title: Characterization and site-directed mutation of a novel aldo-keto reductase from Lodderomyces elongisporus NRRL YB-4239 with high production rate of ethyl (R)-4-chloro-3-hydroxybutanoate.
abstract: A novel aldo-keto reductase (LEK) from Lodderomyces elongisporus NRRL YB-4239 (ATCC 11503) was discovered by genome database mining for carbonyl reduction. LEK was overexpressed in Escherichia coli BL21 (DE3), purified to homogeneity and the catalytic properties were studied. Among the substrates, ethyl 4-chloro-3-oxobutanoate was converted to ethyl (R)-4-chloro-3- hydroxybutanoate ((R)-CHBE), an important pharmaceutical intermediate, with an excellent enantiomeric excess (e.e.) (>99 %). The mutants W28A and S209G obtained by site-directed mutation were identified with much higher molar conversion yields and lower Km values. Further, the constructed coenzyme regeneration system with glucose as co-substrate resulted in a yield of 100 %, an enantioselectivity of >99 %, and the calculated production rate of 56.51 mmol/L/H. These results indicated the potential of LEK for the industrial production of (R)-CHBE and other valuable chiral alcohols.
authors.affiliation: College of Material, Chemistry and Chemical Engineering, Hangzhou Normal University, No. 1378, Wenyixi Road, Hangzhou, 311121, People's Republic of China.
authors.forename: Qiuyan
authors.initials: Q
authors.lastname: Wang
authors.forename: Tingting
authors.initials: T
authors.lastname: Ye
authors.forename: Zhuanzhuan
authors.initials: Z
authors.lastname: Ma
authors.forename: Rong
authors.initials: R
authors.lastname: Chen
authors.forename: Tian
authors.initials: T
authors.lastname: Xie
authors.forename: Xiaopu
authors.initials: X
authors.lastname: Yin
mesh_terms: D000090:Acetoacetates / Q000378:metabolism; D000449:Aldehyde Reductase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000074408:Aldo-Keto Reductases; D002087:Butyrates / Q000378:metabolism*; D003067:Coenzymes / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D004718:Saccharomycetales / Q000201:enzymology* / Q000235:genetics
chemical_list: D000090:Acetoacetates; D002087:Butyrates; D003067:Coenzymes; C121244:ethyl 4-chloro-3-hydroxybutanoate; C111982:ethyl 4-chloro-3-oxobutanoate; D000074408:Aldo-Keto Reductases; D000449:Aldehyde Reductase