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Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis

A recombinant Saccharomyces cerevisiae strain transformed with xylose reductase (XR) and xylitol dehydrogenase (XDH) genes from Pichia stipitis (PsXR and PsXDH, respectively) has the ability to convert xylose to ethanol together with the unfavourable excretion of xylitol, which may be due to interc...

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Main Authors: Seiya, Watanabe, Abu Saleh, Ahmed, Narayana, Visu Annaluru
Format: Article
Language:English
Published: CSIRO Publishing 2007
Subjects:
TP Chemical technology
Online Access:http://ir.unimas.my/id/eprint/13992/1/Seiya.pdf
http://ir.unimas.my/id/eprint/13992/
https://www.researchgate.net/publication/6072782
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spelling my.unimas.ir.139922021-07-12T15:40:25Z http://ir.unimas.my/id/eprint/13992/ Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis Seiya, Watanabe Abu Saleh, Ahmed Narayana, Visu Annaluru TP Chemical technology A recombinant Saccharomyces cerevisiae strain transformed with xylose reductase (XR) and xylitol dehydrogenase (XDH) genes from Pichia stipitis (PsXR and PsXDH, respectively) has the ability to convert xylose to ethanol together with the unfavourable excretion of xylitol, which may be due to intercellular redox imbalance caused by the different coenzyme specificity between NADPH-preferring XR and NAD+-dependent XDH. In this study, we focused on the effect(s) of mutated NADH-preferring PsXR in fermentation. The R276H and K270R/N272D mutants were improved 52- and 146-fold, respectively, in the ratio of NADH/NADPH in catalytic efficiency [(kcat/Km with NADH)/(kcat/Km with NADPH)] compared with the wild-type (WT), which was due to decrease of kcat with NADPH in the R276H mutant and increase of Km with NADPH in the K270R/N272D mutant. Furthermore, R276H mutation led to significant thermostabilization in PsXR. The most positive effect on xylose fermentation to ethanol was found by using the Y-R276H strain, expressing PsXR R276H mutant and PsXDH WT: 20% increase of ethanol production and 52% decrease of xylitol excretion, compared with the Y-WT strain expressing PsXR WT and PsXDH WT. Measurement of intracellular coenzyme concentrations suggested that maintenance of the of NADPH/NADP+ and NADH/NAD+ ratios is important for efficient ethanol fermentation from xylose by recombinant S. cerevisiae. CSIRO Publishing 2007 Article PeerReviewed text en http://ir.unimas.my/id/eprint/13992/1/Seiya.pdf Seiya, Watanabe and Abu Saleh, Ahmed and Narayana, Visu Annaluru (2007) Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis. Microbiology Australia, 153. pp. 3044-3054. ISSN 1324-4272 https://www.researchgate.net/publication/6072782 DOI: 10.1099/mic.0.2007/007856-0 ·
institution Universiti Malaysia Sarawak
building Centre for Academic Information Services (CAIS)
collection Institutional Repository
continent Asia
country Malaysia
content_provider Universiti Malaysia Sarawak
content_source UNIMAS Institutional Repository
url_provider http://ir.unimas.my/
language English
topic TP Chemical technology
spellingShingle TP Chemical technology
Seiya, Watanabe
Abu Saleh, Ahmed
Narayana, Visu Annaluru
Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis
description A recombinant Saccharomyces cerevisiae strain transformed with xylose reductase (XR) and xylitol dehydrogenase (XDH) genes from Pichia stipitis (PsXR and PsXDH, respectively) has the ability to convert xylose to ethanol together with the unfavourable excretion of xylitol, which may be due to intercellular redox imbalance caused by the different coenzyme specificity between NADPH-preferring XR and NAD+-dependent XDH. In this study, we focused on the effect(s) of mutated NADH-preferring PsXR in fermentation. The R276H and K270R/N272D mutants were improved 52- and 146-fold, respectively, in the ratio of NADH/NADPH in catalytic efficiency [(kcat/Km with NADH)/(kcat/Km with NADPH)] compared with the wild-type (WT), which was due to decrease of kcat with NADPH in the R276H mutant and increase of Km with NADPH in the K270R/N272D mutant. Furthermore, R276H mutation led to significant thermostabilization in PsXR. The most positive effect on xylose fermentation to ethanol was found by using the Y-R276H strain, expressing PsXR R276H mutant and PsXDH WT: 20% increase of ethanol production and 52% decrease of xylitol excretion, compared with the Y-WT strain expressing PsXR WT and PsXDH WT. Measurement of intracellular coenzyme concentrations suggested that maintenance of the of NADPH/NADP+ and NADH/NAD+ ratios is important for efficient ethanol fermentation from xylose by recombinant S. cerevisiae.
format Article
author Seiya, Watanabe
Abu Saleh, Ahmed
Narayana, Visu Annaluru
author_facet Seiya, Watanabe
Abu Saleh, Ahmed
Narayana, Visu Annaluru
author_sort Seiya, Watanabe
title Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis
title_short Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis
title_full Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis
title_fullStr Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis
title_full_unstemmed Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing proteinengineered NADH-preferring xylose reductase from Pichia stipitis
title_sort ethanol production from xylose by recombinant saccharomyces cerevisiae expressing proteinengineered nadh-preferring xylose reductase from pichia stipitis
publisher CSIRO Publishing
publishDate 2007
url http://ir.unimas.my/id/eprint/13992/1/Seiya.pdf
http://ir.unimas.my/id/eprint/13992/
https://www.researchgate.net/publication/6072782
_version_ 1706961332321910784
score 13.211869

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