Biosynthesis of cobalamin in Salmonella typhimurium : transformation of riboflavin into the 5,6-dimethylbenzimidazole moiety

Keck, Bettina; Munder, Michael; Renz, Paul

doi:10.1007/s002030050679

Cited by 27 publications

(30 citation statements)

References 12 publications

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“…It was reported that wild-type S. enterica synthesizes pseudo-B 12 during growth under strictly anaerobic conditions and standard B 12 under microaerobic conditions (21,22). Nutritional experiments suggested that wild-type S. enterica uses a ligand other than DMB during growth on propanediol.…”

Section: Conditions That Prevent Synthesis or Installation Of Any ␣-Amentioning

confidence: 99%

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B ₁₂ Cofactors in Salmonella enterica

et al. 2008

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Coenzyme B 12 (Ado-Cbl) and vitamin B 12 (CN-B 12 ) (Fig. 1) include the base 5,6-dimethylbenzimidazole (DMB) as an ␣-axial ligand. Prokaryotes, the only producers of vitamin B 12 , make "complete" corrinoid cofactors with a variety of alternative ␣-axial ligands, including benzimidazoles, phenols, and purines (16, 52). These alternative corrinoids may be functional equivalents of coenzyme B 12 for many bacterial enzymes, but functional differences are suggested by the selectivity of the human assimilatory protein, intrinsic factor for cobalamin, the DMB-containing vitamin B 12 (53).Under strictly anaerobic growth conditions, Salmonella enterica synthesizes corrinoids de novo (44) and installs either adenine (to form pseudo-coenzyme B 12 [Ado-pseudo-B 12 ]) or 2-methyl-adenine (to form adenosyl-factor A) as an ␣-axial ligand (22). If even trace amounts of oxygen are present, DMB-containing B 12 (cobalamin [Cbl]) is also made (39). When grown aerobically on glucose, S. enterica cannot synthesize the corrin moiety, which must be supplied as an "incomplete" corrinoid, such as cobinamide (Cbi), with its corrin ring and aminopropanol side chain. Under these conditions, coenzyme B 12 is made, with DMB as the ␣-axial ligand (20). Figure 2 diagrams the de novo (anaerobic) synthetic pathway and assimilation of exogenous Cbi or . Notice that the ␤-ligand adenosyl can be added (by CobA) to three different substrates and that the (CN) 2 Cbi supplied as the corrin ring source is not a normal CobA substrate.Here, the origins and installation of ␣-axial ligands are approached genetically using an unexpected feature of S. enterica. While S. enterica can use exogenous Cbi to produce B 12 cofactors (cobalamins) during aerobic growth on glucose (20), it makes only about 100 molecules per cell (2). This is apparently insufficient B 12 coenzyme to support growth on ethanolamine (5) unless the DMB base is also supplied. That is, under these conditions, cells neither make sufficient DMB nor install an alternative ligand to allow corrinoid-dependent aerobic growth on ethanolamine (5).This situation allowed positive selection of mutants that can grow on ethanolamine plus Cbi without added DMB. These mutants were expected to show either increased endogenous DMB production or to install an alternative base as an ␣-axial ligand. All of the mutants made pseudo-B 12 cofactors, which have adenine as an ␣-axial ligand, and most of these mutations affected purine metabolism so as to increase the intracellular level of free adenine base. The same set of enzymes (CobUSTC) installs either adenine base (to form pseudo-B 12 ) or DMB (to form vitamin B 12 ). A model suggests how the choice is made. MATERIALS AND METHODSBacterial strains and transposons. Strains were derived from S. enterica (serovar Typhimurium) LT2

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Section: Conditions That Prevent Synthesis or Installation Of Any ␣-Amentioning

confidence: 99%

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B ₁₂ Cofactors in Salmonella enterica

et al. 2008

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“…DMB synthesis in some anaerobes appears to branch from the purine biosynthetic pathway, although enzymes involved in this process have not been isolated (11,18). Interestingly, S. enterica synthesizes the corrin ring anaerobically and derives DMB aerobically from flavin mononucleotide (FMN), although no ortholog of BluB appears to exist in the genome (1,12).…”

mentioning

confidence: 99%

“…The observation that S. enterica produces B 12 [DMB], but that no DMB biosynthetic enzyme has been identified in S. enterica, has puzzled researchers for many years (23). Anderson et al explored this issue by selecting for mutants of S. enterica that grow under conditions that normally require exogenous DMB (1).…”

mentioning

confidence: 99%

“…Anderson et al explored this issue by selecting for mutants of S. enterica that grow under conditions that normally require exogenous DMB (1). One key observation that made this selection possible is that traditional agar contains traces of DMB, and thus, much of the DMB in the B 12 [DMB] produced by S. enterica can be attributed to its presence in agar. Using purified agar enabled the authors to study only the endogenously produced corrinoids.…”

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confidence: 99%

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Pseudo-B ₁₂ Joins the Cofactor Family

Taga

Walker

2008

J Bacteriol

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“…This facultative anaerobic enterobacterium can salvage incomplete corrinoids [e.g. cobyric acid (Cby) and cobinamide (Cbi)] from its environment and converts them into AdoCba under both aerobic and anaerobic conditions (Brushaber et al, 1998;Jeter et al, 1984;Keck et al, 1998). The resulting AdoCba functions as coenzyme for 1,2-propanediol dehydrase and ethanolamine ammonia-lyase enzymes in this bacterium, and is the source of the non-adenosylated cobamide used by methionine synthase (Babior, 1982;Taylor & Weissbach, 1973;Toraya & Fukui, 1982).…”

Section: Introductionmentioning

confidence: 99%

The last step in coenzyme B12 synthesis is localized to the cell membrane in bacteria and archaea

2004

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In Salmonella enterica, the last step of the synthesis of adenosylcobamide is catalysed by the cobalamin synthase enzyme encoded by the cobS gene of this bacterium. Overexpression of the S. enterica cobS gene in Escherichia coli elicited the accumulation of the phage shock protein PspA, a protein whose expression has been linked to membrane stress. Resolution of inner and outer membranes of S. enterica by isopycnic density ultracentrifugation showed CobS activity associated with the inner membrane, a result that was confirmed using antibodies against CobS. Computer analysis of the predicted amino acid sequence of CobS suggested it was an integral membrane protein.Results of experiments performed with strains carrying plasmids encoding CobS-alkaline phosphatase or CobS-b-galactosidase protein fusions were consistent with the membrane localization of the CobS protein. Modifications to the predicted model were made based on data obtained from experiments using protein fusions. The function encoded by the cobS orthologue in the methanogenic archaeon Methanobacterium thermoautotrophicum strain DH compensated for the lack of CobS during cobalamin synthesis in cobS strains of S. enterica. Cobalamin synthase activity was also detected in a membrane preparation of M. thermoautotrophicum. It was concluded that the assembly of the nucleotide loop of adenosylcobamides in archaea and bacteria is a membrane-associated process. Possible reasons for the association of adenosylcobamide biosynthetic enzymes with the cell membrane are discussed.

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Biosynthesis of cobalamin in Salmonella typhimurium : transformation of riboflavin into the 5,6-dimethylbenzimidazole moiety

Cited by 27 publications

References 12 publications

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B ₁₂ Cofactors in Salmonella enterica

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B ₁₂ Cofactors in Salmonella enterica

Pseudo-B ₁₂ Joins the Cofactor Family

The last step in coenzyme B12 synthesis is localized to the cell membrane in bacteria and archaea

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Biosynthesis of cobalamin in Salmonella typhimurium : transformation of riboflavin into the 5,6-dimethylbenzimidazole moiety

Cited by 27 publications

References 12 publications

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B 12 Cofactors in Salmonella enterica

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B 12 Cofactors in Salmonella enterica

Pseudo-B 12 Joins the Cofactor Family

The last step in coenzyme B12 synthesis is localized to the cell membrane in bacteria and archaea

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One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B ₁₂ Cofactors in Salmonella enterica

One Pathway Can Incorporate either Adenine or Dimethylbenzimidazole as an α-Axial Ligand of B ₁₂ Cofactors in Salmonella enterica

Pseudo-B ₁₂ Joins the Cofactor Family