Correlation between the Level of Vitamin-B12-Dependent Methionine Synthetase and Intracellular Concentration of Vitamin B12 in Some Bacteria.

This paper presents several lines of evidence indicating that the synthesis of vitamin-B12-dependent 5-methyltetrahydrofolate-homocysteine transmethylase ("methionine synthetase") is closely correlated to the intracellular level of vitamin B12 in manners different between bacteria which do and do not produce vitamin B12. 1. The intracellular level of vitamin B12 in Rhodospirillum rubrum was reduced to one fifth that of the normal cells when the organism was cultivated in a medium lacking cobalt. In accord with the decrease in vitamin B12 production, the specific activity of methionine synthetase was lowered to about 50%. It was of much interest that the enzyme was always present as the holoenzyme and the apoenzyme could not be detected even in the vitamin-B12-deficient cells containing a markedly decreased level of the holoenzyme. When the vitamin-B12-deficient cells were incubated in a cyano- cobalamin-supplemented medium, the specific activity of methionine synthetase began to increase after 2 h lag time and reached the maximum in about 6 h. This enhancement of the specific activity was suppressed if an appropriate inhibitor of protein synthesis was added to the medium. These facts strongly suggest that synthesis of protein de novo is essential for the enhancement of the specific activity by incubation with cyanocobalamin. Similar results were obtained in other vitamin B12 producers such as Streptomyces olivaceus and Corynebacterium simplex. 2. Chromatium D which is known to form cobalt-free corrinoids but not produce complete vitamin B12, was found to possess methionine synthetase as apoenzyme. When the vitamin-B12-free cells were incubated in the presence of cyanocobalamin, all the apoenzyme was readily converted to the holoenzyme within 2 h. As the formation of the holoenzyme proceeded, the level of the total enzyme (holo + apo) increased to a certain degree. The enhancement, however, was not affected by the inhibitors of protein synthesis, indicating that the increment could not have resulted from de novo synthesis of protein. A similar phenomenon was observed in Escherichia coli which does not produce vitamin B12 but possesses apoprotein of methionine synthetase. It is suggested that vitamin B12 synthesis is closely correlated to that of methionine synthetase in bacteria producing the vitamin, such as Rhodospirllum rubrum, Streptomyces olivaceus and Corynebacterium simplex. On the contrary, the apoprotein of the enzyme is synthesized independently of vitamin B12 in bacteria not producing the vitamin, such as Chromatium D and Escherichia coli. [ABSTRACT FROM AUTHOR]