Process for the stereoselective enzymatic reduction of keto compounds

摘要

In a process for the stereoselective, in particular enantioselective enzymatic reduction of keto compounds to the corresponding chiral hydroxy compounds, wherein the keto compounds are reduced with an enantioselective, NADH-specific oxidoreductase, a polypeptide is used for reducing the keto compounds, which polypeptide exhibits an R-ADH-signature H-[P; A]-[I; A; Q; V; L]-[G; K]-R at positions 204-208 and the following further structural features in their entirety:
(i) an N-terminal Rossmann-Fold GxxxGxG,(ii) an NAG-motif at position 87,(iii) a catalytic triad consisting of S 139, Y 152 and K 156,(iv) a negatively charged amino acid moiety at position 37,(v) two C-terminal motifs in the dimerization domain [A; S]-S-F and [V; I]-DG-[G; A]-Y-[T; C; L]-[A; T; S]-[Q; V; R; L; P],(vi) Val or Leu at position 159 (4 positions downstream of K 156),(vii) Asn at position 178, and(viii) a proline moiety at position 188.

主权项

1. A process for stereoselective enzymatic reduction of keto compounds to corresponding chiral hydroxy compounds, wherein the keto compounds are reduced with an enantioselective, NADH-specific oxidoreductase, the process comprising:
reducing the keto compounds using a polypeptide which exhibits an R-ADH-signature H-[P; A]-[I; A; Q; V; L]-[G; K]-R at positions 205-209 and has the following further structural features in their entirety: (i) an N-terminal Rossmann-Fold GxxxGxG, (ii) an NAG-motif at position 87, (iii) a catalytic triad consisting of S 139, Y 152 and K 156, (iv) a negatively charged amino acid moiety at position 37, (v) two C-terminal motifs in the dimerization domain [A; S]-S-F and [V; I]-DG-[G; A]-Y-[T; C; L]-[A; T; S]-[Q; V; R; L; P], (vi) Val or Leu at position 159 (4 positions downstream of K 156), (vii) Asn at position 178, and (viii) a proline moiety at position 188, wherein the position of the R-ADH-signature H-[P; A]-[I; A; Q; V; L]-[G; K]-R and structural features (i) to (viii) refer to 2bhd_Strex by multi-way amino acid sequence comparison and wherein:(a) the polypeptide comprises one of the amino acid sequences SEQ ID NO:1 to SEQ ID NO:7, or(b) the polypeptide is encoded by a nucleic acid sequence from the group consisting of SEQ ID NO:8 to SEQ ID NO:14, or(c) the polypeptide is encoded by a nucleic acid sequence which hybridizes to one of the nucleic acid sequences mentioned in (b) and has at least 80% sequence identity to one of the nucleic acid sequences mentioned in (b).