Reaction and strain engineering for improved stereo-selective whole-cell reduction of a bicyclic diketone.
Reduction of bicyclo[2.2.2]octane-2,6-dione to (1R, 4S, 6S)-6-hydroxy-bicyclo[2.2.2]octane-2-one by whole cells of Saccharomyces cerevisiae was improved using an engineered recombinant strain and process design. The substrate inhibition followed a Han-Levenspiel model showing an effective concentration window between 12 and 22 g/l, in which the activity was kept above 95%. Yeast growth stage, substrate concentration and a stable pH were shown to be important parameters for effective conversion. The over-expression of the reductase gene YDR368w significantly improved diastereoselectivity compared to previously reported results. Using strain TMB4110 expressing YDR368w in batch reduction with pH control, complete conversion of 40 g/l (290 mM) substrate was achieved with 97% diastereomeric excess (de) and >99 enantiomeric excess (ee), allowing isolation of the optically pure ketoalcohol in 84% yield.
Johanson
Ted
T
Department of Applied Microbiology, Lund University, P.O. Box 124, SE-22100, Lund, Sweden.
Carlquist
Magnus
M
Olsson
Cecilia
C
Rudolf
Andreas
A
Frejd
Torbjörn
T
Gorwa-Grauslund
Marie F
MF
D001643:Bridged Bicyclo Compounds / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D010084:Oxidation-Reduction; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013237:Stereoisomerism; D013379:Substrate Specificity / Q000235:genetics
C487950:6-hydroxybicyclo(2.2.2)octane-2-one; D001643:Bridged Bicyclo Compounds; D029701:Saccharomyces cerevisiae Proteins; C487951:bicyclo(2.2.2)octane-2,6-dione