title: Reassessment of the late steps of coenzyme B12 synthesis in Salmonella enterica: evidence that dephosphorylation of adenosylcobalamin-5'-phosphate by the CobC phosphatase is the last step of the pathway.
abstract: We report that cobC strains of Salmonella enterica serovar Typhimurium are impaired in the ability to salvage cobyric acid (Cby), a de novo corrin ring biosynthetic intermediate, under aerobic growth conditions. In vivo and in vitro evidence support the conclusion that this new phenotype of cobC strains is due to the inability of serovar Typhimurium to dephosphorylate adenosylcobalamin-5'-phosphate (AdoCbl-5'-P), the product of the condensation of alpha-ribazole-5'-phosphate (alpha-RP) and adenosylcobinamide-GDP by the AdoCbl-5'-P synthase (CobS, EC 2.7.8.26) enzyme. Increased flux through the 5,6-dimethylbenzimidazole and cobinamide (Cbi) activation branches of the nucleotide loop assembly pathway in cobC strains restored AdoCbl-5'-P synthesis from Cby in a cobC strain. The rate of the CobS-catalyzed reaction was at least 2 orders of magnitude higher with alpha-RP than with alpha-ribazole as substrate. On the basis of the data reported herein, we conclude that removal of the phosphoryl group from AdoCbl-5'-P is the last step in AdoCbl biosynthesis in serovar Typhimurium and that the reaction is catalyzed by the AdoCbl-5'-P phosphatase (CobC) enzyme. Explanations for the correction of the Cby salvaging phenotype are discussed.
authors.affiliation: Department of Bacteriology, University of Wisconsin, 144A Enzyme Institute, 1710 University Avenue, Madison, WI 53726-4087, USA.
authors.forename: Carmen L
authors.initials: CL
authors.lastname: Zayas
authors.forename: Jorge C
authors.initials: JC
authors.lastname: Escalante-Semerena
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003038:Cobamides / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism*; D003470:Culture Media; D004251:DNA Transposable Elements; D013058:Mass Spectrometry; D016254:Mutagenesis, Insertional; D010710:Phosphates / Q000378:metabolism*; D012486:Salmonella typhimurium / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D000637:Transaminases / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D003038:Cobamides; D003470:Culture Media; D004251:DNA Transposable Elements; D010710:Phosphates; C079403:CobC protein, Bacteria; C033344:cobamamide 5'-phosphate; D000637:Transaminases; C000913:cobamamide