title: Enantioselective bioconversion using Escherichia coli cells expressing Saccharomyces cerevisiae reductase and Bacillus subtilis glucose dehydrogenase.
abstract: Ethyl (R, S)-4-chloro-3-hydroxybutanoate (ECHB) is a useful chiral building block for the synthesis of L-carnitine and hypercholesterolemia drugs. The yeast reductase, YOL151W (GenBank locus tag), exhibits an enantioselective reduction activity, converting ethyl-4-chlorooxobutanoate (ECOB) exclusively into (R)-ECHB. YOL151W was generated in Escherichia coli cells and purified via Ni- NTA and desalting column chromatography. It evidenced an optimum temperature of 45 degrees C and an optimum pH of 6.5-7.5. Bacillus subtilis glucose dehydrogenase (GDH) was also expressed in Escherichia coli, and was used for the recycling of NADPH, required for the reduction reaction. Thereafter, Escherichia coli cells co-expressing YOL151W and GDH were constructed. After permeablization treatment, the Escherichia coli whole cells were utilized for ECHB synthesis. Through the use of this system, the 30 mM ECOB substrate could be converted to (R)-ECHB.
authors.affiliation: Division of Biotechnology, The Catholic University of Korea, Bucheon 420-743, Korea.
authors.forename: Hyun Joo
authors.initials: HJ
authors.lastname: Park
authors.forename: Jihye
authors.initials: J
authors.lastname: Jung
authors.forename: Hyejeong
authors.initials: H
authors.lastname: Choi
authors.forename: Ki-Nam
authors.initials: KN
authors.lastname: Uhm
authors.forename: Hyung Kwoun
authors.initials: HK
authors.lastname: Kim
mesh_terms: D000090:Acetoacetates / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology*; D001709:Biotechnology / Q000379:methods*; D002087:Butyrates / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D015870:Gene Expression; D005818:Genetic Engineering; D042843:Glucose 1-Dehydrogenase / Q000235:genetics*; D009249:NADP / Q000378:metabolism; D010088:Oxidoreductases / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics*
chemical_list: D000090:Acetoacetates; D002087:Butyrates; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C121244:ethyl 4-chloro-3-hydroxybutanoate; D009249:NADP; C111982:ethyl 4-chloro-3-oxobutanoate; D010088:Oxidoreductases; C432953:GRE2 protein, S cerevisiae; D042843:Glucose 1-Dehydrogenase