Relevant bibliographies by topics / Sigme proteins

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Academic literature on the topic 'Sigme proteins'

Author: Grafiati
Published: 6 September 2023

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Journal articles on the topic "Sigme proteins"

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Raman, Sahadevan, Xiaoling Puyang, Tan-Yun Cheng, David C. Young, D. Branch Moody, and Robert N. Husson. "Mycobacterium tuberculosis SigM Positively Regulates Esx Secreted Protein and Nonribosomal Peptide Synthetase Genes and Down Regulates Virulence-Associated Surface Lipid Synthesis." Journal of Bacteriology 188, no. 24 (October 6, 2006): 8460–68. http://dx.doi.org/10.1128/jb.01212-06.

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ABSTRACT The Mycobacterium tuberculosis genome encodes 12 alternative sigma factors, several of which regulate stress responses and are required for virulence in animal models of acute infection. In this work we investigated M. tuberculosis SigM, a member of the extracytoplasmic function subfamily of alternative sigma factors. This sigma factor is expressed at low levels in vitro and does not appear to function in stress response regulation. Instead, SigM positively regulates genes required for the synthesis of surface or secreted molecules. Among these are genes encoding two pairs of Esx secreted proteins, a multisubunit nonribosomal peptide synthetase operon, and genes encoding two members of the proline-proline-glutamate (PPE) family of proteins. Genes up regulated in a sigM mutant strain include a different PPE gene, as well as several genes involved in surface lipid synthesis. Among these are genes involved in synthesis of phthiocerol dimycocerosate (PDIM), a surface lipid critical for virulence during acute infection, and the kasA-kasB operon, which is required for mycolic acid synthesis. Analysis of surface lipids showed that PDIM synthesis is increased in a sigM-disrupted strain and is undetectable in a sigM overexpression strain. These findings demonstrate that SigM positively and negatively regulates cell surface and secreted molecules that are likely to function in host-pathogen interactions.
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Nakunst, Diana, Christof Larisch, Andrea T. Hüser, Andreas Tauch, Alfred Pühler, and Jörn Kalinowski. "The Extracytoplasmic Function-Type Sigma Factor SigM of Corynebacterium glutamicum ATCC 13032 Is Involved in Transcription of Disulfide Stress-Related Genes." Journal of Bacteriology 189, no. 13 (May 4, 2007): 4696–707. http://dx.doi.org/10.1128/jb.00382-07.

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ABSTRACT The gene for the extracytoplasmic function (ECF) sigma factor SigM was deleted from the chromosome of the gram-positive soil bacterium Corynebacterium glutamicum to elucidate the role of the SigM protein in the regulation of gene expression. Comparative DNA microarray hybridizations of the C. glutamicum wild type and sigM-deficient mutant C. glutamicum DN1 revealed 23 genes with enhanced expression in the sigM-proficient strain, encoding functions in the assembly of iron-sulfur clusters (suf operon), thioredoxin reductase (trxB), thioredoxins (trxC, trxB1), chaperones (groES, groEL, clpB), and proteins involved in the heat shock response (hspR, dnaJ, grpE). Deletion of the sigM gene rendered the C. glutamicum cells more sensitive to heat, cold, and the presence of the thiol oxidant diamide. Transcription of the sigM gene increased under different stress conditions, including heat shock, cold shock, and disulfide stress caused by diamide treatment, suggesting a regulatory role for SigM under thiol-oxidative stress conditions. Stress-responsive promoters were determined upstream of the suf operon and of the trxB, trxC, and trxB1 genes. The deduced SigM consensus promoter is characterized by the −35 hexamer gGGAAT and the −10 hexamer YGTTGR. Transcription of the sigM gene is apparently controlled by the ECF sigma factor SigH, since a sigH mutant was unable to enhance the expression of sigM and the SigM regulon under thiol-oxidative stress conditions. A typical SigH-responsive promoter was mapped upstream of the sigM gene. The ECF sigma factor SigM is apparently part of a regulatory cascade, and its transcription is controlled by SigH under conditions of thiol-oxidative stress.
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Fernandes, Norvin D., Qi-long Wu, Dequan Kong, Xiaoling Puyang, Sumeet Garg, and Robert N. Husson. "A Mycobacterial Extracytoplasmic Sigma Factor Involved in Survival following Heat Shock and Oxidative Stress." Journal of Bacteriology 181, no. 14 (July 15, 1999): 4266–74. http://dx.doi.org/10.1128/jb.181.14.4266-4274.1999.

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ABSTRACT Extracytoplasmic function (ECF) sigma factors are a heterogeneous group of alternative sigma factors that regulate gene expression in response to a variety of conditions, including stress. We previously characterized a mycobacterial ECF sigma factor, SigE, that contributes to survival following several distinct stresses. A gene encoding a closely related sigma factor, sigH, was cloned fromMycobacterium tuberculosis and Mycobacterium smegmatis. A single copy of this gene is present in these and other fast- and slow-growing mycobacteria, including M. fortuitum and M. avium. While the M. tuberculosis and M. smegmatis sigH genes encode highly similar proteins, there are multiple differences in adjacent genes. The single in vivo transcriptional start site identified inM. smegmatis and one of two identified in M. bovis BCG were found to have −35 promoter sequences that match the ECF-dependent −35 promoter consensus. Expression from these promoters was strongly induced by 50°C heat shock. In comparison to the wild type, an M. smegmatis sigH mutant was found to be more susceptible to cumene hydroperoxide stress but to be similar in logarithmic growth, stationary-phase survival, and survival following several other stresses. Survival of an M. smegmatis sigH sigE double mutant was found to be markedly decreased following 53°C heat shock and following exposure to cumene hydroperoxide. Expression of the second gene in the sigH operon is required for complementation of the sigH stress phenotypes. SigH is an alternative sigma factor that plays a role in the mycobacterial stress response.
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Singh, Rakesh Kumar, Lav Kumar Jaiswal, Tanmayee Nayak, Ravindra Singh Rawat, Sanjit Kumar, Sachchida Nand Rai, and Ankush Gupta. "Expression, Purification, and In Silico Characterization of Mycobacterium smegmatis Alternative Sigma Factor SigB." Disease Markers 2022 (May 20, 2022): 1–11. http://dx.doi.org/10.1155/2022/7475704.

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Sigma factor B (SigB), an alternative sigma factor (ASF), is very similar to primary sigma factor SigA (σ70) but dispensable for growth in both Mycobacterium smegmatis (Msmeg) and Mycobacterium tuberculosis (Mtb). It is involved in general stress responses including heat, oxidative, surface, starvation stress, and macrophage infections. Despite having an extremely short half-life, SigB tends to operate downstream of at least three stress-responsive extra cytoplasmic function (ECF) sigma factors (SigH, SigE, SigL) and SigF involved in multiple signaling pathways. There is very little information available regarding the regulation of SigB sigma factor and its interacting protein partners. Hence, we cloned the SigB gene into pET28a vector and optimized its expression in three different strains of E. coli, viz., (BL21 (DE3), C41 (DE3), and CodonPlus (DE3)). We also optimized several other parameters for the expression of recombinant SigB including IPTG concentration, temperature, and time duration. We achieved the maximum expression of SigB at 25°C in the soluble fraction of the cell which was purified by affinity chromatography using Ni-NTA and further confirmed by Western blotting. Further, structural characterization demonstrates the instability of SigB in comparison to SigA that is carried out using homology modeling and structure function relationship. We have done protein-protein docking of RNA polymerase (RNAP) of Msmeg and SigB. This effort provides a platform for pulldown assay, structural, and other studies with the recombinant protein to deduce the SigB interacting proteins, which might pave the way to study its signaling networks along with its regulation.
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White, Mark J., Hongjun He, Renee M. Penoske, Sally S. Twining, and Thomas C. Zahrt. "PepD Participates in the Mycobacterial Stress Response Mediated through MprAB and SigE." Journal of Bacteriology 192, no. 6 (January 8, 2010): 1498–510. http://dx.doi.org/10.1128/jb.01167-09.

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ABSTRACT Currently, one-third of the world's population is believed to be latently infected with Mycobacterium tuberculosis. The mechanisms by which M. tuberculosis establishes latent infection remain largely undefined. mprAB encodes a two-component signal transduction system required by M. tuberculosis for aspects of persistent infection. MprAB regulates a large and diverse group of genetic determinants in response to membrane stress, including the extracytoplasmic function (ECF) sigma factor sigE and the HtrA-like serine protease pepD. Recent studies have demonstrated that PepD functions as both a protease and chaperone in vitro. In addition, inactivation of pepD alters the virulence of M. tuberculosis in a mouse model system of infection. Here, we demonstrate that PepD plays an important role in the stress response network of Mycobacterium mediated through MprAB and SigE. In particular, we demonstrate that the protease activity of PepD requires the PDZ domain, in addition to the catalytic serine at position 317. pepD expression initiates from at least three promoters in M. tuberculosis, including one that is regulated by SigE and is located upstream of the mprA coding sequence. Deletion of pepD or mprAB in Mycobacterium smegmatis and M. tuberculosis alters the stress response phenotypes of these strains, including increasing sensitivity to SDS and cell wall antibiotics and upregulating the expression of stress-responsive determinants, including sigE. Taking these data together, we hypothesize that PepD utilizes its PDZ domain to recognize and process misfolded proteins at the cell membrane, leading to activation of the MprAB and SigE signaling pathways and subsequent establishment of a positive feedback loop that facilitates bacterial adaptation.
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Kim, Eun Sook, Ju Yeon Song, Dae Wi Kim, Keith F. Chater, and Kye Joon Lee. "A Possible Extended Family of Regulators of Sigma Factor Activity in Streptomyces coelicolor." Journal of Bacteriology 190, no. 22 (September 12, 2008): 7559–66. http://dx.doi.org/10.1128/jb.00470-08.

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ABSTRACT SCO4677 is one of a large number of similar genes in Streptomyces coelicolor that encode proteins with an HATPase_c domain resembling that of anti-sigma factors such as SpoIIAB of Bacillus subtilis. However, SCO4677 is not located close to genes likely to encode a cognate sigma or anti-anti-sigma factor. SCO4677 was found to regulate antibiotic production and morphological differentiation, both of which were significantly enhanced by the deletion of SCO4677. Through protein-protein interaction screening of candidate sigma factor partners using the yeast two-hybrid system, SCO4677 protein was found to interact with the developmentally specific σF, suggesting that it is an antagonistic regulator of σF. Two other proteins, encoded by SCO0781 and SCO0869, were found to interact with the SCO4677 anti-σF during a subsequent global yeast two-hybrid screen, and the SCO0869-SCO4677 protein-protein interaction was confirmed by coimmunoprecipitation. The SCO0781 and SCO0869 proteins resemble well-known anti-anti-sigma factors such as SpoIIAA of B. subtilis. It appears that streptomycetes may possess an extraordinary abundance of anti-sigma factors, some of which may influence diverse processes through interactions with multiple partners: a novel feature for such regulatory proteins.
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Kazmierczak, Mark J., Martin Wiedmann, and Kathryn J. Boor. "Alternative Sigma Factors and Their Roles in Bacterial Virulence." Microbiology and Molecular Biology Reviews 69, no. 4 (December 2005): 527–43. http://dx.doi.org/10.1128/mmbr.69.4.527-543.2005.

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SUMMARY Sigma factors provide promoter recognition specificity to RNA polymerase holoenzyme, contribute to DNA strand separation, and then dissociate from the core enzyme following transcription initiation. As the regulon of a single sigma factor can be composed of hundreds of genes, sigma factors can provide effective mechanisms for simultaneously regulating expression of large numbers of prokaryotic genes. One newly emerging field is identification of the specific roles of alternative sigma factors in regulating expression of virulence genes and virulence-associated genes in bacterial pathogens. Virulence genes encode proteins whose functions are essential for the bacterium to effectively establish an infection in a host organism. In contrast, virulence-associated genes can contribute to bacterial survival in the environment and therefore may enhance the capacity of the bacterium to spread to new individuals or to survive passage through a host organism. As alternative sigma factors have been shown to regulate expression of both virulence and virulence-associated genes, these proteins can contribute both directly and indirectly to bacterial virulence. Sigma factors are classified into two structurally unrelated families, the σ70 and the σ54 families. The σ70 family includes primary sigma factors (e.g., Bacillus subtilis σA) as well as related alternative sigma factors; σ54 forms a distinct subfamily of sigma factors referred to as σN in almost all species for which these proteins have been characterized to date. We present several examples of alternative sigma factors that have been shown to contribute to virulence in at least one organism. For each sigma factor, when applicable, examples are drawn from multiple species.
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S Thompson, L., and E. J Harry. "Alternative sigma factors: the master regulators." Microbiology Australia 27, no. 3 (2006): 118. http://dx.doi.org/10.1071/ma06118.

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When a bacterial cell encounters a change in environmental conditions, it responds by producing a different complement of cellular proteins. Which proteins are produced and maintained is regulated in a number of ways, including regulation of gene transcription, stabilising or degrading mRNA transcripts, post translational modifications and targeted degradation of proteins.
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Yang, Fumeng, Wenjun Wang, Qian Liu, Xizhen Wang, Guangrong Bian, Shijie Teng, and Wei Liang. "The application of Six Sigma to perform quality analyses of plasma proteins." Annals of Clinical Biochemistry: International Journal of Laboratory Medicine 57, no. 2 (December 4, 2019): 121–27. http://dx.doi.org/10.1177/0004563219892023.

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Background The Six Sigma theory is an important tool for laboratory quality management. It has been widely used in clinical chemistry, haematology and other disciplines. The aim of our study was to evaluate the analytical performance of plasma proteins by application of Sigma metric and to compare the differences among three different allowable total errors in evaluating the analytical performance of plasma proteins. Methods Three different allowable total error values were used as quality goals. Data from an external quality assessment were used as bias, and the cumulative coefficient of variation in internal quality control data was used to represent the amount of imprecision during the same period. Sigma metric of analytes was calculated using the above data. The quality goal index was calculated to provide corrected measures for continuous improvements in analytical quality. Results The Sigma metric was highest using the external quality assessment standards of China: it was sigma ≥6 or higher in 57.1% of plasma proteins. But Sigma metric was lower by using RiliBÄK or biological variation standards. IgG, C3 and C-reactive protein all required quality improvements in imprecision. A single-rule 13s for internal quality control was recommended for IgA, IgM, C4 and rheumatoid factor, whereas multiple rules (13s/22s/R4s) were recommended for IgG, C3 and C-reactive protein, according to the external quality assessment standards of China. Conclusions Different quality goals can lead to different Sigma metric for the same analyte. As the lowest acceptable standard in clinical practice, the external quality assessment standard of China can guide laboratories to formulate reasonable quality improvement programmes.
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Mortillaro, N. A., and A. E. Taylor. "Microvascular permeability to endogenous plasma proteins in the jejunum." American Journal of Physiology-Heart and Circulatory Physiology 258, no. 6 (June 1, 1990): H1650—H1654. http://dx.doi.org/10.1152/ajpheart.1990.258.6.h1650.

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Steady-state lymph flow and lymph (CL) and plasma (CP) protein concentrations were measured at venous outflow pressures of 0, 10, 20, and 30 mmHg in an autoperfused segment of cat jejunum. In addition to determining total protein concentrations in lymph and plasma, polyacrylamide gradient gel electrophoresis was used to determine lymph and plasma protein concentrations of albumin and nine other plasma proteins. The osmotic reflection coefficient (sigma d) for total proteins, albumin, and each of the nine protein fractions was estimated using CL/CP at a capillary filtration rate independent state, when 1 - CL/CP = sigma d. A sigma d of 0.83 was obtained for total proteins, a value similar to that reported for both dog jejunum and descending colon (0.85) but appreciably different from that reported for both cat stomach (0.78) and ileum (0.92). Additionally, sigma d for albumin and each of the nine plasma protein fractions (molecular radii ranging from 37 to 120 A) increased as molecular radius increased. A two-pore model composed of a ratio of 3,750 48-A radius small pores to one 250-A radius large pore describes the data obtained, with 82% of the total volume flow occurring through the small pores and 16% through the large pores.
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Dissertations / Theses on the topic "Sigme proteins"

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Sharma, Khushbu. "Studying sigma family of transcription factors in mycobacteria: In-silico and functional analyses of conserved sigma proteins of mip by in-vitro and in-vivo methods." Thesis, IIT Delhi, 2019. http://eprint.iitd.ac.in:80//handle/2074/8065.

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Lee, S. F. K. "Understanding protein tyrosine phosphatase sigma function : dimer formation and interacting proteins." Thesis, University College London (University of London), 2007. http://discovery.ucl.ac.uk/1444958/.

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Cytoplasmic and transmembrane protein tyrosine phosphatases (PTPs) provide the enzymatic counterbalance to protein tyrosine kinase activity. PTP sigma (PTPa) is an adhesion molecule-like receptor PTP (RPTP) that is expressed on the growth cones of developing axons. PTPa binds ligands located in the basement membrane (heparan sulphate proteoglycans: agrin and collagen 18) and developing muscle (Nucleolin). Disruption of ligand-PTPa interactions affects axon guidance, although neither the role of PTPa in neurons nor the effect of ligand binding on PTPa activity is well understood. Further characterisation of PTPa function remains difficult in the absence of an understanding of PTPa biochemistry that would allow a functional assay to measure the effects of an experimental manipulation on PTPa function. PTPa was shown to be dimeric using a combination of disulphide cross-linking and co-immunoprecipitation techniques. This dimeric form of PTPa was principally cell surface localised according to its accessibility to trypsinisation. However, neither co-immunoprecipitation nor glutathione S-transferase (GST) pull-down techniques allowed the identification of proteins that interact with wild type or recombinant substrate-trapping PTPa. Secondly, H202 treatment of PTPa-expressing cells induced the formation of reduction-sensitive, high molecular weight species. In contrast to the formation of PTPa dimers under the same conditions, this did not require the tyrosine phosphatase domain catalytic site cysteines. It may be possible to utilise the formation of high molecular weight species on non-reducing SDS-PAGE analysis as a proxy measure of PTPa oligomerisation. Moreover, unlike co-immunoprecipitation it can be used on wild type and even endogenously expressed proteins. Finally, the type Ila RPTP family consists of PTPa, PTPS and LAR. To allow the simultaneous disruption of type Ila RPTPs, chicken LAR was cloned from embryonic chick whole body mRNA and characterised.
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Donà, Valentina. "Caratterizzazione dei fattori sigma micobatterici SigE e SigF Caratterizzazione del dominio PPE della proteina PPE17 di Mycobacterium tuberculosis." Doctoral thesis, Università degli studi di Padova, 2010. http://hdl.handle.net/11577/3422362.

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Abstract Mycobacterium tuberculosis (MTB) is the causative of tuberculosis, a disease, which causes 2 millions of death every year, with a dramatic incidence especially in developing countries. To find new drug and vaccine strategies against MTB, it is of fundamental importance to study the mechanisms, that allow its survival to environmental stresses, to which it is subjected during the period of infection and latency in the host macrophages. The fine transcriptional regulation of specific genes in response to stress conditions and the peculiar structure of its wall play a key role on this. In the first part of the PhD project two mycobacterial sigma factors, SigE and SigF, which regulate the transcription of specific genes in response to various environmental stresses such as surface stress, oxidative stress, alkaline pH and thermal shock, have been characterized. First the transcriptional regulation, translational and post-translational regulation of the extracytoplasmic function (ECF) transcription factor SigE were studied. Regarding the study of the transcriptional regulation, it was possible to confirm by 5'RACE PCR and RT-PCR experiments the presence of three promoters of sigE, and to determine the contribution of each promoter in the transcription of this gene, depending on the environmental conditions of bacterial growth. The fact, that the transcriptional start codon of one of these promoters is located 63 base pairs downstream of the start codon annotated in MTB genome opened the possibility of the existence of two isoforms of Sige. By translational fusions between specific sequences of sigE with lacZ, deprived of its own translational initiation codon, and subsequent site-specific mutagenesis, it was possible to confirm, based on further beta-galactosidase activity detection, the existence of two alternative start codons, an ATC and a TTG, coding for an isoform of respectively 218 and 215 of amino acids, in addition to the ATG already annotated in MTB genome, which encodes for an isoform of 257 amino acids. Finally, it was possible to confirm, that the gene downstream sigE encodes for the anti-sigma factor of SigE, called RseA, capable of binding both isoforms of SigE. In a second project also the role of the factor SigF M. smegmatis in the biosynthesis of carotenoid pigments, resistance to hydrogen peroxide and in the efficiency of bacterial transformation was studied. By RT-PCR it has been shown, that SigF controls the transcription of genes involved in the biosynthesis of carotenoid pigments, and, assuming that they serve as protection against free radicals, it was verified that the sigF mutant strain is actually more sensitive compared to the wild type strain to treatment with hydrogen peroxide. Finally, we also demonstrated, that the mutant strain has a higher transformation efficiency than the wild type strain, indicating that SigF regulates the transcription of genes possibly involved in the permeability of the cell wall. In the second part of the project, the localization of the protein on the surface PPE17 mycobacteria was characterized. Like other members of the PPE family, the PPE17 has a highly conserved N-terminal domain, which, based on different evidences in literature, is assumed to play an important role in their translocation to the mycobacterial surface. Moreover, it was investigated the possible influence of the presence of PE11 in the translocation process or in the stability of PPE17, as the PE11 coding sequence is in tandem and co-transcribed with that encoding the PPE17, and there is a specific interaction between these two proteins. The data obtained by proteinase K sensitivity assays performed on M. smegmatis strains, expressing the entire PPE17 or only its domain PPE (dPPE17) fused with the HA epitope, confirm, that the entire PPE17 is exposed on the surface, both in the presence and absence of PE11. According to data obtained, the possibility to translocate the MTB model antigen (Mpt64) on the surface of the vaccine strain M. bovis BCG, by fusing them with the dPPE17 was tested. Proteinase K and whole cell ELISA assays performed on cultures of M. bovis BCG expressing this chimeric protein indicate, that it is indeed localized at the mycobacterial surface. Similarly, another two fusions with dPPE17 were constructed to express on the mycobacterial surface the multimeric MTB antigen AG85-ESAT6 of MTB and the Csp C3 antigen of Plasmodium bergii. According to the proteinase K sensitivity assays carried out on strains of M. smegmatis expressing the two chimeric proteins indicate that also in this case both are localized at the surface. The strains of M. bovis BCG expressing these antigens on their surface will be tested in future in the mouse model to measure any increase in protection compared to the wild type strain.
Riassunto Mycobacterium tuberculosis (MTB) è l’agente eziologico della tubercolosi, patologia che nel mondo causa ogni anno due milioni di morti, con un’incidenza drammatica specie nei Paesi in via di sviluppo. Per poter trovare nuove strategie farmacologiche e vaccinali contro MTB è di fondamentale importanza lo studio dei meccanismi, che permettono la sua sopravvivenza ai vari stress ambientali, ai quali è sottoposto durante il periodo di infezione e latenza nei macrofagi dell’ospite. La fine regolazione della trascrizione di geni specifici in risposta a condizioni di stress e la peculiare struttura della sua parete giocano in merito un ruolo fondamentale. Nella prima parte del progetto di dottorato sono stati caratterizzati due fattori di trascrizione sigma micobatterici, SigE e SigF, che regolano la trascrizione di geni specifici in risposta a vari tipi di stress ambientali, come lo stress di superficie, lo stress ossidativo, il pH alcalino e lo shock termico. Anzitutto è stata studiata la regolazione trascrizionale, traduzionale e posttraduzionale del fattore di trascrizione con funzione extracitoplasmatica (ECF) SigE. Per quanto riguarda lo studio della regolazione trascrizionale, è stato possibile confermare tramite esperimenti di 5’RACE PCR e RT-PCR la presenza di tre promotori di sigE, e a dosare, a seconda delle condizioni ambientali di crescita batterica, il contributo di ciascun promotore nella trascrizione di questo gene. Dato che l’inizio della trascrizione di uno di questi promotori è sito 63 paia di basi a valle del codone di start annotato nel genoma, si è aperta l’ipotesi dell’esistenza di due isoforme di SigE. Mediante fusioni traduzionali tra specifiche sequenze di sigE con lacZ, private del proprio codone di inizio della traduzione, e successive mutagenesi sito-specifiche, è stato possibile confermare, in base all’attività beta-galattosidasica rilevata, l’esistenza di due codoni di start alternativi, un ATC ed un TTG, che codificano per un’isoforma di rispettivamente 218 e 215 di amminoacidi, oltre all’ATG già annotato nel genoma di MTB, che codifica per un’isoforma di 257 amminoacidi. Infine è stato possibile confermare, che il gene a valle di sigE codifica per il fattore anti-sigma di SigE, denominato RseA, in grado di legare entrambe le isoforme di SigE. In un secondo progetto è stato studiato anche il ruolo del fattore SigF di M. smegmatis nella biosintesi di pigmenti carotenoidi, nella resistenza a perossido d’idrogeno e nell’efficienza di trasformazione batterica. Tramite RT-PCR è stato dimostrato che SigF controlla la trascrizione di geni coinvolti nella biosintesi dei pigmenti carotenoidi, e, partendo dal presupposto che essi fungono da protezione contro i radicali liberi, è stato verificato che il mutante per il gene sigF è effettivamente più sensibile rispetto al ceppo selvatico al trattamento con perossido d’idrogeno. Infine è stato dimostrato anche, che il ceppo mutante possiede una maggiore efficienza di trasformazione rispetto al ceppo selvatico, indicando che SigF regola probabilmente la trascrizione di geni coinvolti nella permeabilità della parete. Nella seconda parte del progetto è stata caratterizzata la localizzazione della proteina PPE17 sulla superficie micobatterica. Come altri membri della famiglia PPE, la PPE17 presenta un dominio N-terminale altamente conservato, il quale, in base a diverse evidenze in letteratura, si suppone svolgere un ruolo importante per la loro traslocazione in superficie. Inoltre, si è voluto verificare un’eventuale influenza della presenza della proteina PE11 nel processo di traslocazione in o nella stabilità della PPE17, in quanto la sequenza codificante la PE11 è in tandem e co-trascritta con quella codificante la PPE17, e vi è un’interazione specifica tra queste due proteine. I dati ottenuti mediante saggi di sensibilità alla proteinasi K su ceppi di M. smegmatis, esprimenti la PPE17 intera o solo il suo dominio PPE (dPPE17) fuse all’epitopo HA, confermano che la PPE17 intera sia esposta in superficie, sia in presenza che in assenza di PE11. In base ai dati ottenuti si è infine tentato di veicolare un antigene modello (Mpt64) di MTB sulla superficie del ceppo vaccinale M. bovis BCG fondendolo con il dPPE17. Saggi di sensibilità alla proteinasi K e ELISA su cellule intere effettuati su culture di M. bovis BCG esprimenti questa proteina chimerica indicano, che essa sia effettivamente localizzata a livello superficiale. Allo stesso modo sono state costruite due ulteriori fusioni con il dPPE17 per esprimere sulla superficie micobatterica l’antigene multimerico Ag85-ESAT6 di MTB e l’antigene Csp C3 di Plasmodium berghii. In base a saggi di sensibilità alla proteinasi K svolti su ceppi di M. smegmatis esprimenti le due fusioni anche in questo caso entrambe localizzano in superficie. I ceppi di M. bovis BCG esprimenti questi antigeni sulla loro superficie saranno testati in futuro nel modello del topo per misurare un eventuale aumento della protezione rispetto al ceppo selvatico.
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Morikawa, Kazuya. "Regulation of Plastid Gene Transcription by Sigma Factors and Sigma Factor Binding Proteins." Kyoto University, 2001. http://hdl.handle.net/2433/150743.

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Kyoto University (京都大学)
0048
新制・課程博士
博士(人間・環境学)
甲第9028号
人博第121号
12||123(吉田南総合図書館)
新制||人||30(附属図書館)
UT51-2001-F358
京都大学大学院人間・環境学研究科文化・地域環境学専攻
(主査)教授 豊島 喜則, 教授 藤堂 剛, 助教授 瀬戸口 浩彰
学位規則第4条第1項該当
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Chadsey, Meggen Shepherd. "Regulation of the flagellar specific sigma factor, sigma28, of Salmonella typhimurium by the anti-sigma factor FlgM /." Thesis, Connect to this title online; UW restricted, 1998. http://hdl.handle.net/1773/11490.

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Ruan, Qingguo. "Aire regulates central and peripheral tolerance through direct control of autoantigens and other key genes in thymus epithelial cells and dendritic cells." [Gainesville, Fla.] : University of Florida, 2004. http://purl.fcla.edu/fcla/etd/UFE0006464.

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Thesis (Ph.D.)--University of Florida, 2004.
Typescript. Title from title page of source document. Document formatted into pages; contains 100 pages. Includes Vita. Includes bibliographical references.
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Yeung, Shu-wai. "Analysis of 14-3-3 [sigma] protein in nasopharyngeal tissues." Click to view the E-thesis via HKUTO, 2003. http://sunzi.lib.hku.hk/hkuto/record/B31971398.

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Smillie, David Andrew. "Genes encoding sigma cross-reacting proteins of Escherichia coli." Thesis, University of Edinburgh, 1994. http://hdl.handle.net/1842/14435.

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In the course of work aimed at discovering new sigma transcription initiation factors in Escherichia coli, two unknown sigma cross-reacting proteins (SCRP-23 and SCRP-27A) were identified by cross-reaction with antibodies raised against region 2.2 of sigma 70. This thesis describes the mapping, sequencing and characterisation of the corresponding genes. The gene encoding SCRP-23 was located near 652kb on the physical map of the E. coli chromosomes. Its sequence and that of part of a downstream open reading frame were determined, and found to be closely similar to the ahpC and F genes (respectively) of Salmonella typhimurium. These encode the C22 and F52a subunits of the anti-oxidant enzyme, alkylhydroperoxide reductase. The identity of the E. coli genes was further established by their ability when introduced on plasmids into an ahp deletion strain to restore cumene hydroperoxide resistance. Transcription of ahpCF was found to be driven by two promoters: ahpP1 is dependent on activation by the OxyR transcriptional regulator, whilst ahpP2 is independent of this factor. Indeed ahpP2 is located within the OxyR target site, and is repressed when OxyR is activated by oxidation. The gene encoding SCRP-27A, scrP, was located near 3416 kb on the physical map, just 250 bp downstream of arcB. arcB encodes a transmembrane sensor-regulator of respiratory functions.
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楊舒瑋 and Shu-wai Yeung. "Analysis of 14-3-3 [sigma] protein in nasopharyngeal tissues." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2003. http://hub.hku.hk/bib/B31971398.

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Chagnon, Mélanie J. 1977. "Physiological and molecular functions of the murine receptor protein tyrosine phosphatase sigma (RPTP[sigma])." Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=115661.

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The control of cellular tyrosine phosphorylation levels is of great importance in many biological systems. Among the kinases and phosphatases that modulate these levels, the LAR-RPTPs have been suggested to act in several key aspects of neural development, and in a dysfunctional manner in various pathologies from diabetes to cancer. The aim of this thesis is to describe the physiological functions of one of the members of this subfamily of RPTPs, namely RPTPsigma. First, we showed that glucose homeostasis is altered in RPTPsigma null mice. They are hypoglycemic and more sensitive to exogenous insulin and we proposed that the insulin hypersensitivity observed in RPTPsigma-null mice is likely secondary to their neuroendocrine dysplasia and GH/IGF-1 deficiency. In addition to regulating nervous system development, RPTPsigma was previously shown to regulate axonal regeneration after injury. In the absence of RPTPsigma, axonal regeneration in the sciatic, facial and optical nerves was enhanced following nerve crush. However, myelin-associated growth inhibitory proteins and components of the glial scar such as CSPGs (chondroitin sulfate proteoglycans) have long been known to inhibit axonal regeneration in the CNS, making spinal cord injury irreversible. In collaboration with Dr Samuel David, we unveiled that RPTPsigma null mice are able to regenerate their corticospinal tract following spinal cord hemisections as opposed to their WT littermates. We then isolated primary neurons from both sets of animals and found that the absence of RPTPsigma promotes the ability of the neurons to adhere to certain inhibitory substrates. Finally, in order to better understand the physiological role of RPTPsigma, we used a yeast substrate-trapping approach, to screen a murine embryonic library for new substrates. This screen identified the RhoGAP p250GAP as a new substrate, suggesting a downstream role for RPTPsigma in RhoGTPase signaling. We also identified p130Cas and Fyn as new binding partners. All these proteins have clear functional links to neurite extension. The characterization of RPTPsigma and its signaling partners is essential for understanding its role in neurological development and may one day translate into treatments of neural diseases and injuries.
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Books on the topic "Sigme proteins"

1

Kim, Felix J., and Gavril W. Pasternak, eds. Sigma Proteins: Evolution of the Concept of Sigma Receptors. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/978-3-319-65853-7.

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Watling, Keith J. The Sigma-RBI handbook of receptor classification and signal transduction. 4th ed. Natick, MA: Sigma-RBI, 2001.

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Mason, S. Studies on the cell-binding properties of the reovirus [sigma]1 protein. [s.l.]: typescript, 1992.

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Pasternak, Gavril W., and Felix J. Kim. Sigma Proteins: Evolution of the Concept of Sigma Receptors. Springer, 2017.

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Pasternak, Gavril W., and Felix J. Kim. Sigma Proteins: Evolution of the Concept of Sigma Receptors. Springer, 2018.

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Batt, Jane. Characterization of the role of protein tyrosine phosphatase sigma (PTP[sigma]) in mammalian development. 2002.

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Farone, Mary Bosch. The use of reovirus type 3 protein sigma one in the therapy of murine EL4 lymphoma. 1993.

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Narlikar, A. V., and Y. Y. Fu, eds. Oxford Handbook of Nanoscience and Technology. Oxford University Press, 2017. http://dx.doi.org/10.1093/oxfordhb/9780199533060.001.0001.

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This volume highlights engineering and related developments in the field of nanoscience and technology, with a focus on frontal application areas like silicon nanotechnologies, spintronics, quantum dots, carbon nanotubes, and protein-based devices as well as various biomolecular, clinical and medical applications. Topics include: the role of computational sciences in Si nanotechnologies and devices; few-electron quantum-dot spintronics; spintronics with metallic nanowires; Si/SiGe heterostructures in nanoelectronics; nanoionics and its device applications; and molecular electronics based on self-assembled monolayers. The volume also explores the self-assembly strategy of nanomanufacturing of hybrid devices; templated carbon nanotubes and the use of their cavities for nanomaterial synthesis; nanocatalysis; bifunctional nanomaterials for the imaging and treatment of cancer; protein-based nanodevices; bioconjugated quantum dots for tumor molecular imaging and profiling; modulation design of plasmonics for diagnostic and drug screening; theory of hydrogen storage in nanoscale materials; nanolithography using molecular films and processing; and laser applications in nanotechnology. The volume concludes with an analysis of the various risks that arise when using nanomaterials.
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Placencia López, Bárbara Miladys, Fernanda Vanessa Alcívar Macías, José Aníbal Sánchez Saltos, Mayra Alejandra Cedeño Mera, Silvia Beatriz Alarcón Barreiro, María Gabriela Pertuz Alarcón, Jacqueline Beatriz Delgado Molina, José Roberto Rodríguez Mera, Agustina Elizabeth Cedeño Casanova, and Christian Paúl Vera Zambrano. Covid 19. Mawil Publicaciones de Ecuador, 2022, 2022. http://dx.doi.org/10.26820/978-9942-602-37-4.

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El siglo XXI se ha caracterizado desde sus inicios por una problemática de salud que ha afectado al mundo, que va desde un incremento de la resistencia microbiana, aumento de las enfermedades oncológicas hasta la aparición de nuevas enfermedades infecciosas emergentes y reemergentes, como ha sido la aparición de la COVID-19 a finales del pasado año. Los coronavirus son una extensa familia de virus que pueden causar enfermedades tanto en animales como en humanos. En los humanos, se sabe que varios coronavirus causan infecciones respiratorias que pueden ir desde el resfriado común hasta enfermedades más graves como el síndrome respiratorio de Oriente Medio (MERS) y el síndrome respiratorio agudo severo (SRAS). La COVID-19 (coronavirus disease 2019) también conocida como enfermedad por nuevo coronavirus es causada por el coronavirus del síndrome respiratorio agudo severo (SARS-CoV-2), su forma es redonda u ovalada y a menudo polimórfica, tiene un diámetro de 60 a 140 nm, la proteína espiga que se encuentra en la superficie del virus y forma una estructura en forma de barra, es la estructura principal utilizada para la tipificación, la proteína de la nucleocápside encapsula el genoma viral y puede usarse como antígeno de diagnóstico. Produce síntomas similares a los de la gripe, entre los que se incluyen fiebre, tos, disnea, mialgia y fatiga. También se ha observado la pérdida súbita del olfato y el gusto (sin que la mucosidad fuese la causa). En casos graves se caracteriza por producir neumonía, síndrome de dificultad respiratoria aguda, sepsis y choque séptico que conduce a alrededor del 3 % de los infectados a la muerte, aunque la tasa de mortalidad se encuentra en 4,48 % y sigue ascendiendo.
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Book chapters on the topic "Sigme proteins"

1

Kim, Felix J. "Introduction to Sigma Proteins: Evolution of the Concept of Sigma Receptors." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 1–11. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/164_2017_41.

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Katz, Jonathan L., Takato Hiranita, Weimin C. Hong, Martin O. Job, and Christopher R. McCurdy. "A Role for Sigma Receptors in Stimulant Self-Administration and Addiction." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 177–218. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/164_2016_94.

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Kruse, Andrew. "Structural Insights into Sigma1 Function." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 13–25. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/164_2016_95.

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Zeng, Chenbo, Elizabeth S. McDonald, and Robert H. Mach. "Molecular Probes for Imaging the Sigma-2 Receptor: In Vitro and In Vivo Imaging Studies." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 309–30. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/164_2016_96.

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Sabino, Valentina, and Pietro Cottone. "Sigma Receptors and Alcohol Use Disorders." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 219–36. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/164_2016_97.

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Maurice, Tangui, and Nino Goguadze. "Sigma-1 (σ1) Receptor in Memory and Neurodegenerative Diseases." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 81–108. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/164_2017_15.

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Weber, Frauke, and Bernhard Wünsch. "Medicinal Chemistry of σ1 Receptor Ligands: Pharmacophore Models, Synthesis, Structure Affinity Relationships, and Pharmacological Applications." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 51–79. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/164_2017_33.

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Pasternak, Gavril W. "Allosteric Modulation of Opioid G-Protein Coupled Receptors by Sigma1 Receptors." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 163–75. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/164_2017_34.

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Laurini, Erik, Domenico Marson, Maurizio Fermeglia, and Sabrina Pricl. "3D Homology Model of Sigma1 Receptor." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 27–50. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/164_2017_35.

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Kim, Felix J., and Christina M. Maher. "Sigma1 Pharmacology in the Context of Cancer." In Sigma Proteins: Evolution of the Concept of Sigma Receptors, 237–308. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/164_2017_38.

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Conference papers on the topic "Sigme proteins"

1

Simin, Faria Anjum, and Joseph Schober. "Subcellular Localization Analysis of Sigma 1 Receptor (S1R) and Binding Immunoglobulin Protein (BiP) for Identification of Antagonists versus Agonists." In ASPET 2023 Annual Meeting Abstracts. American Society for Pharmacology and Experimental Therapeutics, 2023. http://dx.doi.org/10.1124/jpet.122.176040.

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GOMES, ARTUR BRUNO SILVA, FELIPE JATOBÁ LEITE NONATO, JULIANA MATOS FERREIRA BERNARDO, and MARCOS REIS GONÇALVES. "BIOMARCADORES EMPREGADOS NA MONITORIZAÇÃO DA IMUNOTERAPIA PARA A RINITE ALÉRGICA." In II Congresso Brasileiro de Imunologia On-line. Revista Multidisciplinar em Saúde, 2022. http://dx.doi.org/10.51161/ii-conbrai/5097.

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Introdução: Imunoterapia com alérgenos é uma abordagem potencialmente curativa para a rinite alérgica. Assim, a investigação de marcadores moleculares é fundamental para aprimorar o manejo clínico. Objetivo: Avaliar o uso dos biomarcadores na monitorização da imunoterapia para rinite alérgica. Material e Método: Revisão integrativa realizada na base de dados Pubmed com estratégia de pesquisa: allergen immunotherapy, biomarker e rhinitis, combinados com o operador booleano AND. Aplicou-se filtro de 5 anos, artigos em inglês e português, em humanos, retornando 97 resultados dos quais se selecionaram 14. Como critérios de inclusão escolheram-se artigos que avaliavam biomarcadores, enquanto os de exclusão foram os que abordaram outras doenças. Resultados e Discussão: Biomarcadores mais utilizados são sIgE e IgG4 e apresentam-se elevados na resposta ao tratamento. Por sua vez, a IgG4 para Dermatophagoides aumenta 30 vezes mais na imunoterapia subcutânea em relação à sublingual. Na imunoterapia subcutânea IgA e IgG4 específicas salivares servem como indicadores da resposta terapêutica, juntamente com o aumento do Interferon γ, que é inversamente proporcional aos sintomas de rinite. Ademais, na mucosa nasal observou-se a expressão de CD137 em células TCD8+. Os novos biomarcadores Leucotrieno A4 Hidrolase e Molécula de Adesão de Células Leucocitárias Ativadas mostram-se potencialmente preditivos à resposta terapêutica, enquanto a Periostina apresenta acurácia superior ao sIgE. Observou-se aumento da tolerabilidade dos basófilos, associado à diminuição da gravidade dos sintomas, medida pelos marcadores proteína CD63 e ectoenzima CD203c. Por fim, a análise metaboloma-proteoma possibilita melhor compreensão da doença e viabiliza a construção de painel molecular para a monitorização do tratamento. Conclusão: Biomarcadores indicam resposta para imunoterapia específica na rinite alérgica, todavia, novos estudos são essenciais para consolidar e padronizar essa análise laboratorial.
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SeungSang, Ko, Ji Young Kim, Joon Jeong, Jong Eun Lee, Woo Ick Yang, Hy De Lee, and Woo Hee Jung. "Abstract A68: The role of estrogen‐responsive finger protein (Efp) and 14‐3‐3 sigma (σ) in breast carcinogenesis and their prognostic implication." In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics--Nov 15-19, 2009; Boston, MA. American Association for Cancer Research, 2009. http://dx.doi.org/10.1158/1535-7163.targ-09-a68.

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Reports on the topic "Sigme proteins"

1

Coplin, David, Isaac Barash, and Shulamit Manulis. Role of Proteins Secreted by the Hrp-Pathways of Erwinia stewartii and E. herbicola pv. gypsophilae in Eliciting Water-Soaking Symptoms and Initiating Galls. United States Department of Agriculture, June 2001. http://dx.doi.org/10.32747/2001.7580675.bard.

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Many bacterial pathogens of plants can inject pathogenicity proteins into host cells using a specialized type III secretion system encoded by hrpgenes. This system deliver effector proteins, into plant cells that function in both susceptible and resistant interactions. We have found that the virulence of Erwinia stewartii(Es; syn. Pantoea stewartii) and Erwinia herbicola pv. gypsophilae (Ehg, syn. Pantoea agglomerans), which cause Stewart's wilt of corn and galls on Gypsophila, respectively, depends on hrpgenes. The major objectives of this project were: To increase expression of hrpgenes in order to identify secreted proteins; to identify genes for proteins secreted by the type-III systems and determine if they are required for pathogenicity; and to determine if the secreted proteins can function within eukaryotic cells. We found that transcription of the hrp and effector genes in Es and Ehg is controlled by at least four genes that constitute a regulatory cascade. Environmental and/or physiological signaling appears to be mediated by the HrpX/HrpY two component system, with HrpX functioning as a sensor-kinase and HrpY as a response regulator. HrpYupregulateshrpS, which encodes a transcriptional enhancer. HrpS then activates hrpL, which encodes an alternate sigma factor that recognizes "hrp boxes". All of the regulatory genes are essential for pathogenicity, except HrpX, which appears only to be required for induction of the HR in tobacco by Es. In elucidating this regulatory pathway in both species, we made a number of significant new discoveries. HrpX is unusual for a sensor-kinase because it is cytoplasmic and contains PAS domains, which may sense the redox state of the bacterium. In Es, a novel methyl-accepting protein may function upstream of hrpY and repress hrp gene expression in planta. The esaIR quorum sensing system in Es represses hrp gene expression in Es in response to cell-density. We have discovered six new type III effector proteins in these species, one of which (DspE in Ehg and WtsE in Es) is common to both pathogens. In addition, Es wtsG, which is a homolog of an avrPpiB from P. syringae pv. pisi, and an Ehg ORF, which is a homolog of P. syringae pv. phaseolicola AvrPphD, were both demonstrated to encode virulence proteins. Two plasmidborne, Ehg Hop proteins, HsvG and PthG, are required for infection of gypsophilia, but interestingly, PthG also acts as an Avr elicitor in beets. Using a calmodulin-dependent adenylate cyclase (cyaA) reporter gene, we were successful in demonstrating that an HsvG-CyaA fusion protein can be transferred into human HeLa cells by the type-III system of enteropathogenic E. coli. This is a highly significant accomplishment because it is the first direct demonstration that an effector protein from a plant pathogenic bacterium is capable of being translocated into a eukaryotic cell by a type-III secretion system. Ehg is considered a limiting factor in Gypsophila production in Israel and Stewart’s Wilt is a serious disease in the Eastern and North Central USA, especially on sweet corn in epidemic years. We believe that our basic research on the characterization of type III virulence effectors should enable future identification of their receptors in plant cells. This may lead to novel approaches for genetically engineering resistant plants by modifying their receptors or inactivating effectors and thus blocking the induction of the susceptible response. Alternatively, hrp gene regulation might also provide a target for plant produced compounds that interfere with recognition of the host by the pathogen. Such strategies would be broadly applicable to a wide range of serious bacterial diseases on many crops throughout the USA and Israel.
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Shapira, Roni, Judith Grizzle, Nachman Paster, Mark Pines, and Chamindrani Mendis-Handagama. Novel Approach to Mycotoxin Detoxification in Farm Animals Using Probiotics Added to Feed Stuffs. United States Department of Agriculture, May 2010. http://dx.doi.org/10.32747/2010.7592115.bard.

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T-2 toxin, a toxic product belongs to the trichothecene mycotoxins, attracts major interest because of its severe detrimental effects on the health of human and farm animals. The occurrence of trichothecenes contamination is global and they are very resistant to physical or chemical detoxification techniques. Trichothecenes are absorbed in the small intestine into the blood stream. The hypothesis of this project was to develop a protecting system using probiotic bacteria that will express trichothecene 3-O-acetyltransferase (Tri101) that convert T-2 to a less toxic intermediate to reduce ingested levels in-situ. The major obstacle that we had faced during the project is the absence of stable and efficient expression vectors in probiotics. Most of the project period was invested to screen and isolate strong promoter to express high amounts of the detoxify enzyme on one hand and to stabilize the expression vector on the other hand. In order to estimate the detoxification capacity of the isolated promoters we had developed two very sensitive bioassays.The first system was based on Saccharomyces cerevisiae cells expressing the green fluorescent protein (GFP). Human liver cells proliferation was used as the second bioassay system.Using both systems we were able to prove actual detoxification on living cells by probiotic bacteria expressing Tri101. The first step was the isolation of already discovered strong promoters from lactic acid bacteria, cloning them downstream the Tri101 gene and transformed vectors to E. coli, a lactic acid bacteria strain Lactococcuslactis MG1363, and a probiotic strain of Lactobacillus casei. All plasmid constructs transformed to L. casei were unstable. The promoter designated lacA found to be the most efficient in reducing T-2 from the growth media of E. coli and L. lactis. A prompter library was generated from L. casei in order to isolate authentic probiotic promoters. Seven promoters were isolated, cloned downstream Tri101, transformed to bacteria and their detoxification capability was compared. One of those prompters, designated P201 showed a relatively high efficiency in detoxification. Sequence analysis of the promoter region of P201 and another promoter, P41, revealed the consensus region recognized by the sigma factor. We further attempted to isolate an inducible, strong promoter by comparing the protein profiles of L. casei grown in the presence of 0.3% bile salt (mimicking intestine conditions). Six spots that were consistently overexpressed in the presence of bile salts were isolated and identified. Their promoter reigns are now under investigation and characterization.
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Zhou, Ting, Roni Shapira, Peter Pauls, Nachman Paster, and Mark Pines. Biological Detoxification of the Mycotoxin Deoxynivalenol (DON) to Improve Safety of Animal Feed and Food. United States Department of Agriculture, July 2010. http://dx.doi.org/10.32747/2010.7613885.bard.

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The trichothecene deoxynivalenol (DON, vomitoxin), one of the most common mycotoxin contaminants of grains, is produced by members of the Fusarium genus. DON poses a health risk to consumers and impairs livestock performance because it causes feed refusal, nausea, vomiting, diarrhea, hemolytic effects and cellular injury. The occurrence of trichothecenes contamination is global and they are very resistant to physical or chemical detoxification techniques. Trichothecenes are absorbed in the small intestine into the blood stream. The overall objective of this project was to develop a protecting system using probiotic bacteria that will express trichothecene 3-O-acetyltransferase (Tri101) that convert T-2 to a less toxic intermediate to reduce ingested levels in-situ. The major obstacle that we had faced during the project is the absence of stable and efficient expression vectors in probiotics. Most of the project period was invested to screen and isolate strong promoter to express high amounts of the detoxify enzyme on one hand and to stabilize the expression vector on the other hand. In order to estimate the detoxification capacity of the isolated promoters we had developed two very sensitive bioassays.The first system was based on Saccharomyces cerevisiae cells expressing the green fluorescent protein (GFP). Human liver cells proliferation was used as the second bioassay system.Using both systems we were able to prove actual detoxification on living cells by probiotic bacteria expressing Tri101. The first step was the isolation of already discovered strong promoters from lactic acid bacteria, cloning them downstream the Tri101 gene and transformed vectors to E. coli, a lactic acid bacteria strain Lactococcuslactis MG1363, and a probiotic strain of Lactobacillus casei. All plasmid constructs transformed to L. casei were unstable. The promoter designated lacA found to be the most efficient in reducing T-2 from the growth media of E. coli and L. lactis. A prompter library was generated from L. casei in order to isolate authentic probiotic promoters. Seven promoters were isolated, cloned downstream Tri101, transformed to bacteria and their detoxification capability was compared. One of those prompters, designated P201 showed a relatively high efficiency in detoxification. Sequence analysis of the promoter region of P201 and another promoter, P41, revealed the consensus region recognized by the sigma factor. We further attempted to isolate an inducible, strong promoter by comparing the protein profiles of L. casei grown in the presence of 0.3% bile salt (mimicking intestine conditions). Six spots that were consistently overexpressed in the presence of bile salts were isolated and identified. Their promoter reigns are now under investigation and characterization.
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