Methylobacterium rhodesianumMB 126 possesses two stereospecific crotonyl-CoA hydratases

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Abstract
Two distinct crotonyl-CoA hydratases of Methylobacterium rhodesianum MB 126 were separated by column chromatography on DEAE-Sepharose CL-6B. The two enzymes were further purified by chromatography on red 120 agarose and Mono Q. Enzyme A was specific for L(+)-hydroxybutyryl-CoA. It had an apparent molecular weight of 160 000 with two identical subunits of 43 000 and 34 000. The apparent Kmvalues for L(+)-hydroxybutyryl-CoA and crotonyl-CoA were 83 and 90 μM, respectively. Enzyme B was specific for D(−)-hydroxybutyryl-CoA. It had an apparent molecular weight of 39 000 with identical subunits of 12 500. It showed sigmoidal kinetics for crotonyl-CoA, and the Hill coefficient was about 2.5. The apparent Kmvalue for D(−)-hydroxybutyryl-CoA was 0.5 mM. The possible contribution of a sequence including β-ketothiolase, NADH-linked L(+)-specific acetoacetyl-CoA reductase, and two stereospecific crotonyl-CoA hydratases to PHB synthesis in methylotrophic serine-pathway bacteria is discussed.Key words: poly(β-hydroxybutyrate), PHB, crotonyl-CoA hydratase, methylotroph, Methylobacterium rhodesianum.