# US 9,040,265 B2 · Oxidoreductases for the stereoselective reduction of keto compounds > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 9,040,265 B2 - Document ID: US9040265B2 - Status: Granted - Kind: B2 - Application number: 13/280,876 - Filed: 2011-10-25 - Published: 2014-01-16 - Granted: 2015-05-26 - Earliest claimed: 2005-07-27 - Assignees: Iep GmbH - Applicants: Anke Tschentscher; Antje Gupta; Maria Bobkova - Inventors: Anke Tschentscher; Antje Gupta; Maria Bobkova - Web page: https://patentgrep.com/patent/US9040265B2 ## CPC - C12P 7/62: Carboxylic acid esters - C12N 9/0004: {Oxidoreductases (1.)} - C12N 9/0008: {acting on the aldehyde or oxo group of donors (1.2)} - C12P 7/02: containing a hydroxy group - C12P 7/04: acyclic - C12P 7/42: Hydroxy-carboxylic acids - C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)} - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract The invention relates to a process for the enantioselective enzymatic reduction of a keto compound to the corresponding chiral hydroxy compound, wherein the keto compound is reduced with an oxidoreductase in the presence of a cofactor, and is characterized in that an oxidoreductase is used which has an amino acid sequence in which (a) at least 70% of the amino acids are identical to the amino acids of one of the amino acid sequences SEQ ID No 1, SEQ ID No 6 and SEQ ID No 8, or (b) at least 55% of the amino acids are identical to the amino acids of the amino acid sequence SEQ ID No 2, or (c) at least 65% of the amino acids are identical to the amino acids of the amino acid sequence SEQ ID No 3, or (d) at least 75% of the amino acids are identical to the amino acids of the amino acid sequence SEQ ID No 4, or (e) at least 65% of the amino acids are identical to the amino acids of the amino acid sequence SEQ ID No 5, or (0 at least 50% of the amino acids are identical to the amino acids of the amino acid sequence SEQ ID No 7, or (g) at least 72% of the amino acids are identical to the amino acids of the amino acid sequence SEQ ID No 129. ## Claims ### Claim 1 A recombinant host cell which is a bacterial, insect, plant or isolated mammalian cell and has been transformed or transfected with an expression vector comprising: a nucleic acid sequence encoding an oxidoreductase and which is linked to an expression control sequence, wherein the oxidoreductase has an amino acid sequence at least 70% identical to SEQ ID No:6 or fragment thereof and reduces 2-octanone to (S)-2-octanol and/or 4-haloacetoacetate ester to (R)-4-halo-3-hydroxybutyric ester in the presence of NADH or NADPH with enantiomeric purity of more than 95%. ### Claim 2 (depends on 1) The recombinant host cell according to claim 1, wherein the oxidoreductase is enzymatically active for the enantioselective enzymatic reduction of 2-octanone to (S)-2-octanol with enantiomeric purity of 100%. ### Claim 3 (depends on 1) The recombinant host cell according to claim 1, wherein the oxidoreductase is enzymatically active for the enantioselective enzymatic reduction of 4-haloacetoacetate ester to (R)-4-halo-3-hydroxybutyric ester with enantiomeric purity of 100%. ### Claim 4 An expression vector comprising: a nucleic acid sequence encoding an oxidoreductase and is linked to an expression control sequence, wherein the codes for an oxidoreductase has an amino acid sequence at least 70% identical to SEQ ID No:6 or fragment thereof and reduces 2-octanone to (S)-2-octanol and/or 4-haloacetoacetate ester to (R)-4-halo-3-hydroxybutyric ester in the presence of NADH or NADPH with enantiomeric purity of more than 95%. ### Claim 5 (depends on 4) The expression vector of claim 4, wherein the oxidoreductase is enzymatically active for the enantioselective enzymatic reduction of 2-octanone to (S)-2-octanol with enantiomeric purity of 100%. ### Claim 6 (depends on 4) The expression vector according to claim 4, wherein the oxidoreductase is enzymatically active for the enantioselective enzymatic reduction of 4-haloacetoacetate ester to (R)-4-halo-3-hydroxybutyric ester with enantiomeric purity of 100%. ### Claim 7 A cloning vector comprising: a nucleic acid sequence encoding an oxidoreductase having an amino acid sequence at least 70% identical to SEQ ID No:6 or fragment thereof, wherein said oxidoreductase reduces 2-octanone to (S)-2-octanol and/or 4-haloacetoacetate ester to (R)-4-halo-3-hydroxybutyric ester in the presence of NADH or NADPH with enantiomeric purity of more than 95%. ### Claim 8 (depends on 7) The cloning vector of claim 7, wherein the oxidoreductase is enzymatically active for the enantioselective enzymatic reduction of 2-octanone to (S)-2-octanol with enantiomeric purity of 100%. ### Claim 9 (depends on 7) The cloning vector of claim 7, wherein the oxidoreductase is enzymatically active for the enantioselective enzymatic reduction of 4-haloacetoacetate ester to (R)-4-halo-3-hydroxybutyric ester with enantiomeric purity of 100%. ### Claim 10 A recombinant host cell which is a bacterial, insect, plant or isolated mammalian cell and has been transformed or transfected with an expression vector comprising a nucleic acid sequence encoding an oxidoreductase having an amino acid sequence of at least 95% identical to SEQ ID No:6 or enzymatically active fragment thereof. ### Claim 11 An expression vector comprising a nucleic acid sequence encoding an oxidoreductase having an amino acid sequence at least 95% identical to SEQ ID No:6 or enzymatically active fragment thereof. ### Claim 12 A cloning vector comprising a nucleic acid sequence encoding an oxidoreductase having an amino acid sequence at least 95% identical to SEQ ID No:6 or enzymatically active fragment thereof. ### Claim 13 A process for the enantioselective enzymatic reduction of a keto compound to the corresponding chiral hydroxy compound comprising contacting the keto compound with an oxidoreductase having an amino acid sequence at least 95% identical to SEQ ID NO:6 or enzymatically active fragment thereof and NADP or NADPH, wherein the keto compound is reduced to the corresponding alcohol.