Carbonyl reductase, gene thereof and method of using the same
AssigneeKaneka Corporation
InventorsNoriyuki Kizaki, Yukio Yamada, Yoshihiko Yasohara, Junzo Hasegawa
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Abstract
A novel polypeptide for producing tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate, a gene encoding the polypeptide, and a method of using the polypeptide, are provided. The polypeptide has an enzyme activity to asymmetrically reduce tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate to tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate. The polypeptide comprises the amino acid sequence represented by SEQ ID NO. 1 in the Sequence Listing, or the amino acid sequence having at least one amino acid substitution, insertion, deletion, or addition thereof.
Claims15
An isolated polypeptide having an enzyme activity to asymmetrically reduce tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate to tert-butyl (3R, 5S)-6-chloro-3,5-dihydroxyhexanoate, wherein the polypeptide comprises the amino acid sequence of SEQ ID NO:
- 1.
An isolated polynucleotide encoding a polypeptide having an enzyme activity to asymmetrically reduce tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate to tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate, wherein the polynucleotide hybridizes to a nucleotide sequence represented by SEQ ID NO:
- 2 under stringent conditions;
- whereby the stringent conditions comprise hybridization conditions having about 0.7 to about 1.0M sodium chloride concentration at about 65° C. and wash conditions having about 0.015M sodium chloride concentration to about 0.3M sodium chloride concentration at about 65° C.
An expression vector comprising a polynucleotide encoding a polypeptide having an enzyme activity to asymmetrically reduce tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate to tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate, wherein the polynucleotide hybridizes to a nucleotide sequence represented by SEQ ID NO:
- 2 under stringent conditions;
- whereby the stringent conditions comprise hybridization conditions having about 0.7 to about 1.0M sodium chloride concentration at about 65° C. and wash conditions having about 0.015M sodium chloride concentration to about 0.3M sodium chloride concentration at about 65° C.
An isolated polypeptide having an enzyme activity to asymmetrically reduce tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate to tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate, wherein the polypeptide is encoded by a polynucleotide that hybridizes to a nucleotide sequence represented by SEQ ID NO:
- 2 under stringent conditions;
- whereby the stringent conditions comprise hybridization conditions having about 0.7 to about 1.0M sodium chloride concentration at about 65° C. and wash conditions having about 0.015M sodium chloride concentration to about 0.3M sodium chloride concentration at about 65° C.
A method for producing a tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate comprising the steps:
- a) reacting a host cell, or cell lysate thereof, said cell comprising an expression vector further comprising a polynucleotide encoding a polypeptide having an enzyme activity to asymmetrically reduce tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate to tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate wherein the polynucleotide hybridizes to a nucleotide sequence represented by SEQ ID NO:
- 2 under stringent conditions, whereby the stringent conditions comprise hybridization conditions having about 0.7 to about 1.0 M sodium chloride concentration at about 65° C. and wash conditions having about 0.015 M sodium chloride concentration to about 0.3 M sodium chloride concentration at about 65° C., with tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate;
- and
- b) obtaining said tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate.
Dependencies are inferred from the claim text.