BIOSYNTHESIS OF VITAMIN B12 AND ANAEROBIC METABOLISM

维生素 B12 的生物合成和无氧代谢

• 批准号: 2902588
• 负责人: JOHN R ROTH
• 金额: $ 37.97万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2902588
• 关键词: Salmonella typhimurium; anaerobiosis; bacterial genetics; carbon dioxide; cobalt; ethanolamines; gene expression; iron; microorganism metabolism; mutant; operon; propylene glycols; transposon /insertion element; vitamin B12; vitamin biosynthesis

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.

沙门氏菌将其1％的基因组献给了维生素B12（钴胺素）的合成大型辅助因子（MW 1570），由沙门氏菌（Salmonella）从头开始，而不是大肠杆菌。 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综合导入和使用的几个方面进行工作，包括B12的下配体的来源（Dimethylbenzimidazole）。 我们有遗传证据表明涉及转运的周质钴胺素结合蛋白，并具有突变体，表明转运蛋白可能有助于抑制COB（生物合成）操纵子，而不是效应子的重要性。 B12依赖性降解途径的分析揭示了几个意外的特征。 （1）B12的生理重要性可能是厌氧（唯一制造B12的条件）。 电子受体四硫酸盐对丙二醇和乙醇胺的钴胺依赖性厌氧生长发生，而不是富马酸或硝酸盐。 我们将追求遗传学，生物化学和减少四硫酸盐的调节，并试图了解为什么该受体是独一无二的以及它在自然界中的位置。 （2）非常大的操纵子编码用于使用乙醇胺（EUT； 17个基因）和丙二醇（PDU；大于20基因）的酶。 （3）这些操纵子的大多数基因在所使用的条件下没有突变表型，表明它们在新的条件下起作用或对其他代谢功能是多余的。 （4）两个操纵子编码多种蛋白质与羧化体的壳蛋白同源的细胞器蛋白，我们最近已经看到了这些蛋白质。 这表明EUT和PDU途径可能涉及二氧化碳固定。 用高二氧化碳生长的细胞具有乙醇胺使用的第二个途径，该途径仅需要17基因（EUT）操纵子产生的一种（EUTE）酶。二氧化碳途径取决于B12，但不需要B12依赖性酶乙醇胺氨基酶（EUTBC）。 我们正在表征这些途径。 我们认为B12代谢是沙门氏菌（一种病原体）及其姐妹大肠杆菌之间的主要差异。 尽管这些生物在实验室中看起来非常相似，但它们在分类学上很容易区分。 B12（COB，CBI）的合成，丙二醇（PDU）的使用和降低多硫化物（TTR，PHS，ASR）的合成都是将沙门氏菌与大肠杆菌区分开的特性。 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。