Stereoselective synthesis of (R)-3-quinuclidinol through asymmetric reduction of 3-quinuclidinone with 3-quinuclidinone reductase of Rhodotorula rubra.
A novel nicotinamide adenine dinucleotide phosphate-dependent carbonyl reductase, 3-quinuclidinone reductase, was isolated from Rhodotorula rubra JCM3782. The enzyme catalyzes the asymmetric reduction of 3-quinuclidinone to (R)-3-quinuclidinol. The gene encoding the enzyme was also cloned and sequenced. A 819-bp nucleotide fragment was confirmed to be the gene encoding the 3-quinuclidinone reductase by agreement of the internal amino acid sequences of the purified enzyme. The gene encodes a total of 272 amino acid residues, and the deduced amino acid sequence shows similarity to those of several short-chain dehydrogenase/reductase family proteins. An expression vector, pWKLQ, which contains the full length 3-quinuclidinone reductase gene was constructed. Using Escherichia coli cells coexpressing the 3-quinuclidinone reductase and glucose dehydrogenase (cofactor regeneration enzyme) genes, 618 mM 3-quinuclidinone was almost stiochiometrically converted to (R)-3-quinuclidinol with an >99.9% enantiomeric excess within 21 h of reaction.
Uzura
Atsuko
A
Research & Development Center, Nagase & Co., Ltd., Kobe, Japan. atsuko.uzura@nagase.co.jp
Nomoto
Fumiki
F
Sakoda
Akiko
A
Nishimoto
Yukifumi
Y
Kataoka
Michihiko
M
Shimizu
Sakayu
S
D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D001711:Biotransformation; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015870:Gene Expression; D042843:Glucose 1-Dehydrogenase / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011812:Quinuclidines / Q000378:metabolism*; D012248:Rhodotorula / Q000201:enzymology* / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013237:Stereoisomerism
D004271:DNA, Fungal; D005656:Fungal Proteins; D011812:Quinuclidines; C090998:3-quinuclidinone; C474540:3-quinuclidinol; D000429:Alcohol Oxidoreductases; C019642:alcohol dehydrogenase (NADP+); D042843:Glucose 1-Dehydrogenase