Cobalamin (vitamin B12) biosynthesis: functional characterization of the Bacillus megaterium cbi genes required to convert uroporphyrinogen III into cobyrinic acid a,c-diamide.

The function of individual genes of the Bacillus megaterium cobI operon genes in cobalamin (vitamin B12) biosynthesis was investigated by their ability to complement defined Salmonella typhimurium cob mutants. This strategy confirmed the role of cbiA, -D, -F, -J, -L and cysGA. Furthermore the operon as a whole was used to restore corrin biosynthesis in Escherichia coli, which, although closely related to S. typhimurium, does not possess the CobI pathway. When the B. megaterium cob operon was cloned into a plasmid and transformed into an E. coli strain containing the S. typhimurium cbiP, it conferred upon the host strain the ability to make the cobyric acid de novo. However, cobyric acid synthesis was observed only when the strain was grown anaerobically. Derivatives of the corrin-producing E. coli strain were constructed in which genes of the B. megaterium cob operon had been inactivated. These strains were used to demonstrate that, whereas B. megaterium cbiD, -G and -X are essential for cobyric acid synthesis, the cbiW and -Y genes could be deleted without detriment to cobyric acid production in E. coli.