Salmonella dedicates 1 percent of its genome to synthesis of vitamin B12 (cobalamin) a large cofactor (MW 1570) synthesized de novo by Salmonella, but not E. coli. Another 1 percent of the Salmonella genome encodes metabolic pathways (including degradation of ethanolamine and propanediol) that require the B12. We propose work on several aspects of B12 synthesis import and use, including the source of the lower ligand of B12 (dimethylbenzimidazole). We have genetic evidence for a periplasmic cobalamin binding protein involved in transport and have mutants that suggest the transport proteins may contribute to repression of the cob (biosynthetic) operon beyond mere important of the effector. Analysis of the B12-dependent degradative pathways has revealed several unexpected features. (1) Physiological importance of B12 is likely to be anaerobic (the only conditions under which B12 is made). Cobalamin- dependent anaerobic growth on propanediol and ethanolamine occurs with the electron acceptor tetrathionate, but not with fumarate or nitrate. We will pursue the genetics, biochemistry and regulation of tetrathionate reduction and will try to learn why this acceptor is unique and where it appears in nature. (2) Very large operons encode enzymes for use of ethanolamine (eut; 17 genes) and propanediol (pdu; greater than 20 genes). (3) Most of the genes of these operons have no mutant phenotype under the conditions used, suggesting that they function under novel conditions or are redundant to other metabolic functions. (4) Both operons encode multiple proteins homologous to the shell proteins of carboxysomes, organelles, which we have recently visualized. This suggests that the eut and pdu pathways could involve CO2 fixation. Cells grown with high CO2 have a second pathway for ethanolamine use which requires only one (EutE) enzyme produced by the 17-gene (eut) operon; the CO2-pathway depends on B12 but does not require the B12-dependent enzyme ethanolamine ammonia lyase(EutBC). We are characterizing these pathways. We feel that B12 metabolism underlies major differences between Salmonella (a pathogen) and its sister- species E. coli. While these organisms appear very similar in the lab, they are easily distinguished taxonomically. Synthesis of B12 (cob, cbi), use of propanediol (pdu) and reduction of polysulfide (ttr, phs, asr) are all properties used to distinguish Salmonella from E. coli. Understanding this metabolism may contribute to understanding the natural lifestyle of Salmonellae.

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National Institute of General Medical Sciences (NIGMS)
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Microbial Physiology and Genetics Subcommittee 2 (MBC)
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Ikeda, Richard A
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University of California Davis
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Anderson, Peter J; Lango, Jozsef; Carkeet, Colleen et al. (2008) One pathway can incorporate either adenine or dimethylbenzimidazole as an alpha-axial ligand of B12 cofactors in Salmonella enterica. J Bacteriol 190:1160-71
Penrod, Joseph T; Roth, John R (2006) Conserving a volatile metabolite: a role for carboxysome-like organelles in Salmonella enterica. J Bacteriol 188:2865-74
Carkeet, Colleen; Dueker, Stephen R; Lango, Jozsef et al. (2006) Human vitamin B12 absorption measurement by accelerator mass spectrometry using specifically labeled (14)C-cobalamin. Proc Natl Acad Sci U S A 103:5694-9
Price-Carter, Marian; Fazzio, Thomas G; Vallbona, Ester Ibanez et al. (2005) Polyphosphate kinase protects Salmonella enterica from weak organic acid stress. J Bacteriol 187:3088-99
Roth, John R; Andersson, Dan I (2004) Amplification-mutagenesis--how growth under selection contributes to the origin of genetic diversity and explains the phenomenon of adaptive mutation. Res Microbiol 155:342-51
Penrod, Joseph T; Mace, Christopher C; Roth, John R (2004) A pH-sensitive function and phenotype: evidence that EutH facilitates diffusion of uncharged ethanolamine in Salmonella enterica. J Bacteriol 186:6885-90
Sheppard, David E; Penrod, Joseph T; Bobik, Thomas et al. (2004) Evidence that a B12-adenosyl transferase is encoded within the ethanolamine operon of Salmonella enterica. J Bacteriol 186:7635-44
Slechta, E Susan; Bunny, Kim L; Kugelberg, Elisabeth et al. (2003) Adaptive mutation: general mutagenesis is not a programmed response to stress but results from rare coamplification of dinB with lac. Proc Natl Acad Sci U S A 100:12847-52
Bunny, Kim; Liu, Jing; Roth, John (2002) Phenotypes of lexA mutations in Salmonella enterica: evidence for a lethal lexA null phenotype due to the Fels-2 prophage. J Bacteriol 184:6235-49
Price-Carter, M; Tingey, J; Bobik, T A et al. (2001) The alternative electron acceptor tetrathionate supports B12-dependent anaerobic growth of Salmonella enterica serovar typhimurium on ethanolamine or 1,2-propanediol. J Bacteriol 183:2463-75

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