BIOSYNTHESIS OF VITAMIN B12 AND ANAEROBIC METABOLISM

沙门氏菌将其基因组的1%用于合成维生素B12（钴胺素），这是一种由沙门氏菌从头合成的大型辅因子（MW 1570），而不是大肠杆菌。另外1%的沙门氏菌基因组编码需要维生素B12的代谢途径（包括乙醇胺和丙二醇的降解）。我们对B12的合成、进口和利用的几个方面提出了建议，包括B12的下配体（二甲基苯并咪唑）的来源。我们有一种参与转运的质周钴胺结合蛋白的遗传证据，并且有突变表明转运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子。对b12依赖性降解途径的分析揭示了几个意想不到的特征。(1) B12在生理上的重要性可能是厌氧的（这是制造B12的唯一条件）。丙二醇和乙醇胺上依赖钴胺素的厌氧生长发生在电子受体四硫酸盐上，而不发生在富马酸盐或硝酸盐上。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特的，以及它在自然界中出现的地方。(2)非常大的操纵子编码用于乙醇胺（17个基因）和丙二醇（超过20个基因）的酶。(3)这些操纵子的大部分基因在所使用的条件下没有突变表型，这表明它们在新的条件下起作用或对其他代谢功能是冗余的。(4)这两个操纵子都编码了多个与羧基体、细胞器壳蛋白同源的蛋白质，我们最近已经看到了这些蛋白质。这表明eut和pdu通路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种途径利用乙醇胺，只需要17基因操纵子产生的一种（EutE）酶；二氧化碳途径依赖于B12，但不需要依赖于B12的酶乙醇胺解氨酶（EutBC）。我们正在描述这些途径。我们认为，B12代谢是沙门氏菌（一种病原体）和它的姊妹物种大肠杆菌之间主要差异的基础。虽然这些生物在实验室里看起来非常相似，但在分类学上很容易区分。B12的合成（cob, cbi），丙二醇的使用（pdu）和多硫化物的还原（ttr, phs, asr）都是用来区分沙门氏菌和大肠杆菌的特性。了解这种新陈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.