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- Gururajan, Vasudevan Thanvanthri, et al.
(författare)
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Development and characterisation of a recombinant Saccharomyces cerevisiae mutant strain with enhanced xylose fermentation properties
- 2007
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Ingår i: Annals of Microbiology. - 1590-4261. ; 57:4, s. 599-607
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Tidskriftsartikel (refereegranskat)abstract
- The purpose of this study was to help lay the foundation for further development of xylose-fermenting Saccharomyces cerevisiae yeast strains through an approach that combined metabolic engineering and random mutagenesis in a recombinant haploid strain that overexpressed only two genes of the xylose pathway. Previously, S. cerevisiae strains, overexpressing heterologous genes encoding xylose reductase, xylitol dehydrogenase and the endogenous XKS1 xylulokinase gene, were randomly mutagenised to develop improved xylose-fermenting strains. In this study, two gene cassettes (ADH1(p)-PsXYL1-ADH1(T) and PGK1(p)-PsXYL2-PGK1(T)) containing the xylose reductase (PsXYL1) and xylitol dehydrogenase (PsXYL2) genes from the xylose-fermenting yeast, Pichia stipitis, were integrated into the genome of a haploid S. cerevisiae strain (CEN.PK 2-1D). The resulting recombinant strain (YUSM 1001) overexpressing the P. stipitis XYL1 and XYL2 genes (but not the endogenous XKS1 gene) was subjected to ethyl methane sulfonate (EMS) mutagenesis. The resulting mutants were screened for faster growth rates on an agar medium containing xylose as the sole carbon source. A mutant strain (designated Y-X) that showed 20-fold faster growth in xylose medium in shake-flask cultures was isolated and characterised. In anaerobic batch fermentation, the Y-X mutant strain consumed 2.5-times more xylose than the YUSM 1001 parental strain and also produced more ethanol and glycerol. The xylitol yield from the mutant strain was lower than that from the parental strain, which did not produce glycerol and ethanol from xylose. The mutant also showed a 50% reduction in glucose consumption rate. Transcript levels of XYL1, XYL2 and XKS1 and the GPD2 glycerol 3-phosphate dehydrogenase gene from the two strains were compared with real-time reverse transcription polymerase chain reaction (RT-PCR) analysis. The mutant showed 10-40 times higher relative expression of these four genes, which corresponded with either the higher activities of their encoded enzymes or by-product formation during fermentation. Furthermore, no mutations were observed in the mutant's promoter sequences or the open reading frames of some of its key genes involved in carbon catabolite repression, glycerol production and redox balancing. The data suggest that the enhancement of the xylose fermentation properties of the Y-X mutant was made possible by increased expression of the xylose pathway genes, especially the XKS1 xylulokinase gene.
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- Shah, Nihir, et al.
(författare)
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Determination of an antimicrobial activity of Weissella confusa, Lactobacillus fermentum, and Lactobacillus plantarum against clinical pathogenic strains of Escherichia coli and Staphylococcus aureus in co-culture
- 2016
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Ingår i: Annals of Microbiology. - : Springer Science and Business Media LLC. - 1590-4261 .- 1869-2044. ; 66:3, s. 1137-1143
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Tidskriftsartikel (refereegranskat)abstract
- Lactic acid bacteria (LAB) have long been used to produce safe and high quality products as they are potential producers of a wide range of antimicrobial compounds that exert either narrow or wide spectrum antimicrobial activity towards spoilage or disease-causing organisms. The present investigation aimed to study the antimicrobial effect of three LAB strains, viz., Lactobacillus plantarum (86), Lactobacillus fermentum (AI2) and Weissella confusa (AI10), against two clinical pathogenic strains viz., Escherichia coli NG 502121 and Staphylococcus aureus AY 507047 in co-culture. Effects of change in inoculum size, and growth measurement at different time intervals were also studied. The pH and viable count were measured for initial as well as 24 h incubated samples. A significant (P < 0.05) reduction (2–3 log cycles) in growth of both pathogens while co-cultured with LAB strains was observed. The nonsignificant (P < 0.05) pH difference revealed the action of other metabolites apart from organic acids. LAB strains overruled the growth of E. coli and S. aureus within 10 and 6 h of the initial growth stage, respectively, compared to controls. These results led us to further characterize and purify the antimicrobial compound produced by the studied strains, so that they can be exploited in the production of safe foods with longer shelf life.
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- Thanvanthri, Vasu, et al.
(författare)
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A constitutive catabolite repression mutant of a recombinant Saccharomyces cerevisiae strain improves xylose consumption during fermentation
- 2007
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Ingår i: Annals of Microbiology. - 1590-4261. ; 57:1, s. 85-92
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Tidskriftsartikel (refereegranskat)abstract
- Efficient xylose utilisation by microorganisms is of importance to the lignocellulose fermentation industry. The aim of this work was to develop constitutive catabolite repression mutants in a xylose-utilising recombinant Saccharomyces cerevisiae strain and evaluate the differences in xylose consumption under fermentation conditions. S. cerevisiae YUSM was constitutively catabolite repressed through specific disruptions within the MIG1 gene. The strains were grown aerobically in synthetic complete medium with xylose as the sole carbon source. Constitutive catabolite repressed strain YCR17 grew four-fold better on xylose in aerobic conditions than the control strain YUSM. Anaerobic batch fermentation in minimal medium with glucose-xylose mixtures and N-limited chemostats with varying sugar concentrations were performed. Sugar utilisation and metabolite production during fermentation were monitored. YCR17 exhibited a faster xylose consumption rate than YUSM under high glucose conditions in nitrogen-limited chemostat cultivations. This study shows that a constitutive catabolite repressed mutant could be used to enhance the xylose consumption rate even in the presence of high glucose in the fermentation medium. This could help in reducing fermentation time and cost in mixed sugar fermentation.
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