2-Haloacrylate reductase, a novel enzyme of the medium chain dehydrogenase/reductase superfamily that catalyzes the reduction of a carbon-carbon double bond of unsaturated organohalogen compounds.
A soil bacterium, Burkholderia sp. WS, grows on 2-chloroacrylate as the sole carbon source. To identify the enzymes metabolizing 2-chloroacrylate, we carried out comparative two-dimensional gel electrophoresis of the proteins from 2-chloroacrylate- and lactate-grown bacterial cells. As a result, we found that a protein named CAA43 was inducibly synthesized when the cells were grown on 2-chloroacrylate. The CAA43 gene was cloned and shown to encode a protein of 333 amino acid residues (M(r) 35,788) that shared a significant sequence similarity with NADPH-dependent quinone oxidoreductase from Escherichia coli (38.2% identity). CAA43 was overproduced in E. coli and purified to homogeneity. The purified protein catalyzed the NADPH-dependent reduction of the carbon-carbon double bond of 2-chloroacrylate to produce (S)-2-chloropropionate, which is probably further metabolized to (R)-lactate by (S)-2-haloacid dehalogenase in Burkholderia sp. WS. NADH did not serve as a reductant. Despite the sequence similarity to quinone oxidoreductases, CAA43 did not act on 1,4-benzoquinone and 1,4-naphthoquinone. 2-Chloroacrylate analogs, such as acrylate and methacrylate, were also inert as the substrates. In contrast, 2-bromoacrylate served as the substrate. Thus, we named this novel enzyme 2-haloacrylate reductase. This study revealed a new pathway for the degradation of unsaturated organohalogen compounds. It is also notable that the enzyme is useful for the production of (S)-2-chloropropionate, which is used for the industrial production of aryloxyphenoxypropionic acid herbicides.
Kurata
Atsushi
A
Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan.
Kurihara
Tatsuo
T
Kamachi
Harumi
H
Esaki
Nobuyoshi
N
D000179:Acrylates / Q000378:metabolism*; D000594:Amino Acid Oxidoreductases / Q000737:chemistry; D000595:Amino Acid Sequence; D019117:Burkholderia / Q000201:enzymology*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D006843:Hydrocarbons, Chlorinated; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000737:chemistry; D009249:NADP / Q000494:pharmacology; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011422:Propionates / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D021241:Spectrometry, Mass, Electrospray Ionization; D013379:Substrate Specificity; D013696:Temperature
C502742:2-chloroacrylic acid; D000179:Acrylates; D029968:Escherichia coli Proteins; D006843:Hydrocarbons, Chlorinated; D011422:Propionates; D009249:NADP; C021974:2-chloropropionic acid; D010088:Oxidoreductases; D000594:Amino Acid Oxidoreductases; C031955:L-aspartate oxidase, E coli; D016660:NAD(P)H Dehydrogenase (Quinone)