BIOSYNTHESIS OF VITAMIN B12 AND ANAEROBIC METABOLISM

沙门氏菌将其1%的基因组用于合成维生素B12(钴胺)，这是一种由沙门氏菌从头合成的大辅助因子(MW 1570)，但不是大肠杆菌。另外1%的沙门氏菌基因组编码需要B12的代谢途径(包括乙醇胺和丙二醇的降解)。我们建议在B12的合成、进口和使用的几个方面进行工作，包括B12的低配体(二甲基苯并咪唑)的来源。我们有参与转运的周质钴胺结合蛋白的遗传证据，并有突变表明运输蛋白可能有助于抑制COB(生物合成)操纵子，而不仅仅是重要的效应器。对B12依赖的降解途径的分析揭示了几个意想不到的特征。(1)B12的生理重要性可能是厌氧的(这是制造B12的唯一条件)。依赖钴胺的厌氧生长在丙二醇和乙醇胺上与电子受体四硫酸盐发生，而不与富马酸或硝酸盐发生。我们将继续研究四硫酸盐还原的遗传学、生物化学和调控，并试图了解为什么这种受体是独一无二的，以及它在自然界中出现在哪里。(2)非常大的操纵子编码乙醇胺(EUT；17个基因)和丙二醇(PDU；多于20个基因)使用的酶。(3)这些操纵子的大部分基因在所用条件下没有突变表型，表明它们在新的条件下发挥功能或对其他代谢功能是多余的。(4)这两个操纵子都编码与我们最近观察到的羧基体、细胞器的外壳蛋白同源的多个蛋白质。这表明EUT和PDU途径可能涉及二氧化碳固定。在高二氧化碳环境下生长的细胞有第二条利用乙醇胺的途径，该途径只需要17基因(Eut)操纵子产生的一个(EutE)酶；CO2-途径依赖于B12，但不需要B12依赖的酶乙醇胺解氨酶(EutBC)。我们正在描述这些路径的特征。我们认为B12代谢是沙门氏菌(一种病原体)与其姊妹种大肠杆菌的主要区别所在。虽然这些生物在实验室中看起来非常相似，但它们很容易在分类学上区分开来。合成B12(COB，CBI)、使用丙二醇(PDU)和还原多硫化物(TTR、PHS、ASR)都是区分沙门氏菌和大肠杆菌的特征。了解这种代谢可能有助于了解沙门氏菌的自然生活方式。

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.