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Efficient Anaerobic Whole Cell Stereoselective Bioreduction with Recombinant Saccharomyces cerevisiae.

Katz, Michael LU ; Frejd, Torbjörn LU ; Hahn-Hägerdal, Bärbel LU and Gorwa-Grauslund, Marie-Francoise LU (2003) In Biotechnology and Bioengineering 84(5). p.573-582
Abstract
In this study we investigate the NADPH-dependent stereoselective reduction of the bicyclic diketone bicyclo[2.2.2]octane-2,6-dione (BCO2,6D) to the chiral ketoalcohol (1R,4S,6S)-6-hydroxybicyclo[2.2.2]octane-2-one (BCO2one6ol). Our aim was to develop a whole cell batch process for reduction of carbonyl substrates with (i) a high cosubstrate yield (formed product/consumed cosubstrate) and (ii) a high conversion rate under anaerobic conditions with Saccharomyces cerevisiae as biocatalyst and glucose as cosubstrate. Five open reading frames (ORFs), YMR226c, YDR368w, YOR120w, YGL157w, and YGL039w, encoding reductases involved in the conversion of BCO2,6D were identified using cell-free extract from strains belonging to the ExClone collection... (More)
In this study we investigate the NADPH-dependent stereoselective reduction of the bicyclic diketone bicyclo[2.2.2]octane-2,6-dione (BCO2,6D) to the chiral ketoalcohol (1R,4S,6S)-6-hydroxybicyclo[2.2.2]octane-2-one (BCO2one6ol). Our aim was to develop a whole cell batch process for reduction of carbonyl substrates with (i) a high cosubstrate yield (formed product/consumed cosubstrate) and (ii) a high conversion rate under anaerobic conditions with Saccharomyces cerevisiae as biocatalyst and glucose as cosubstrate. Five open reading frames (ORFs), YMR226c, YDR368w, YOR120w, YGL157w, and YGL039w, encoding reductases involved in the conversion of BCO2,6D were identified using cell-free extract from strains belonging to the ExClone collection (yeast ORF expression clones; ResGen, Invitrogen Corp., UK). We report the one-step purification and characterization of three major BCO2,6D reductases, YMR226cp, YDR368wp (YPR1p), and YOR120wp (GCY1p). The reductases were overexpressed under a strong constitutive promoter and the impact on cosubstrate yield, conversion time, glucose consumption rate, and reduction rate was investigated when reductases were overexpressed either alone or in combination with low phosphoglucose isomerase activity (encoded by YBR196c). Combining overexpression of BCO2,6D reductase with reduced glycolytic rate (low phosphoglucose isomerase activity) offers a fast whole cell stereoselective bioreduction system useful for facilitated anaerobic batch conversions. © 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 84: 573-582, 2003. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Biotechnology and Bioengineering
volume
84
issue
5
pages
573 - 582
publisher
John Wiley & Sons
external identifiers
  • wos:000186316800007
  • pmid:14574691
  • scopus:0242291798
ISSN
1097-0290
DOI
10.1002/bit.10824
language
English
LU publication?
yes
id
964e6dd3-3e91-47aa-b913-6f92572bfe32 (old id 128578)
date added to LUP
2007-06-28 14:10:04
date last changed
2018-05-29 11:34:24
@article{964e6dd3-3e91-47aa-b913-6f92572bfe32,
  abstract     = {In this study we investigate the NADPH-dependent stereoselective reduction of the bicyclic diketone bicyclo[2.2.2]octane-2,6-dione (BCO2,6D) to the chiral ketoalcohol (1R,4S,6S)-6-hydroxybicyclo[2.2.2]octane-2-one (BCO2one6ol). Our aim was to develop a whole cell batch process for reduction of carbonyl substrates with (i) a high cosubstrate yield (formed product/consumed cosubstrate) and (ii) a high conversion rate under anaerobic conditions with Saccharomyces cerevisiae as biocatalyst and glucose as cosubstrate. Five open reading frames (ORFs), YMR226c, YDR368w, YOR120w, YGL157w, and YGL039w, encoding reductases involved in the conversion of BCO2,6D were identified using cell-free extract from strains belonging to the ExClone collection (yeast ORF expression clones; ResGen, Invitrogen Corp., UK). We report the one-step purification and characterization of three major BCO2,6D reductases, YMR226cp, YDR368wp (YPR1p), and YOR120wp (GCY1p). The reductases were overexpressed under a strong constitutive promoter and the impact on cosubstrate yield, conversion time, glucose consumption rate, and reduction rate was investigated when reductases were overexpressed either alone or in combination with low phosphoglucose isomerase activity (encoded by YBR196c). Combining overexpression of BCO2,6D reductase with reduced glycolytic rate (low phosphoglucose isomerase activity) offers a fast whole cell stereoselective bioreduction system useful for facilitated anaerobic batch conversions. © 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 84: 573-582, 2003.},
  author       = {Katz, Michael and Frejd, Torbjörn and Hahn-Hägerdal, Bärbel and Gorwa-Grauslund, Marie-Francoise},
  issn         = {1097-0290},
  language     = {eng},
  number       = {5},
  pages        = {573--582},
  publisher    = {John Wiley & Sons},
  series       = {Biotechnology and Bioengineering},
  title        = {Efficient Anaerobic Whole Cell Stereoselective Bioreduction with Recombinant Saccharomyces cerevisiae.},
  url          = {http://dx.doi.org/10.1002/bit.10824},
  volume       = {84},
  year         = {2003},
}