J. Biochem, 1982, Vol. 91, No. 4 1163-1171
© 1982 Japanese Biochemical Society
research-article |
Methionine Biosynthesis in Brevibacterium flavum: Properties and Essential Role of O-Acetylhomoserine Sulfhydrylase
Central Research Laboratories Ajinomoto Co , Inc, Kawasaki-ku, Kawasaki, Kanagawa 210
Out of 27 strains of methionine auxotrophs ofBrevibacterium flavum, 14 strains did not grow on homoserine but grew on O-acetylhomoserine, and all were found to lack homoserine O-acetyltransferase [EC 2.3.1.31
[EC]
] alone. Another 3 strains did not grow on O-acetylhomoserine but grew on homocysteine, and the two strains tested were found to lack O-acetylhomoserine sulfhydrylase (AHS) alone, without any changes in the activities of cystathionine 
-synthase [EC 4.2.99.9
[EC]
] and ß-cystath-ionase [EC 4.4.1.8
[EC]
]. Prototrophic revertants of the AHS-lacking mutants showed concomitant reversion of AHS activity. None of the methionine auxotrophs grew on cystathionine. From these results it was concluded that the methionine biosyn-thetic pathway of this bacterium involves formation of O-acetylhomoserine from homoserine by the action of homoserine O-acetyltransferase, and direct formation of homocysteine from O-acetylhomoserine by the AHS reaction. AHS synthesis was strongly repressed by methionine. AHS was purified to 70% purity. The purified preparation was activated by pyridoxal phosphate after treatment with hydroxylamine. The enzyme showed a molecular weight of 360,000, an optimum pH of 8.7 for activity, and specifically reacted with O-acetyl-L-homoserine and showed with O-acetyl-L-serine one hundredth as much activity as that with O-acetylhomoserine, but did not show activity with O-succinyl-L-homoserine, homoserine, or serine. The Km values for O-acetylhomoserine and H2S were 2.0 mM and 0.08 mM, respectively. The enzyme was inhibited 50, 23, and 29% by 10 mM L-meth-ionine, L-homoserine, and O-acetyl-L-serine, respectively, but it was not inhibited by cystathionine or S-adenosyl-L-methionine.
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