se. Five yeast strains of wine and four yeast strains of beer were fermented for 24 h and tested for their capacity to incorporate Se. The amount of 12.6 mmol Se was added in the growth medium. A maximum of 642.6 +/- 3.6 mug Se/g yeast was found to be incorporated in Uvaferm BC wine strain. Uvaferm windsor of beer strain was able to incorporate a maximum of 826.8 +/- 10.4 mug Se/g yeast. These yeast strains could be used as alternatives for Se supplementation.
Se speciation was carried out on the bakery yeast strain containing 1550 +/- 35 mug Se/g yeast, using Fast Phase Liquid Chromatography (FPLC) and amino acid analysis. Out of 1550 +/- 35 mug Se/g yeast, 57.5% Se was present as selenoaminoacids. The yeast extract contained 147 +/- 14 mug/g of SeCys, 248 +/- 13 mug/g of SeCyst and 295 +/- 17 mug/g of SeMet. Yeast cell walls contained 65 +/- 8 mug/g of SeMet; 69 +/- 5 mug/g of SeCyst and 67 +/- 9 mug/g of SeCys. These selenoaminoacids are known for their beneficial health effects. The produced Se-enriched bakery yeast could be used, after evaluated to be toxicologically safe, as an efficient dietary supplement.
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Eder, Matthias. "Deciphering the genetic and metabolic basis of yeast aroma properties." Thesis, Montpellier, SupAgro, 2017. http://www.theses.fr/2017NSAM0054.
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La levure Saccharomyces cerevisiae joue un rôle essentiel dans la production de composés aromatiques, tels que les esters, les alcools supérieurs et les acides organiques, ainsi que dans la transformation de précurseurs d'arômes du raisin pendant la fermentation du vin. Afin d'identifier les bases génomiques et métaboliques de ces propriétés, un croisement a été réalisé entre deux souches de levures de vin, sélectionnées pour leurs besoins en azote différents lors de la fermentation. 130 ségrégants de génération F2 ont été génotypés par séquençage complet du génome et individuellement phénotypés pendant la fermentation en mesurant les métabolites extracellulaires par HPLC et GC-MS. Les flux métaboliques intracellulaires ont été estimés à l’aide d’un modèle à base de contraintes. Une analyse QTL (quantitative trait locus) a été utilisée pour identifier les allèles influençant les variations d'arômes et de flux métaboliques. Plus de 80 QTL expliquant la variation de 59 caractères quantitatifs ont été détectés. Ces caractères comprennent des paramètres fermentaires, de consommation de substrat, la production de principaux métabolites et d’arômes fermentaires, ainsi que le métabolisme de composés aromatiques du raisin. L’intérêt de la cartographie QTL pour identifier les déterminants génétiques de variations de flux intracellulaires (f-QTLs) a par ailleurs été démontrée. Les QTL détectés ont été disséqués et des gènes dont les allèles contribuent spécifiquement aux variations phénotypiques ont été identifiés. Ces résultats soulignent la complexité génomique et métabolique de la synthèse et de la transformation d'arômes par la levure. L'identification de ces déterminants génétiques permet de mieux comprendre les liens entre variation génétique des levures et traits technologiques et fournit une base précieuse pour le développement de souches optimisées par des stratégies génétiques de croisement assisté par marqueursThe yeast Saccharomyces cerevisiae plays a vital role in the production of aroma compounds, such as esters, higher alcohols and organic acids, and the conversion of grape-derived aroma precursors during wine fermentation. To identify the genomic and metabolic bases for these processes, a cross was performed between two wine yeast strains selected because of their different nitrogen requirement during fermentation. 130 F2-segregants were genotyped by whole genome sequencing and individually phenotyped during wine fermentation by measuring extracellular metabolites using HPLC and GC-MS. Intracellular metabolic fluxes were estimated by constraint-based modeling. Quantitative trait locus (QTL) mapping was used to identify allelic variants influencing variations in the aroma profile and metabolic fluxes. More than 80 QTLs explaining variation in 59 quantitative traits were detected. These traits consisted of general fermentation parameters, substrate consumption, the production of main metabolites and fermentative aromas and the metabolism of grape aroma compounds. The applicability of QTL mapping to detect regions influencing intracellular fluxes (f-QTLs) was furthermore demonstrated. Found QTLs were dissected and genes with allele specific contributions to the phenotype were identified. These results emphasize the genomic and metabolic complexity of yeast aroma formation. In addition, the identification of genetic determinants increases knowledge about the links between genetic variation and industrial traits and provides a valuable foundation for the development of optimized strains by marker-assisted selection breeding strategies
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Mir, Nadeem Hussain. "The synthesis of β-1,3-glucanase in the native Oerskovia and in recombinant Bacillus subtilis : characterisation of system stability and comparison to native expression." Thesis, University of Reading, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308054.
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Godfrey, Valerie. "Production of Biodiesel from Oleaginous Organisms Using Underutilized Wastewaters." DigitalCommons@USU, 2012. https://digitalcommons.usu.edu/etd/1337.
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Driven by the rising costs, decreasing convenience, and increased demand of fossil fuels, the need for alternative, sustainable energy sources has caused a spark in interest in biomass-based fuels. Oleaginous organisms such as yeast, algae, and bacteria have been considered as microscopic biofactories for oils that can be converted into biodiesel. The process of growing such organisms using current technology requires an alarming amount of freshwater, which is another resource of growing concern. The research detailed within explains how several sources of underutilized wastewater can serve as growth medium in the biodiesel production process. Using only nitrogen and in one case phosphorus as external supplements, algae were shown to grow on produced water from oil and gas industry waste, local municipal wastewater, environmental brackish water from the Great Salt Lake, and wastewater from the potato processing industry. In each case, growth and biodiesel production in wastewaters was as good as or better than laboratory media. The bacterial organism Rhodococcus opacus PD630 and the yeast organism Cryptococcus curvatus were also used to grow on the dairy manufacturing wastewater whey permeate, a large source of underutilized fixed carbon, with successful lipid production. C. curvatus was also used to successfully grow and form large amounts of biodiesel from ice cream factory wastewater and from wheat straw hydrolysate. In each case, the need for freshwater and outside nutrients was nearly entirely replaced, with the exception of some nitrogen supplementation, with a wastewater nutrient source, thus adding to the sustainability of biomass-based fuels.
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Lorencez, Gonzalez Isaac Lorencez Gonzales Isaac. "Aerobic ethanol production with a flocculent yeast in a biomass recycling system /." [S.l.] : [s.n.], 1985. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=7702.
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Mokiou, Stella. "Ecophysiological approaches to production and formulation of the biocontrol yeast Pichia anomala." Thesis, Cranfield University, 2004. http://hdl.handle.net/1826/932.
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To produce commercial biocontrol agents (BCAs) successfully, it is important that cheap and economic substrates are used which support high numbers of good quality inoculum. Production of formulations conserving ecological competence and shelf-life should also be ensured. With this in mind, studies focusing on yeast ecophysiology were conducted to produce and formulate ecologically competent P. anomala cells for controlling spoilage of moist cereal grain. The liquid culture systems used were synthetic, nutrient yeast dextrose broth (NYDB), and a complex (industry byproduct, cane molasses) media. Manipulation of cultural conditions by means of imposing water-stress with several solute additions to the media had an impact on yield, cell water potentials (Ψc), viability and endogenous sugar/polyol accumulation. Glucose addition resulted in higher yeast yield (6.15 and 3.4 mg cell ml-1 medium for NYDB and molasses, respectively). Water activity (aw) modifications of the media resulted in modification of Ψc so that Ψc ≤ Ψw (medium water potential). The change in yeast Ψc was attributed to the intracellular accumulation/synthesis of polyols, mostly glycerol and arabitol and sugars, mostly trehalose. In molasses-based medium cells accumulated/synthesized trehalose [32 mg g-1 fresh weight (f.w.) yeast cell]. Higher amounts of endogenous trehalose (up to 140 mg g-1 f.w. yeast cell) were retained intracellularly when modified yeast cells were isotonically washed compared to those subjected to hypo-osmotic shock by washing with water. The pattern was similar for endogenous arabitol. Trehalose retention doubled and quadrupled, and arabitol increased by 65 and 100% in proline and NaCl treatments respectively. The molasses control medium gave high [>1010 colony forming units (CFU) ml-1] cell viability, which was further increased by addition of NaCl and proline (≈ 3x1010CFU ml-1).
Fluidised bed drying of yeast cells showed that drying at 50oC for 20 min resulted in high cell viability (67%) and low moisture content (7%). Osmoprotection and several carriers and adjuvants affected viability and moisture content. Cotton seed flour (CSF) + 10% skimmed milk (SM) resulted in the highest cell protection (74%) during the drying process, with a final moisture content of about 5% and this was easy to resuspend. Storage stability of the formulation was 50% at 4oC and ambienta temperatures for up to 150 days. P. anomala cells grown in NaCl
ii
modified molasses-media, when osmoprotected, retained four times more trehalose and resulted in significantly increased survival after drying and storage stability for 150 days. When SM + sucrose at 10% (w/v) was used as a protective solution, P. anomala cells were highly resistant to freezing, thawing and freeze drying processes. Storage stability at 4oC of freeze dried P. anomala cells was particularly high (>86%) over a period of 150 days while storage at 22oC resulted in a rapid decrease in cell viability to <35% over a period of 30 days. Osmoprotection using post-harvest isotonic washing treatment had no effect on storage stability. P. anomala produced high β-(1-3)-glucanase (>2.2 activity units), low chitinase (<0.9 activity units) and β-glucosidase (<3 nmol 4-nitrophenol min µg protein of specific enzyme activity)-1-1 amounts. The role of the first hydrolase in biocontrol activity is possibly important while that of the other two is not clear for P. anomala. In lab-scale bioassays using wheat grain under aerobic conditions, populations of P. verrucosum 22625 were significantly reduced by formulated P. anomala cells at both 0.93 and 0.95 aw levels while OTA production was significantly reduced at 0.93 aw only.
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Chantasuban, Tanakorn. "Assessing the wine yeast Metschnikowia pulcherrima for the production of 2-phenylethanol." Thesis, University of Bath, 2017. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.715273.
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2-phenylethanol (2PE) is a valuable fragrance compound which gives a rose-like aroma. As such 2PE is one the highest used fragrances globally. While 2PE is predominantly produced from petrochemical resources, there is a growing market for a naturally derived alternative for food products. 2PE from natural sources is priced so highly due to limited supply from rose petals. Recently, a few reports have demonstrated the production in yeasts through both the de novo production from glucose and ex novo biosynthesis with L-phenylalanine as a precursor. While these are promising most of the yeasts used can only produce low titres under optimal conditions, and the fermentation still appears to be too expensive. In this investigation the wine yeast M. pulcherrima was selected to be assessed for 2PE production. M. pulcherrima is known to produce 2PE in small titres in wine production though has yet to be explored as a platform for this product. M. pulcherrima has several advantages as a yeast platform, in that it produces a range of antimicrobials which can ward off invasive species, allowing for less sterile control in any large scale fermentation. M. pulcherrima was demonstrated to be able to produce 2PE in high titres in the batch mode through de novo synthesis of glucose, producing up to 1 g/L in shake flasks on the lab scale. Arabitol was also observed in the fermentation broth and was produced up to 20 g/L. The fermentation was then scaled up to 2L in batch mode. From these experiments, up to 700 mg/L of 2PE was produced. This is substantially more than any other yeast in the literature to date. Though when xylose or glycerol was present then 2PE production was severely limited. M. pulcherrima was also demonstrated to be able to produce 2PE by bioconversion from phenylalanine up to 1.5 g/L. This 2PE concentration is suggested to be threshold of toxicity to M. pulcherrima by the toxicity study. The production of 2PE could be increased substantially by introducing an absorbent into the process. Liquid solvents and solid adsorbents were assessed to increase 2PE production, used as in-situ 2PE adsorbents. Oleyl alcohol was found to be a good solvent for in-situ extractive solvent in M. pulcherrima culture and increase the production to 3.3 g/L which is higher than 2PE tolerance threshold of the yeast. Activated carbon was also found to be an excellent 2PE adsorbents, with maximum Langmuir adsorption capacity up to 0.807 g/g. 2PE synthesis with activated carbon as an in-situ adsorbent can increase 2PE production to 14 g 2PE/L. Finally, the process was scaled to 2L and run in batch, continuous and semi-continuous modes. This study demonstrates that not only is M. pulcherrima a viable organism to produce 2PE but it has the potential to be scaled up and run in a more cost effective semi-continuous mode when coupled to a continuous extraction technique.
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Babapour, Ayda Barid, and Maryam Nadalipour Gavitar. "Investigation of yeast Grown in SSF Dring Biothanol Production from Lignocellusosic Material." Thesis, Högskolan i Borås, Institutionen Ingenjörshögskolan, 2012. http://urn.kb.se/resolve?urn=urn:nbn:se:hb:diva-16498.
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Ethanol produced from lignocellulosic biomass has the potential to become a promisingalternative to gasoline. In this work the simultaneous saccharification and fermentation (SSF)technology was applied for ethanol production from hardwood with focus on cell growth,ethanol production and contamination.The SSF was performed at PH 5.5 and 35°C for different suspended solid concentrations(8%, 10% and 12%) of pretreated birch slurry which contained 16 % total suspended solids.Two different hexose fermenting yeast strain (Ethanol Red) and pentose fermenting yeaststrain were used.Quantifying the concentration of chemical components and metabolites in the fermentationmedium demonstrated that glucose and xylose are the major fermentable sugars in the slurry.The higher load of slurry (12%) represents a higher content of carbohydrates and potentiallyhigher end concentration of ethanol. Moreover, more lactic acid is produced with the lowerload of slurry (8 % or 10 %), presumably due to a result of a less inhibitory environment forbacterial growth. In this context, acetic acid sticks out as the most important inhibitor withconcentrations of 15.2 and 12.5 and 9.7 g/l respectively in the 12 %, 10 % and 8 % (ofsuspended solids) trials. Using pentose fermenting yeast may lead to higher ethanolproduction, lower xylose uptake and lower lactic acid formation. Cell viability and cellvitality determination from fermentation media in all the trails represented a sharplydecreasing trend during the fermentation for both Ethanol Red yeast strain and the pentosefermenting strain yeast strain apparently due to cell decomposition.Program: MSc in Resource Recovery - Industrial Biotechnology
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Reynders, Michael Barry. "Studies on growth, modelling and pigment production by the yeast Phaffia rhodozyma." Master's thesis, University of Cape Town, 1995. http://hdl.handle.net/11427/21500.
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Bibliography: pages 117-123.Within the aquaculture industry a potential has been identified for the pigment astaxanthin. Astaxanthin is the carotenoid responsible for the distinctive coloration of salmonids, crustaceans and certain birds. Due to the fact that animals cannot synthesize carotenoids themselves, it is necessary for these pigments to be present in their food source. In the case of farm-raised salmonids and crustaceans, supplementation of their food with astaxanthin is required. The chemical synthesis of astaxanthin is very costly and complicated. As a result natural, microbial sources of astaxanthin are being investigated. Phaffia rhodozyma is the only yeast known to synthesize astaxanthin as its principle carotenoid. The aim of this dissertation is to present a study investigating the growth and pigmentation of P. rhodozyma, with a view to its commercial production. A P. rhodozyma mutant (UCT-1 N-3693) with a 50% increased total carotenoid content was selected after NTG mutagenesis of the wild strain.
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Gough, Suzanne. "Production of ethanol from molasses using the thermotolerant yeast strain Kluyveromyces marxianus IMB3." Thesis, University of Ulster, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.284833.
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Swana, Jeffrey Ross. "Construction and Analysis of a Modified Yeast Strain for Next Generation Biofuel Production." Digital WPI, 2013. https://digitalcommons.wpi.edu/etd-theses/52.
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Current research efforts are focused on 'second generation biofuels', which includes biofuels produced from lignocellulosic material. Lignocellulosic material is primarily composed of cellulose, a glucose polymer, xylose rich hemicellulose and non-fermentable lignin. Saccharomyces cerevisiae is widely used on an industrial scale for the production of ethanol from glucose; however, native S. cerevisiae does not contain the genes required for fermentation of xylose into ethanol. Others have sequentially expressed trans-genes from xylose fermenting organisms to engineer strains of S. cerevisiae capable of fermenting this pentose. The goal of this thesis was to generate a single cassette of 9 genes which have been shown to ferment xylose and arabinose. The 17 kb DNA fragment harboring all the genes necessary was introduced into the yeast genome using one-step homologous recombination based transformation. Expression of this cassette was verified by demonstrating that the first and last genes on this cassette were transcribed. The modified strain exhibited xylose utilization under microaerobic fermentation conditions. Further genetic and process engineering methods may be employed to improve the yield. The experiments described here demonstrate that generating a functional cassette of pentose fermenting genes is still achievable.
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Grbin, Paul R. "Physiology and metabolism of Dekkera/Brettanomyces yeast in relation to mousy taint production." Adelaide, 1998. http://hdl.handle.net/2440/21644.
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Title page, contents and abstract only. The complete thesis in print form is available from the University Library.Thesis (Ph.D.)--University of Adelaide, Dept. of Horticulture, Viticulture, and Oenology, 1998
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Pilkington, Heather. "Development of a fluidized bioreactor system using immobilized yeast for continuous beer production." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape3/PQDD_0031/NQ68119.pdf.
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Alwan, Zeena. "Neutral lipid production by the yeast Debaryomyces hansenii NCYC102 under different stress conditions." Thesis, University of Sheffield, 2017. http://etheses.whiterose.ac.uk/18660/.
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Oleaginous yeasts are very efficient in the accumulation of triacylglycerol, and are expected to be one of the most important feedstocks for the biofuel industry in the future. Lipid content can be enhanced through physiological stress or genetic manipulation. Debaryomyces hansenii NCYC102 was selected from three different yeast species (also including Yarrowia lipolytica NCYC476 and Cryptococcus curvatus NCYC2904) due to the highest neutral lipid content. The growth rate, osmolytes and neutral lipids were measured in cells grown under different concentrations (0, 0.8, 1.6 M) of NaCl. The maximum content of total osmolytes was found in 0.8 M NaCl YM medium. However, the highest level of glycerol was measured in 1.6 M NaCl grown cells. The main osmolytes identified by 1H NMR spectroscopy were glycerol, arabitol, glucose and trehalose. Debaryomyces. hansenii cells were grown in minimal medium with different carbon/nitrogen ratios using either glucose or glycerol as the sole carbon source along with ammonium sulphate as nitrogen source. Maximal neutral lipid production was observed in 48:0.5 glucose/ammonium sulphate ratio which achieved 1.4-fold increase compared with glycerol-based medium (8 glycerol: 0.25 ammonium sulphate). GC-MS analysis of the transesterified fatty acids showed that palmitic, oleic and stearic acids were the main fatty acids present, under normal and stress conditions (high salt and limited nitrogen source). Deletion of the GUT2 encoding for G3P dehydrogenase increased neutral lipid production up to a 1.4-fold compared to wild type strains. The mutant strains displayed slightly higher cell densities in medium with glucose when compared with wild type strains, while they failed to grow on glycerol as a sole carbon source. Collectively, these results indicate that D. hansenii is a good organism to produce biofuels as it has an intrinsic ability to accumulate neutral lipids and this can be further enhanced by genetic and metabolic engineering.
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Sonq, Jyh Yeong, and 宋至勇. "Autolysis of baker''s yeast for the production of yeast extract." Thesis, 1995. http://ndltd.ncl.edu.tw/handle/16918393562740375602.
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