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1
Josefsson, Peter. « Biochemical modification of wood components ». Licentiate thesis, Stockholm : Fibre and Polymer Technology, KTH, the Royal Institute of Technology, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-4171.
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Gosh, D. « Structural and biochemical investigation of tat components ». Thesis(Ph.D.), CSIR-National Chemical Laboratory Pune, 2019. http://dspace.ncl.res.in:8080/xmlui/handle/20.500.12252/6031.
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Chavan, Uttam Dnyanu. « Chemical and biochemical components of Beach pea (Lathyrus maritimus L.) / ». Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape10/PQDD_0021/NQ54844.pdf.
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Hutchinson, Lisa. « Biochemical analysis of components of the wingless/WNT signalling pathway ». Thesis, Institute of Cancer Research (University Of London), 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.395976.
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Anderson, Shona M. L. « Biochemical and immunological studies of surface components of Fasciola hepatica throughout development ». Thesis, Queen's University Belfast, 1989. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.335984.
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Liu, Jing. « Biochemical and structural studies of key components in the Wnt signaling pathway / ». Thesis, Connect to this title online ; UW restricted, 2008. http://hdl.handle.net/1773/5685.
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Delgado, Callisaya Pedro Angel. « Biochemical components of seminal plasma of llamas (Lama glama) at three ages ». BYU ScholarsArchive, 2002. https://scholarsarchive.byu.edu/etd/5357.
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This study was conducted at the installation of the Rural Academic Unit-Tiahuanaco of the BCU, located in the community of Achaca, third municipal section of Ingavi province, department of La Paz. It is 57 km from the La Paz-Desaguadero international highway, at 68 degrees 42 minutes 28 seconds latitude South by 16 degrees 35 minutes 41 seconds longitude West, at an altitude of 3856 meters above sea level. The study went from October 2000 to September 2001. The study consisted of determining the concentrations of the biochemical components in llama seminal plasma at three ages. Components studied were glucose, inorganic phosphate, creatinine, total protein, albumin, globulins, cholesterol, calcium, potassium, sodium, and magnesium. Twelve male llamas of 3, 4, and 5 years were selected and acquired from the Choquecota area of the Carangas province, department of Oruro. Four animals were chosen at each age and were subjected to a training period of semen collection during 2 months, using the artificial hindquarters designed for this effect. The 6 that best responded to the training were used for the investigation. Eight collections were obtained from each animal over the course of the study, and they were used for laboratory analysis. The results were analyzed using a hierarchical factorial design that involved a mixed analysis (nested and crossed) of the factors of age and collections. (The averages of two collections corresponding to each week were analyzed.) Each weekly collection average per age was an experimental unit. Four experimental units were obtained for each age, and the analysis of the data was done with the SAS statistics package version 6.12. From the analyses done the following results were obtained: the concentrations of glucose (6.246 [plus or minus] 0.716 mg/dl), creatinine (3.459 [plus or minus] 1.27 mg/dl), cholesterol (67.28 [plus or minus] 18.21 mg/dl), potassium (8.249 [plus or minus] 1.78 mEq/L), and sodium (123.187 [plus or minus] 18.39 mEq/L) did not show significant differences between ages or collections (p>0.05). The concentrations of calcium (12.138 [plus or minus] 3.64 mg/dl) and magnesium (1.943 [plus or minus] 0.52 mEq/L) showed significant (p<0.05) differences in age only and not in collections. Globulins (1.574 [plus or minus] 0.51 g/dl) showed differences between collections (p<0.05) but not between ages. Total protein (3.732 [plus or minus] 0.45 g/dl), albumin (2.158 [plus or minus] 0.46 g/dl), and inorganic phosphate (9.42 [plus or minus] 2.42 mg/dl) showed differences both between ages and between collections (p<0.05).
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Shao, Dongmin. « Structural and biochemical studies of the components of the NADPH oxidase in human neutrophils ». Thesis, University College London (University of London), 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.398014.
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Chen, Jui-Lin. « Biochemical Identification of Molecular Components Required for Cyanide Assimilation in Pseudomonas fluorescens NCIMB 11764 ». Thesis, University of North Texas, 1998. https://digital.library.unt.edu/ark:/67531/metadc278624/.
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Utilization of cyanide as a nutritional nitrogen source in P. fluorescens NCIMB 11764 was shown to involve a novel metabolic mechanism involving nonenzymatic neutralization outside of cells prior to further enzymatic oxidation within. Several cyanide degrading enzymes were produced by NCIMB 11764 in response to growth or exposure to cyanide, but only one of these cyanide, oxygenase (CNO), was shown to be physiologically required for assimilation of cyanide as a growth substrate.
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Choudhury, Jayanta. « Studies on the antioxidative and other beneficial properties of some plants from wetlands of North Bengal ». Thesis, University of North Bengal, 2017. http://ir.nbu.ac.in/hdl.handle.net/123456789/2662.
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Hilbi, Hubert Franz Pius Hilbi Hubert Franz Pius Hilbi Hubert Franz Pius Hilbi Hubert Franz Pius. « The malonate decarboxylase enzyme system of Malonomonas rubra : identification, purification and biochemical characterization of components / ». [S.l.] : [s.n.], 1994. http://e-collection.ethbib.ethz.ch/show?type=diss&nr=10766.
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Breit, Claudia Verena [Verfasser], Andrea [Akademischer Betreuer] Musacchio et Daniel [Gutachter] Rauh. « Biochemical studies of spindle assembly checkpoint components / Claudia Verena Breit. Betreuer : Andrea Musacchio. Gutachter : Daniel Rauh ». Dortmund : Universitätsbibliothek Dortmund, 2015. http://d-nb.info/1112268286/34.
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Mekhail-Ishak, Kamilia. « Biochemical components of xenobiotic metabolism in human colon and in murine breast : relation to drug sensitivity ». Thesis, McGill University, 1989. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=74268.
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The biochemical characterization of phase I and phase II components considered to be important in the metabolism of carcinogens and other xenobiotics was examined in two experimental models. This was done to elucidate the relationship between the process of carcinogenesis and the selection of a biochemical phenotype which could result in drug resistance, previously described in rat hepatocarcinogenesis models.A model of mammary carcinogenesis in mice demonstrated no significant biochemical change relating to potential drug resistance among normal, preneoplastic and neoplastic mammary tissues. This is of particular interest since human breast cancer is usually quite drug sensitive at the start of the treatment.
Human colon cancer is probably induced by carcinogenic compounds present in the environmental dietary elements and also exhibits de novo resistance to most antineoplastic drugs. Most of the biochemical elements examined were found to be present in these colon tissues. Additionally, the detoxification pathways including glutathione and its related enzymes, were found to be significantly elevated in colon tumor versus normal adjacent mucosa. The molecular characterization of glutathione S-transferase (GST) isoenzymes using antibodies and cloned molecular probes to specific enzyme forms showed that the anionic form of GST (GST-$ pi)$ is greater in tumors than in the respective normal mucosa. The neutral form (GST-$ mu)$ is conversely decreased in tumor relative to normal. The cationic (GST-$ alpha)$ is present only in 30% of the samples examined, with no difference between tumor and adjacent normal mucosa. The expression of cytochrome P-450 isoenzymes was also examined in a similar fashion; a phenobarbital-inducible form was expressed in most colon tissues examined, and expression of the polycyclic aromatic hydrocarbons-inducible P$ sb3$-450 was present in some colon tissues while mRNA for P$ sb1$-450 was not detected in any. The biochemical alterations found in human colon could be the targets of therapeutic manoeuvers to enhance the efficacy of antineoplastic treatment of human colon cancer.
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Miles, C. M. « Biochemical and developmental studies on protein components of the diacylglcerol transport system in Locusta migratoria migratorioides ». Thesis, University of Hull, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.382977.
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Kunze, Kurt, Hendrik Niemann, Susanne Ueberlein, Renate Schulze, Hermann Ehrlich, Eike Brunner, Peter Proksch et Karl-Heinz van Pée. « Brominated Skeletal Components of the Marine Demosponges, Aplysina cavernicola and Ianthella basta : Analytical and Biochemical Investigations ». Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2013. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-127045.
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Demosponges possess a skeleton made of a composite material with various organic constituents and/or siliceous spicules. Chitin is an integral part of the skeleton of different sponges of the order Verongida. Moreover, sponges of the order Verongida, such as Aplysina cavernicola or Ianthella basta, are well-known for the biosynthesis of brominated tyrosine derivates, characteristic bioactive natural products. It has been unknown so far whether these compounds are exclusively present in the cellular matrix or whether they may also be incorporated into the chitin-based skeletons. In the present study, we therefore examined the skeletons of A. cavernicola and I. basta with respect to the presence of bromotyrosine metabolites. The chitin-based-skeletons isolated from these sponges indeed contain significant amounts of brominated compounds, which are not easily extractable from the skeletons by common solvents, such as MeOH, as shown by HPLC analyses in combination with NMR and IR spectroscopic measurements. Quantitative potentiometric analyses confirm that the skeleton-associated bromine mainly withstands the MeOH-based extraction. This observation suggests that the respective, but yet unidentified, brominated compounds are strongly bound to the sponge skeletons, possibly by covalent bonding. Moreover, gene fragments of halogenases suggested to be responsible for the incorporation of bromine into organic molecules could be amplified from DNA isolated from sponge samples enriched for sponge-associated bacteria.
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Flores, Demetrio Laruta. « Biochemical Components in the Secretion of the Bulb Urethral Glands of Llama (Lama Glama) in Three Ages ». BYU ScholarsArchive, 2002. https://scholarsarchive.byu.edu/etd/5361.
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The present study was performed in the Zoo-Technical Engineering department of the Tiahuanaco Rural Academic Unit of the Bolivian Catholic University. The biochemical components of the secretions produced by the bulb urethral glands of male llama that were studied are glucose, inorganic phosphorus, creatinine, total proteins, albumin, total lipids, cholesterol, and triglycerides. Spectrophotometer standardized techniques were used in nine animals of three, four, and five years of age from communities of the Ingavi province. Groups comprised of three animals each were selected by age criteria. After dissecting and separating the pelvic urethra, gland secretion was obtained by finger pressure. The process was made after beneficiating the animals. The bulb urethral gland secretions present a white color and a hard viscous consistency. The results of biochemical analysis in the three ages are as follows : glucose 535.79 mg/dl; inorganic phosphorus 30.67 mg/dl; creatinine 25.34 mg/dl; total proteins 11.78 g/dl; albumin, 8.596 g/dl; total lipids 1022.55 mg/dl; cholesterol 168.83 mg/dl; and triglycerides, 605.10 mg/dl. Animal age significantly influences on the concentrations of glucose, creatinine, and total lipids with a probability of (p>= 0.05). Animal age has no influence over the concentrations of inorganic phosphorus, total proteins, albumin, cholesterol, and triglycerides with a probability of (p<= 0.05).
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Delgado, Callisaya Pedro Angel. « Componentes bioquimicos del plasma seminal de Llama (Lama glama) en tres edades / ». Diss., La Paz, Bolivia, 2002. http://contentdm.lib.byu.edu/cgi-bin/docviewer.exe?CISOROOT=/Benson&CISOPTR=5495.
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Tesis de grado Presentada a la carrera de Ingenieria Zootecnica, Universidad Catolica Boliviana "San Pablo" Unidad Academica Campesina Tiahuanaco, Carrera Ingeniera Zootechnica.Abstract in Spanish and English.
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Oddone, Anna. « Structural and biochemical characterisation of the yeast exosome component Rrp40 ». Strasbourg 1, 2007. http://www.theses.fr/2007STR13014.
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Chez les cellules eucaryotes, la maturation 3’ et la dégradation de nombreux ARNs nécessitent un complexe protéique, l’exosome. La séquence de l’exosome est conservée de la levure à l’homme et récemment des homologues chez les archaeabactéries ont été décrits. Le complexe est composé de six protéines homologues de la RNase PH et de trois protéines qui contiennent domaines S1 et KH. Bien que la caractérisation fonctionnelle de l’exosome ait progressé au cours de ces dernières années, les structures des sous-unités ainsi que l’architecture du complexe restaient à élucider. L'objectif de ce travail de thèse était de résoudre la structure et caractériser les propriétés biochimiques des sous-unités contenant les domaines S1/KH, et plus particulièrement de la protéine Rrp40, une sous-unité spécifique aux cellules eucaryotes. La structure cristalline de Saccharomyces cerevisiae Rrp40 révèle que Rrp40 comprend un domaine S1 ainsi qu'un domaine KH, modules typiques pour l’interaction avec l’ARN. L’interface entre le domaine S1 et le domaine KH est stabilisée par une séquence GxNG conservée dans les domaines KH de l’exosome uniquement. Des données de résonance magnétique nucléaire révèlent la présence d’un site liant le manganèse à l’interface de ces deux domaines. Expériences de titration par calorimétrie montrent que la protéine Rrp40 de la levure et son homologue Rrp4 de S. Solfataricus ont une très faible affinité intrinsèque pour l’ARN. Toutefois, l’affinité du cœur catalytique de l’exosome de S. Solfataricus pour l’ARN est significativement augmentée en présence de Rrp4, suggérant que de multiples sous-unités pourraient contribuer de façon coopérative à la liaison de l’ARNIn eukaryotes, mRNA turnover, RNA quality control and 3'-end processing of several RNAs require the exosome, a multi-subunit protein complex that is conserved from yeast to human, and of which some archaeal counterparts have recently been described. The exosome core consists of six proteins homologous to bacterial RNase PH nucleases and of three S1-domain containing subunits, predicted to be involved in RNA substrate binding. Although the functional characterisation had considerably progressed since exosome first description, no structural information was available for the complex or its subunits, at the onset of this thesis. This thesis presents the first structure of a component of the eukaryotic exosome core, Rrp40 from Saccharomyces cerevisiae. The crystal structure of the N-terminal deletion construct of Rrp40 reveals that an S1 and an unusual KH domain are tightly packed against each other. The interface between the S1 and the KH domain is stabilised by a GxNG sequence, which is uniquely conserved in exosome KH domains. NMR data reveal the presence of a manganese-binding site at the interface of the two domains. Isothermal titration calorimetry shows that yeast Rrp40 and its archaeal counterpart Rrp4 have alone very low affinity for RNA, despite the fact that both S1 and KH domains are known nucleic acid-binding modules. The affinity of an archaeal core exosome for RNA, however, is significantly increased in the presence of Rrp4, indicating that multiple subunits may contribute to cooperative binding of RNA substrates by the exosome
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Forson, Benedicta. « Biochemical Characterization of the Two-component Monooxygenase System ; Isobutylamine N-hydroxylase (IBAH) and Flavin Reductase (FRED) ». Thesis, Virginia Tech, 2016. http://hdl.handle.net/10919/81454.
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Isobutylamine N-hydroxylase (IBAH) and flavin reductase (FRED) from Streptomyces viridifaciens are part of a two-component flavin-dependent monooxygenase enzyme system that catalyze the conversion of isobutylamine (IBA) to isobutylhydroxylamine (IBHA), a key step in the formation of valanimycin, an azoxy antibiotic. In this work, we present the over-expression, purification and biochemical characterization of this two-component enzyme system. IBAH and FRED were expressed and purified to homogeneity as separate proteins. FRED exhibited the oxidoreductase activity by catalyzing the oxidation of NADPH. The hydroxylation activity of IBAH was confirmed using liquid chromatography – mass spectrometry (LC-MS). Steady state kinetic data showed an oxidation activity of the monooxygenase component which proceeded at 1.97 ± 0.06 s⁻¹ as measured from oxygen consumption and in product formation, the rate was 0.012 ± 0.001 s⁻¹ , suggesting a high degree of uncoupling between product formation and oxygen consumption. In pre-steady state kinetic characterization studies, the FRED-catalyzed reduction of FAD by NADPH occurred at a rate of 10.0 ± 0.2 s⁻¹ and the KM was 490 ± 40 µM. The rate of reduction was ~1.5-fold decreased in the presence of substrate IBA whiles the KM was 500 ± 50 µM. NADH showed a markedly reduced rate of reduction with a kred of 0.34 ± 0.03 s⁻¹ with an apparent KM of 3000 ± 500 µM. The rate of flavin re-oxidation in the absence of monooxygenase IBAH was 4.79 × 10⁻⁹ M-1 s⁻¹. Our results suggest a reaction mechanism for the IBAH monooxygenase system controlled by the oxidation half reaction that may be modulated by a complex formation between the reductase and monooxygenase components.Master of Science in Life Sciences
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Morris, Kristy, et n/a. « Optimisation of the Biocatalytic Component in a Ferricyanide Mediated Approach to Rapid Biochemical Oxygen Demand Analysis ». Griffith University. School of Environmental and Applied Science, 2005. http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20060906.121244.
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A novel rapid method for the determination of biochemical oxygen demand (BOD) has been developed. By replacing oxygen, the terminal electron acceptor in the microbial oxidation of organic substrate, with the ferricyanide ion, a significant increase in the rate of the biochemical reaction could be achieved. This arises from the high solubility of the ferricyanide ion (compared to oxygen); therefore allowing for elevated microbial populations without rapid depletion of the electron acceptor. Therefore, the BOD of a sample can be determined within 1-3 hours compared to 5-days with the standard BOD5 assay. A range of microorganisms were shown to be able to use the ferricyanide ion as an alternative electron acceptor for the biodegradation of a range of organic compounds in the ferricyanide mediated BOD (FM-BOD) assay. The most suitable biocatalyst in the FM-BOD method, however, was shown to be a mixture of microorganisms that was capable of degrading large amounts and types of compounds. These mixed consortia of microorganisms included a synthetic mixture prepared in our laboratory and two commercially available consortia, BODseedTM and Bi-ChemTM. When these seed materials were employed in the FM-BOD assay, the method was shown to closely estimate the BOD5 values of real wastewater samples. The linear dynamic working range of the FM-BOD method was also greatly extended compared to the standard BOD5 assay (nearly 50 times greater) and other oxygen based BOD biosensors. The immobilisation of the microbial consortia by both gel entrapment and freeze-drying methods was shown to greatly reduce the preparation and handling time of the mixed consortia for use in the FM-BOD method. Immobilisation of the mixed microbial consortium in LentiKats®, a PVA hydrogel, resulted in a marked increase in the stability of the biocatalyst. Diffusion limitations resulting from the gel matrix, however, reduced the rate and extent of the bioreaction as well as the linear dynamic working range of the method. Freeze-drying techniques were shown to circumvent some of the limitations identified with gel entrapment for the immobilisation of the mixed consortia. The freeze-dried consortia could be used off-the-shelf and demonstrated reduced diffusional restrictions. A marked decrease in the viability of the microorganisms was observed directly following the freeze-drying process and in subsequent storage. Carrageenan, however, was shown to afford a significant degree a protection to the cells during the freeze-drying process.
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Morris, Kristy. « Optimisation of the biocatalytic component in a ferricyanide mediated approach to rapid biochemical oxygen demand analysis ». Thesis, Griffith University, 2005. http://hdl.handle.net/10072/367552.
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A novel rapid method for the determination of biochemical oxygen demand (BOD) has been developed. By replacing oxygen, the terminal electron acceptor in the microbial oxidation of organic substrate, with the ferricyanide ion, a significant increase in the rate of the biochemical reaction could be achieved. This arises from the high solubility of the ferricyanide ion (compared to oxygen); therefore allowing for elevated microbial populations without rapid depletion of the electron acceptor. Therefore, the BOD of a sample can be determined within 1-3 hours compared to 5-days with the standard BOD5 assay. A range of microorganisms were shown to be able to use the ferricyanide ion as an alternative electron acceptor for the biodegradation of a range of organic compounds in the ferricyanide mediated BOD (FM-BOD) assay. The most suitable biocatalyst in the FM-BOD method, however, was shown to be a mixture of microorganisms that was capable of degrading large amounts and types of compounds. These mixed consortia of microorganisms included a synthetic mixture prepared in our laboratory and two commercially available consortia, BODseedTM and Bi-ChemTM. When these seed materials were employed in the FM-BOD assay, the method was shown to closely estimate the BOD5 values of real wastewater samples. The linear dynamic working range of the FM-BOD method was also greatly extended compared to the standard BOD5 assay (nearly 50 times greater) and other oxygen based BOD biosensors. The immobilisation of the microbial consortia by both gel entrapment and freeze-drying methods was shown to greatly reduce the preparation and handling time of the mixed consortia for use in the FM-BOD method. Immobilisation of the mixed microbial consortium in LentiKats®, a PVA hydrogel, resulted in a marked increase in the stability of the biocatalyst. Diffusion limitations resulting from the gel matrix, however, reduced the rate and extent of the bioreaction as well as the linear dynamic working range of the method. Freeze-drying techniques were shown to circumvent some of the limitations identified with gel entrapment for the immobilisation of the mixed consortia. The freeze-dried consortia could be used off-the-shelf and demonstrated reduced diffusional restrictions. A marked decrease in the viability of the microorganisms was observed directly following the freeze-drying process and in subsequent storage. Carrageenan, however, was shown to afford a significant degree a protection to the cells during the freeze-drying process.Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Environmental and Applied Science
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Ibrahim, Iskander Mohamed. « Biochemical characterisation of the cyanobacterial Hik2-Rre1 two-component regulatory system ». Thesis, Queen Mary, University of London, 2013. http://qmro.qmul.ac.uk/xmlui/handle/123456789/8509.
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Two-component signal transduction systems (TCS) consist of a sensor histidine kinase and a response regulator. TCS are ubiquitous in prokaryotes, but found only in some eukaryotes. TCS mediate adaptation to various environmental changes in bacteria, plants, fungi, and protists. Histidine kinase 2 (Hik2) is a sensor histidine kinase found in all cyanobacteria. The Hik2 homologue known as Chloroplast Sensor Kinase is found in algae and plants, where it is encoded by the nuclear genome and it is targeted to chloroplasts. CSK couples the redox state of the photosynthetic electron transport chain to chloroplast gene transcription. This thesis describes biochemical characterisation of the signal transduction mechanism of Hik2 and its response regulator (Rre) partners in order to clarify the Hik2-Rre two-component signal transduction pathway. Results presented in this thesis illustrate that the autophosphorylation activity of the full-length Hik2 protein is specifically inhibited by sodium ions. An autophosphorylation event of a histidine kinase is the result of homodimerisation and is followed by trans or cis-autophosphorylation of each monomer on its conserved histidine residue. Chemical crosslinking revealed that the Hik2 protein exists predominantly as a phosphorylated (autokinase active) monomer, tetramer, and higher-order oligomeric complexes. The functions of these different oligomeric states of Hik2 are also discussed. From a previous study, which was based on an observation from a yeast two-hybrid assay, Hik2 was proposed to form a two-component pair with Rre1 and RppA. However, no further evidence was presented to support either direct interaction or direct phosphotransfer activity of the Hik2-Rre pair. This thesis confirms interaction of Hik2-Rre1 and Hik2-RppA two-component ! %! pairs using an in vitro pull-down assay and phosphotransfer kinetics. Finally, a model is proposed for the Hik2 based two-component signal transduction pathway.
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Gendron, Joshua Martin. « Genetic, biochemical, and developmental analyses of the brassinosteroid signaling component, BZR1 / ». May be available electronically:, 2008. http://proquest.umi.com/login?COPT=REJTPTU1MTUmSU5UPTAmVkVSPTI=&clientId=12498.
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Chidley, H. G. « Biochemical and molecular characterization of a volatile component of mango (mangifera indica L.Cv.alphanso) flavor ». Thesis(Ph.D.), CSIR-National Chemical Laboratory, Pune, 2015. http://dspace.ncl.res.in:8080/xmlui/handle/20.500.12252/5894.
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Grancell, Adam Scott 1969. « Biochemical characterization of CBF3, an essential DNA-binding component of the yeast kinetochore ». Thesis, Massachusetts Institute of Technology, 1998. http://hdl.handle.net/1721.1/49652.
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Zhang, Lei. « Biophysical and biochemical investigation of the structure of chloroplast twin arginine transport component Hcf106 ». Miami University / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=miami1429539744.
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Cranford-Smith, Tamar. « Genetic, biochemical and structural characterisation of YecA, a novel component of the bacterial Sec machinery ». Thesis, University of Birmingham, 2018. http://etheses.bham.ac.uk//id/eprint/8354/.
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The Sec pathway provides a mechanism for the translocation of proteins across or into the cytoplasmic membrane. In bacteria, SecA is a core component of the Sec machinery. YecA has a 20-amino acid sequence at its carboxy-terminus that has high sequence identity to the zinc-binding domain at the carboxy-terminus of SecA. This study provides evidence to show that YecA is a novel component of the Sec machinery of E. coli. The yecA gene is not essential for the viability of E. coli but the deletion of yecA interferes with Sec-dependent translocation and the combined deletion of the yecA and secB genes results in a severely cold-sensitive phenotype. The genetic investigations were supported by biochemical evidence that suggests that YecA improves the translocation-coupled ATPase activity of SecA. Structural investigations suggest that YecA is a monomer in solution. The α-helical domain that forms the main body of YecA is connected via a short linker with limited flexibility to an independent metal-binding domain that has two conformations. The purification of YecA suggested the presence of iron. Biophysical experiments were used to confirm the interaction of the YecA metal-binding domain with iron. This study provides evidence for an additional component of the translocation machinery.
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Pratt, Elizabeth Stratton. « Genetic and biochemical studies of Adr6, a component of the SWI/SNF chromatin remodeling complex / ». Thesis, Connect to this title online ; UW restricted, 2001. http://hdl.handle.net/1773/10288.
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Wojnowska, M. « Biochemical and structural characterisation of a two-component signalling system downstream of bacteriophytochrome photoreceptor 1 in Rhizobium NT-26 ». Thesis, University College London (University of London), 2013. http://discovery.ucl.ac.uk/1388782/.
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Prokaryotic signal transduction frequently involves two-component systems (TCSs), typically comprising a sensor histidine kinase (HK) and a response regulator (RR). Via a conserved phosphotransfer reaction, TCSs couple the detection of diverse stimuli with appropriate responses. The initial aim of this project was to characterise the two-component “signalome” of an arsenite oxidiser, Rhizobium NT-26, in the context of the environmental niche, and compare it to “signalomes” of other bacterial species. A light-sensing HK, bacteriophytochrome photoreceptor 1 (BphP1), was thus identified. Previous studies indicated that BphP1 and the members of its gene cluster - two single-domain RRs and a hybrid RR/HK protein, ExsG - may constitute a TCS. Functional characterisation revealed that BphP1 initiates a branched signalling pathway; however, the mediated output could not be identified. ExsG, a HWE-type HK, was shown to possess dual HK/RR activity and act as a novel type of signalling switch: phosphorylation of the N-terminal receiver domain downregulates the autokinase activity of the C-terminal kinase core. Ion mobility-mass spectrometry analysis indicated that while Asp62 phosphorylation stabilises the “closed” form of ExsG, nucleotide binding stabilises the “open” conformation. These observations led to a model in which phosphorylation of the receiver domain precludes ATP binding and thus inhibits autokinase activity. Furthermore, ExsG was demonstrated to hexamerise via the HWE core, which makes it the first non-dimeric HK. Notably, however, the basic unit of the hexameric assembly is a homodimer, and the HWE core shares homology with canonical HK cores. The results presented herein broaden the current knowledge on TCSs and identify previously unreported mechanisms involved in two-component signal transduction. The members of the BphP1 signalling cascade enrich the pool of modular communication units that can be exploited in engineering artificial signalling networks, biosensors and microorganisms with novel functionalities.
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Hopkins, Adam P. « Molecular and biochemical characterisation of SiaP as a sialic acid binding protein component of a TRAP transporter of sialic acid ». Thesis, University of York, 2010. http://etheses.whiterose.ac.uk/1030/.
Texte intégralRésumé :
Sialic acid utilisation plays an important role in the growth and persistence of the obligate human mucosal pathogen Haemophilus influenzae, which causes respiratory tract infections, septicaemia and meningitis. Like many other bacteria, H. influenzae can use host-derived sialic acids as carbon, nitrogen and energy sources, but also as a terminal modification on the LPS to better evade the human immune system. H. influenzae takes up exogenous sialic acid via a tripartite ATP-independent periplasmic (TRAP) transporter, SiaPQM. This possesses an extracytoplasmic substrate binding protein (SBP), SiaP, which binds the substrate in the periplasm and delivers it to the specific membrane permease, SiaQM. SiaP contains two globular domains, which close around the substrate upon binding. Here, the mechanism of sialic acid binding by SiaP is investigated using site-directed mutagenesis of residues in the ligand binding site and analogues of sialic acid. These, and several mutations on the surface of SiaP, were investigated for their effect on transport by SiaPQM in vitro, using SiaQM reconstituted into proteoliposomes, and in vivo, using expression of siaPQM in an E. coli strain lacking its native sialic acid transporter, NanT. It is demonstrated that stabilisation of the carboxylate group of sialic acid by the totally conserved Arginine-147 is important for high-affinity ligand binding, but is not essential for transport. Mutation of Aparagine-150 to Aspartate abolishes the function of the transporter without affecting ligand binding, suggesting the existence of a critical interaction between the components of the transporter. The catabolism of the sialic acid analogues was also examined in E. coli expressing different sialic acid transporters. This indicates that a wide variety of sialic acid analogues are potential carbon sources in many pathogenic bacteria.
31
Emmerich, Christoph H. « Biochemical and functional characterisation of the Linear Ubiquitin Chain Assembly Complex (LUBAC) as a component of the TNF-Receptor 1 signalling complex ». Thesis, Imperial College London, 2010. http://hdl.handle.net/10044/1/6186.
Texte intégralRésumé :
Tumour necrosis factor (TNF) plays a critical role in inflammatory processes and is involved in the regulation of immune responses. Depending on the cellular context, TNF initiates a complex cascade of signalling events that can lead to induction of proinflammatory cytokines, cell proliferation, differentiation or cell death. Most of the pleiotropic effects of TNF are mediated by the death-domain (DD) containing TNF-R1. Ligand-induced trimerisation of TNF-R1 leads to the formation of an intracellular multiprotein complex, the TNF-R1 signalling complex (TNF-RSC). To be able to comprehend the complex network of signalling pathways emanating from TNF-R1, the TNF-RSC and its composition need to be understood at the molecular level. Using a modified tandem affinity purification approach, HOIL-1 and HOIP were identified as two novel, functional components of the native TNF-RSC. Together they were shown to form a linear ubiquitin chain assembly complex (LUBAC), catalysing the formation of linear head-to-tail ubiquitin chains. LUBAC mediates ubiquitination of NEMO with linear ubiquitin chains, required for efficient NF-kB activation following TNF stimulation. In this thesis, it could be demonstrated that the stimulation-dependent recruitment of LUBAC to the TNF-RSC is impaired in cIAP1/2-deficient mouse embryonic fibroblast (MEF) cell lines. Furthermore, it was shown that the E3 ligase activity of cIAPs, but not of TRAF2, is required for HOIL-1 recruitment to the TNF-RSC. This result, together with the ability of HOIL-1 and HOIP to bind polyubiquitin chains of various linkage types, suggests that LUBAC is recruited to the TNF-RSC via cIAP-generated ubiquitin chains. It was also found that LUBAC enhances NEMO interaction with the TNF-RSC, that it stabilises this protein complex and that LUBAC is required for efficient TNF-induced activation of NF-kB and JNK, resulting in apoptosis inhibition. Finally, it is shown in this thesis that the catalytic activity of LUBAC is required for stabilisation of the TNF-RSC, thereby adding a novel form of ubiquitin linkage to TNF signalling. The identification of HOIL-1 and HOIP as functional constituents of the TNF-RSC provides evidence that LUBAC is an important regulator at the apex of TNF-induced signalling cascades and increases the combinatorial complexity of ubiquitin modifications within this receptor complex.
32
Newcombe, Anthony Richard. « The biochemical role of the small G protein Rac1 in cell signalling pathways : interaction with RhoGDI and the phagocyte NADPH oxidase component, p67'p'h'o'x ». Thesis, University College London (University of London), 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.342224.
Texte intégral
33
Szwagierczak, Aleksandra. « Biochemical and biophysical characterization of the lyase isomerase PecE/PecF complex, nicastrin the transmembrane component of the gamma-secretase complex and structural investigations of the genomic islands integrases ». kostenfrei, 2009. https://mediatum2.ub.tum.de/node?id=829180.
Texte intégral
34
Kothamachu, Varun Bhaskar. « An investigation into dynamic and functional properties of prokaryotic signalling networks ». Thesis, University of Exeter, 2016. http://hdl.handle.net/10871/26597.
Texte intégralRésumé :
In this thesis, I investigate dynamic and computational properties of prokaryotic signalling architectures commonly known as the Two Component Signalling networks and phosphorelays. The aim of this study is to understand the information processing capabilities of different prokaryotic signalling architectures by examining the dynamics they exhibit. I present original investigations into the dynamics of different phosphorelay architectures and identify network architectures that include a commonly found four step phosphorelay architecture with a capacity for tuning its steady state output to implement different signal-response behaviours viz. sigmoidal and hyperbolic response. Biologically, this tuning can be implemented through physiological processes like regulating total protein concentrations (e.g. via transcriptional regulation or feedback), altering reaction rate constants through binding of auxiliary proteins on relay components, or by regulating bi-functional activity in relays which are mediated by bifunctional histidine kinases. This study explores the importance of different biochemical arrangements of signalling networks and their corresponding response dynamics. Following investigations into the significance of various biochemical reactions and topological variants of a four step relay architecture, I explore the effects of having different types of proteins in signalling networks. I show how multi-domain proteins in a phosphorelay architecture with multiple phosphotransfer steps occurring on the same protein can exhibit multistability through a combination of double negative and positive feedback loops. I derive a minimal multistable (core) architecture and show how component sharing amongst networks containing this multistable core can implement computational logic (like AND, OR and ADDER functions) that allows cells to integrate multiple inputs and compute an appropriate response. I examine the genomic distribution of single and multi domain kinases and annotate their partner response regulator proteins across prokaryotic genomes to find the biological significance of dynamics that these networks embed and the processes they regulate in a cell. I extract data from a prokaryotic two component protein database and take a sequence based functional annotation approach to identify the process, function and localisation of different response regulators as signalling partners in these networks. In summary, work presented in this thesis explores the dynamic and computational properties of different prokaryotic signalling networks and uses them to draw an insight into the biological significance of multidomain sensor kinases in living cells. The thesis concludes with a discussion on how this understanding of the dynamic and computational properties of prokaryotic signalling networks can be used to design synthetic circuits involving different proteins comprising two component and phosphorelay architectures.
35
Jalal, Ahmed Hasnain. « Multivariate Analysis for the Quantification of Transdermal Volatile Organic Compounds in Humans by Proton Exchange Membrane Fuel Cell System ». FIU Digital Commons, 2018. https://digitalcommons.fiu.edu/etd/3886.
Texte intégralRésumé :
In this research, a proton exchange membrane fuel cell (PEMFC) sensor was investigated for specific detection of volatile organic compounds (VOCs) for point-of-care (POC) diagnosis of the physiological conditions of humans. A PEMFC is an electrochemical transducer that converts chemical energy into electrical energy. A Redox reaction takes place at its electrodes whereas the volatile biomolecules (e.g. ethanol) are oxidized at the anode and ambient oxygen is reduced at the cathode. The compounds which were the focus of this investigation were ethanol (C2H5OH) and isoflurane (C3H2ClF5O), but theoretically, the sensor is not limited to only those VOCs given proper calibration.
Detection in biosensing, which needs to be carried out in a controlled system, becomes complex in a multivariate environment. Major limitations of all types of biosensors would include poor selectivity, drifting, overlapping, and degradation of signals. Specific detection of VOCs in multi-dimensional environments is also a challenge in fuel cell sensing. Humidity, temperature, and the presence of other analytes interfere with the functionality of the fuel cell and provide false readings. Hence, accurate and precise quantification of VOC(s) and calibration are the major challenges when using PEMFC biosensor.
To resolve this problem, a statistical model was derived for the calibration of PEMFC employing multivariate analysis, such as the “Principal Component Regression (PCR)” method for the sensing of VOC(s). PCR can correlate larger data sets and provides an accurate fitting between a known and an unknown data set. PCR improves calibration for multivariate conditions as compared to the overlapping signals obtained when using linear (univariate) regression models.
Results show that this biosensor investigated has a 75% accuracy improvement over the commercial alcohol breathalyzer used in this study when detecting ethanol. When detecting isoflurane, this sensor has an average deviation in the steady-state response of ~14.29% from the gold-standard infrared spectroscopy system used in hospital operating theaters.
The significance of this research lies in its versatility in dealing with the existing challenge of the accuracy and precision of the calibration of the PEMFC sensor. Also, this research may improve the diagnosis of several diseases through the detection of concerned biomarkers.
36
Jhan, Jheng-Hao, et 詹正豪. « Studies on flavor components and biochemical changes during processing of raw ham ». Thesis, 2006. http://ndltd.ncl.edu.tw/handle/96752563999431352137.
Texte intégralRésumé :
碩士國立屏東科技大學
食品科學系
94
The changes of contents of chemical composition (moisture, crude protein, crude fat, sodium chloride), flavor components and the value of water activity (Aw), pH were examined during processing of raw ham. Effect of salt on the protein of raw ham during curing period by DSC and texture analyser were also studied. The results obtained were as followed. The value of Aw went down from 0.99 of the fresh ham to 0.91 of the cured ham, as moisture content decreased too, but the crude protein and crude fat content increased relatively during the curing process. The salt contents of ham increased from 0.3% of raw meat to 3.81% after curing day1 by 6% (w/w) NaCl addition, and then its contents was almost constant (6.5%). The value of pH went down from 6.06 of the fresh ham to 5.86 and then up to 5.92 of the finished product. Contents of each free amino acid and ammonia in the ham were increased during the processing. Especially increases of threonine, serine, glutamic acid, alanine, valine, leucine and lysine etc. The DSC analysis, it became clear that denaturation of the meat mainly occurred on myosin by salting. As the denaturation proceed, the enthalpy change, ΔH of 1.212 cal/g rapidly decreased to 0.824 cal/g on curing day3, and then gradually decreased to 0.727 cal/g. Rupture strength of the meat increased during process as a whole, the change of dynamic rigidity of the pork meat pastes by heating exhibited the tendency to decrease at the temperature range from 52℃ to 58℃ after curing day7. The result was considered to come from the structure change of the sample due to thermal denaturation of myosin.
37
Kishore, Kakani Naga. « Molecular and biochemical characterization of viral and vector components required for cucumber necrosis virus transmission ». Thesis, 2004. http://hdl.handle.net/2429/17230.
Texte intégralRésumé :
Natural transmission of Cucumber necrosis virus (CNV) occurs via zoospores of
the chytrid fungus Olpidium bornovanus. Transmission involves specific adsorption of
virus particles onto the zoospore plasmalemma prior to infestation of cucumber roots by
virus bound zoospores. In order to determine if specific regions of the CNV capsid are
involved in transmission, several naturally occurring CNV transmission mutants were
isolated and characterized. Analysis of the mutants showed that the CNV trimer cavity at
the particle quasi three-fold axis plays an important role in transmission, and, moreover
that the reduction in transmission is at least partially due to the reduced ability of
mutants to bind to zoospores. In vitro virus/zoospore binding studies have shown that
pre-treatment of zoospores with trypsin and sodium periodate each decrease CNV
binding by approximately 80%, whereas no reduction in binding was found when
zoospores were treated with phospholipase C. These studies suggest an important role for
zoospore proteins and/or glycoprotein(s) in virus attachment. In virus overlay assays,
CNV virions bound to specific-sized zoospore proteins, but CNV transmission mutants
showed little or reduced binding. Several sugars were used to study their inhibitory
potential on CNV binding to zoospores in vitro. It was found that a variety of mannose-containing
sugars inhibited CNV binding to zoospores whereas several others did not.
These studies suggest that the putative zoospore receptor may be a mannose-containing
glycoprotein.
Many animal virus particles undergo conformational changes upon binding to
their cellular receptors. CNV is an icosahedral virus and like many other isometric plant
viruses, undergoes expansion in the presence of EDTA at an alkaline pH. In the case of CNV, we have demonstrated that during expansion, the internally located coat protein
RNA binding domain (R) and arm domains translocate to the particle exterior, becoming
protease sensitive. Protease digestion experiments of zoospore-bound virus have revealed
that CNV undergoes conformational change upon binding to zoospores and that the
conformationally altered virion resembles the swollen conformation. In addition, we have
found that a CNV mutant defective in vector transmission is unable to undergo this
conformational change. This is the first time that conformational change in a plant virus
particle has been shown to be essential for vector transmission.Land and Food Systems, Faculty of
Graduate
38
林君霏. « Biochemical Changes during Storage and Seasonal Variation of Extractive Components of Hard Clam (Meretrix Lusoria) ». Thesis, 1995. http://ndltd.ncl.edu.tw/handle/98680806149642358192.
Texte intégralRésumé :
碩士國立海洋大學
水產食品科學研究所
83
Biochemical analyses were carried out for understanding the changes of freshness and taste coponents of hard clam (Meretrix lusoria) during storage at different temperatures. The seasonal variations of proximate composition, extractive nitrogenous components and glycogen in hard clam were also studied. The biochemical changes of anaerobic metabolism in hard clam were discussed as well. The pH, volatile basic nitrogen (VBN), trimethylamine (TMA) ammonia, drip and survival showed marked change at the stage of decomposition The pH was a reliable fressness indicator for hard clam, and its value of 7.0 was proposed as a limit of quality acceptability. VBN was also a suitable indicator, and its value of >16 mg/100g for samples stored at 0∼10℃, and >25 mg/100g for samples stored at 1525℃, was the level indicating initial decomposition. The obvious increase of TMA and ammonia, but the marked decrease of glycogen and drip were found at the stage of decomposition. Most of clams also died at this stage. TMA ammonia, glycogen, drip and survival seemed not be good freshness indicators for hard clam. The changes of nucleotides and related compounds during storage were temperature-dependent. The K and K'' value of 40%, and the Kc value of 45% for samples stored at 5, 10, 15 and 25℃ were proposed as the limit values for comsumption. The contents of total free amino acids and taste amino acids (alanine, glycine, glutamic acid and arginine) were increased with the time of storage at 5 and 10℃ while those of other temperatures showed little change or decrease The total organic acids and succinic acid also increased during storage.In conclusion, the temperature around 5 and 10℃ was more suitalble for storing hard clam than other temperatures. Seasonal variations of proximate composition extractive nitrogenous components and glycogen in hard clam were investigated from November 1993 to November 1994. No marked changes of proximate composition in samples were found. The amouts of glycogen in hard clam sampled in summer and autumn were higher than those in other seasons. The contents of total free amino acids in hard clam were ranged from 913 to 1299mg/100g, and the taste amino acids ranged from 289 to 524 mg/100g. Both were found higher in samples of July and September. The levels of nucleotides and related compounds in hard clam of May and July were higher than those of other season samples. The contents of organic acids and succinic acid in samples of july an September were liigher. The clam purchased in summer and autumn were probably more tasty due to the higher amounts of glycogen and free amion acids.
39
CHEN, ZHI-YING, et 陳芝瑩. « Effects of different starters on the biochemical changes and the flavor components of Lao-chao fermentation ». Thesis, 1990. http://ndltd.ncl.edu.tw/handle/27634501326120901395.
Texte intégral
40
Lu, Chung-Hao, et 呂仲浩. « Changes of blood biochemical components and mammary gland function oflactating Holstein cows in response to environmental high temperature ». Thesis, 1998. http://ndltd.ncl.edu.tw/handle/25505694410234118490.
Texte intégral
41
Liu, Hsu-chun, et 劉旭鈞. « Biochemical changes and the flavor components analysis during the fermentation of rice vinegar and red yeast rice vinegar ». Thesis, 2004. http://ndltd.ncl.edu.tw/handle/20757320307469004687.
Texte intégralRésumé :
碩士大同大學
生物工程學系(所)
92
Abstract In this study, rice vinegars and red yeast rice vinegars were prepared. The biochemical changes of rice vinegars and red yeast rice vinegars during koji-making, alcohol fermentation, acetic acid fermentation and aging periods were under investigation. Meanwhile, aromatic volatiles of rice vinegars and red yeast rice vinegars were also identified and quantified. The results revealed that red yeast rice could be obtained after culturing Monascus ruber BCRC31535 on rice for 10 d. The content of monacolin k increased to 45.3 mg/kg dry wt. Addition of red yeast rice to rice koji decreased the α-amylase activity of rice koji. During alcohol fermentation of rice moromi and red yeast rice moromi, the amount of reducing sugar decreased and the amount of alcohol increased with increasing time. The concentration of ethanol in rice moromi and red yeast rice moromi increased to 10% after 5 d of alcohol fermentation. The content of red pigment in red yeast rice moromi also increased. However, the content of red pigment content in rice moromi was much lower. The concentration of monacolin k in red yeast rice moromi was below detection limit. During acetic acid fermentation of rice wine by Acetobacter aceti BCRC 12325, the concentration of ethanol decreased and acetic acid formed. The final concentration of acetic acid of rice vinegar and red yeast rice vinegar was 4.7% and the content of red pigment in red yeast rice vinegar slightly decreased at the end of fermentation. The aroma compounds identified in rice vinegar and red yeast rice vinegar were similar. Twenty-four compounds, including 7 alcohols, 7 esters, 6 acids, 2 aldehydes and 2 ketones, were identified. The amount of 2,3-butanediol was highest in rice vinegar and red yeast rice vinegar, followed by 2-phenylethanol. After acetic acid fermentation for 48h, the amount of 2,3-butanediol significantly increased. However, the amounts of most of the aroma compounds decreased after aging for 2 months.
42
Oppizzi, Maria Luisa [Verfasser]. « Genetic, biochemical, and electron microscopic analysis of components involved in transcription coupled mRNA export / presented by Maria Luisa Oppizzi ». 2004. http://d-nb.info/972361774/34.
Texte intégral
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Urzica, Eugen [Verfasser]. « Biochemical analysis of essential components involved in mitochondrial and cytosolic iron-sulfur protein biogenesis in Saccharomyces cerevisiae / vorgelegt von Eugen Urzica ». 2007. http://d-nb.info/985310200/34.
Texte intégral
44
Rajvanshi, Pradumn K. « Molecular and biochemical studies on components of transcription regulation, tropane alkaloid biosynthesis and signal transduction in the medicinal plant Datura metel L ». Thesis, 2021. https://etd.iisc.ac.in/handle/2005/5552.
Texte intégralRésumé :
Tropane alkaloids (TA) are among the oldest medicines known to humankind as they possess hallucinogenic and poisonous properties. Datura metel, an herbaceous plant, belongs to the family Solanaceae whose members are well known to produce tropane alkaloids. The isolation of the key biosynthetic enzymes, transcription factors, and protein kinases and their corresponding genes, is important in understanding TA biosynthesis and in developing metabolic engineering approaches to enhance the amount of tropane alkaloids in plants. Putrescine-N- methyltransferase (PMT; EC 2.1.1.53) is the first enzyme in the tropane alkaloid and nicotine biosynthetic pathway and it catalyzes the S-adenosylmethionine (SAM) dependent N-methylation of putrescine to methyl-putrescine. In the present study, Datura metel putrescine methyltransferase (DmPMT) protein has been biochemically and biophysically characterized. The initial rate kinetic study for DmPMT showed the Vmax 24.47 nKat/mg, Km for Putrescine 255.5 M and Km for SAM were 128.2M. The ordered Bi Bi kinetic mechanism best describes the N-methyl-transferase reaction catalyzed by DmPMT. Upon structural analysis of this N-terminus β-sheet and putrescine binding pocket, we could predict that the interaction of Trp-64 of N-term to the amino group of Putrescine through Cation-π interaction. Most probably, this interaction is crucial for keeping Putrescine in better orientation towards SAM.
Here we report the isolation of transcription factor DORA and DmMYB from Datura metel. q-RTPCR showed that expression of the DORA gene was strongly induced by treatment with fungal extract and moderately induced by methyl jasmonate plus wounding treatments. The Gel mobility shift assay showed that the recombinant DORA protein could bind specifically to GCC box, a cis-acting element present in the promoter regions of several target genes regulated by AP2 domain-containing transcription factor. DmMYB also exhibited sequence-specific DNA-binding. The binding constant for DmMYB-DNA interaction (Kd) was calculated to be 8.1*10-9 M, and the stoichiometry of the interaction was 1:1. 3D homology model of the DmMYB-DNA complex was constructed, confirming the role of TRP-17 and TRP-89 in complex formation. Further, our model highlighted a possible functional role of DmMYB-Mg2+ binding.
In the present study, we have isolated a full-length CDPK cDNA, DmCDPK1 (GenBank accession number ABY28389), from Datura metel by RT-PCR and rapid amplification of cDNA ends (RACE) methods. The recombinant DmCDPK1 with His-tag was heterologous expressed in E. coli and purified using Ni-NTA affinity chromatography. Biochemical analyses showed that both activities of autophosphorylation and phosphorylation of histone III-S substrate by DmCDPK1 protein are dependent on calcium. The kinase activity of the recombinant enzyme was calmodulin independent and sensitive to CaM antagonists, W7, and calmidazolium.CSIR, IISC, ICMR, DST
45
Chen, Yen-Chi, et 陳彥錡. « Biochemical Binding Characterization of Human Progesterone Receptor Membrane Component 1 ». Thesis, 2013. http://ndltd.ncl.edu.tw/handle/46377342883670419342.
Texte intégralRésumé :
碩士國立屏東教育大學
化學生物系碩士班
101
Human progesterone receptor membrane protein (Progesterone Receptor Membrane Component 1 , PGRCM 1) the study was to investigate the protein; Recent studies have pointed out, PGRMC1 Council and heme binding, but its specific physiological role has not yet been identified in the organism. In addition, although the name PGRCM 1 literature that progesterone receptor membrane protein for the human, but in fact do not directly bind progesterone, but with an unknown protein steroid / drug binding protein (S/D-BP) for Progesterone binding message passing and metabolism; expressed concern for the results of this literature. Therefore, in order to further understand the PGRCM 1 and progesterone combined with the actual situation, this study lab colleagues previously applied transgenic technology in Escherichia coli (E.coli) in a large number of performance PGRMC1; and were added to the culture of Heme precursor ALA (5-Aminolevul Acid Hydrochloride) in the culture medium; and in developing purification after adding hemin (Hemin Chloride) reconstruction, obtained by these two methods PGRMC1 iron containing metal center and comparison. We further use of UV - visible spectroscopy (UV-Vis) confirm PGRMC1 with metal centers of iron oxidation state (Fe3 +), and the use of quartz crystal microbalance QCM (Quartz Crystal Microbala) analysis of the binding constant between progesterone; also Discussion between PGRMC1 and binding of Heme. The results of this thesis show, PGRMC1 does not directly bind with Progesterone, but PGRMC1 active center Heme reconstruction may impact combined with Progesterone situation; while PGRMC1 and Heme Kd value can be achieved between 10-8.
46
Hariharan, Vignesh Narayan. « Biochemical and Functional Characterization of the Mycobacterial PdtaS-PdtaR Two Component System ». Thesis, 2018. https://etd.iisc.ac.in/handle/2005/5468.
Texte intégralRésumé :
Two-Component Systems (TCSs) are primarily prokaryotic modes of cell signaling involving a receptor sensor kinase (SK) that senses stimuli and a response regulator (RR) that effects a response, usually at the level of transcriptional regulation. It has been shown that the human pathogen Mycobacterium tuberculosis uses TCSs to adapt to various conditions inside the host during infection and they play roles in host cell entry, intracellular survival and division as well as hypoxia and dormancy. However, one of the major roadblocks in our understanding of TCSs stem from the lack of identified ligands that activate TCS SKs. Most studies involving TCSs have made use of gene knockouts to study the TCS in its ‘off’ state and derive possible functions of the TCS from this data. Currently, only 3 out of 13 SKs have known ligands and 4 out of a total of 12 paired TCSs are of unknown function.
47
Caria, Sofia. « Biochemical and structural studies on the PhoR-PhoB two-component regulatory system from Deinococcus radiodurans ». Doctoral thesis, 2010. http://hdl.handle.net/10451/2241.
Texte intégralRésumé :
Tese de doutoramento, Farmácia (Microbiologia), Universidade de Lisboa, Faculdade de Farmácia, 2010.This thesis concerns studies of a two-component system (TCS) from Deinococcus radiodurans bacterium. TCSs work based on the activity of two proteins: the histidine kinase and the response regulator. Histidine kinases are responsible for several reactions of this mechanism: autophosphorylation, phosphotransfer and phosphatase activity. The mechanism is initiated by an autophosphorylation, induced by a signal from the environment. These proteins are composed of two domains: the dimerisation and histidine phosphorylation domain (DHpD), and the catalytic ATPase domain (CAD). The response regulator is activated by the histidine kinase, through a phosphotransfer reaction. The response regulators are composed by a receiver domain (RD) that will receive the phosphoryl group from the histidine kinase on the aspartate residue, and an effector domain which commonly consists of a DNA-binding domain (DBD). Upon phosphorylation of the RD of the response regulator the protein suffers a conformation change that increases its binding affinity to specific DNA region. The work focuses on the PhoR-PhoB system, a signalling mechanism that gets activated upon environmental phosphate starvation. The activation of PhoB response regulator, through phosphotransfer from PhoR histidine kinase, leads to activation of the last protein with consequent binding to a specific DNA sequence (Pho box), regulating the Pho regulon. The Pho system has been studied in different bacteria using different kinds of approaches. However, the knowledge of this system in D. radiodurans concerns the annotation of the genes that encode the two proteins of the mechanism: PhoR (DR2244) and PhoB (DR2245). This thesis presents the structural characterisation of both full-length proteins and their domains, through small angle X-ray solution scattering (SAXS) analysis. Biochemical data and the X-ray structure of the PhoRCAD domain (resolution 2.25 Å) complement those studies. The biochemical data demonstrates the PhoR autophosphorylation activity as well as phosphoryl transfer activity to PhoBRD.
Esta dissertação refere-se ao estudo estrutural e bioquímico de um dos sistemas de dois componentes (TCS) de Deinococcus radiodurans. O TCSs baseia-se na actividade de duas proteinas: a quinase de histidina e o regulador de resposta. As quinases de histidinas efectuam diversas reacções destes sistemas como autofosforilação, transferência de fosfato e actividade de fosfatase. O mecanismo é iniciado pela autofosforilação da proteína, sendo esta reacção induzida por um sinal ambiental específico. Estas proteínas são compostas dois domínios: o domínio de dimerização e fosforilação de histidina (DHpD) e o domínio catalítico de ATPase (CAD). O regulador de resposta é activado através de uma reacção de fosfotransferência da quinase de histidina; sendo a proteína composta por domínio receptor (RD), que é fosforilado pela quinase de histidina, no aspartato; e pelo domínio effector que neste e na maioria dos casos consiste num domínio de ligação ao DNA (DBD). Após a fosforilação do regulador de resposta, a proteína sofre uma alteração conformacional, dimerizando e aumentando subsequente a sua afinidade de ligação a uma região específica do DNA. O estudo centra-se no TCS PhoR-PhoB, que é activado após uma limitação da concentração de fosfato no meio. A proteína PhoR activa a proteína PhoB através da tranferência do grupo fosfato com consequente aumento da afinidade de última proteína para uma região específica do DNA (Pho box), regulando a transcrição dos genes do regulão Pho. O sistema Pho tem sido alvo de numerosos estudos em diversas bactérias. No entanto, a informação relativa ao sistema em D. radiodurans resume-se apenas à anotação dos genes codificantes para proteínas PhoR (DR2244) e PhoB (DR2245). Nesta tese, é apresentada a caracterização estrutural das proteínas em solução na sua forma integral bem como dos respectivos domínios; utilizando a técnica de difracção de raios-X de pequenos ângulos (SAXS). O estudo é complementado com análises bioquímicas e determinação de estrutura cristalografica do domínio PhoRCAD (resolução de 2.25 Å). Os resultados bioquímicos demonstram a autofosforilação in vitro da proteína PhoR, bem como a sua capacidade para transferir o grupo fosfato para o domínio PhoBRD.
European Synchrotron Radiation Facility (ESRF)
48
« Proteomic, Genetic, and Biochemical Analyses of Two-Component Regulatory Systems in Porphyromonas gingivalis and Escherichia coli ». Master's thesis, 2013. http://hdl.handle.net/2286/R.I.20978.
Texte intégralRésumé :
abstract: Pathogenic Gram-negative bacteria employ a variety of molecular mechanisms to combat host defenses. Two-component regulatory systems (TCR systems) are the most ubiquitous signal transduction systems which regulate many genes required for virulence and survival of bacteria. In this study, I analyzed different TCR systems in two clinically-relevant Gram-negative bacteria, i.e., oral pathogen Porphyromonas gingivalis and enterobacterial Escherichia coli. P. gingivalis is a major causative agent of periodontal disease as well as systemic illnesses, like cardiovascular disease. A microarray study found that the putative PorY-PorX TCR system controls the secretion and maturation of virulence factors, as well as loci involved in the PorSS secretion system, which secretes proteinases, i.e., gingipains, responsible for periodontal disease. Proteomic analysis (SILAC) was used to improve the microarray data, reverse-transcription PCR to verify the proteomic data, and primer extension assay to determine the promoter regions of specific PorX regulated loci. I was able to characterize multiple genetic loci regulated by this TCR system, many of which play an essential role in hemagglutination and host-cell adhesion, and likely contribute to virulence in this bacterium. Enteric Gram-negative bacteria must withstand many host defenses such as digestive enzymes, low pH, and antimicrobial peptides (AMPs). The CpxR-CpxA TCR system of E. coli has been extensively characterized and shown to be required for protection against AMPs. Most recently, this TCR system has been shown to up-regulate the rfe-rff operon which encodes genes involved in the production of enterobacterial common antigen (ECA), and confers protection against a variety of AMPs. In this study, I utilized primer extension and DNase I footprinting to determine how CpxR regulates the ECA operon. My findings suggest that CpxR modulates transcription by directly binding to the rfe promoter. Multiple genetic and biochemical approaches were used to demonstrate that specific TCR systems contribute to regulation of virulence factors and resistance to host defenses in P. gingivalis and E. coli, respectively. Understanding these genetic circuits provides insight into strategies for pathogenesis and resistance to host defenses in Gram negative bacterial pathogens. Finally, these data provide compelling potential molecular targets for therapeutics to treat P. gingivalis and E. coli infections.Dissertation/Thesis
M.S. Biology 2013
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Erraguntla, Mythili. « Genetic Analysis And Biochemical Activities Of β Protein : A Component Of Bacteriophage λ General Genetic Recombination ». Thesis, 1995. https://etd.iisc.ac.in/handle/2005/1906.
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Erraguntla, Mythili. « Genetic Analysis And Biochemical Activities Of β Protein : A Component Of Bacteriophage λ General Genetic Recombination ». Thesis, 1995. http://etd.iisc.ernet.in/handle/2005/1906.
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