MECHANISM OF AUTOTROPHIC GROWTH WITH CO OR CO2 AND H2

CO 或 CO2 和 H2 的自养生长机制

• 批准号: 3272653
• 负责人: HARLAND G. WOOD
• 金额: $ 16.11万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-04-01 至 1991-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3272653
• 关键词: Clostridium; Mossbauer spectrometry; acetates; acetyl coA; bioenergetics; carbon dioxide; carbon monoxide; chemical binding; circular dichroism; coenzyme A; electron nuclear double resonance spectroscopy; electron spin resonance spectroscopy; enzyme mechanism; enzyme structure; hydrogen; methyl group; methyltransferase; microorganism metabolism; molecular cloning; nuclear magnetic resonance spectroscopy; protein structure; tetrahydrofolates

These investigations have as the ultimate objective elucidation of the pathway by which certain bacteria grow with CO or CO2 and H2 as the source of carbon and energy. This is a pathway of autotrophic growth which differs from any previously described. It involves reduction of CO2 to methyltetrahydrofolate, then combination of this methyl group with CO and CoASH to form acetyl-CoA which is then used as the source of carbon for anabolic growth. The synthesis of acetyl-CoA from methyltetrahydrofolate, CO and CoASH is the unique portion of this pathway. Homogeneous proteins have been isolated from Clostridium thermoaceticum which catalyze this synthesis. They are CO dehydrogenase methyltransferase, a corrinoid enzyme, ferredoxin, and disulfide reductase. The CO dehydrogenase is the master enzyme of this pathway. It forms the C1 intermediate from CO or CO2 that is the precursor of the carbonyl group of acetyl-CoA, is the acceptor of the methyl from the methyl corrinoid enzyme and through the action of the CO dehydrogenase reductase combines with CoA. It then catalyzes the condensation of these three groups to acetyl-CoA. Our main objective is to elucidate how CO dehydrogenase catalyzes this condensation. We will use electron spin resonance (ESR), electron-nuclear double resonance, Mossbauer, and magnetic circular dichroism techniques to characterize the geometry and redox state of the Ni-Fe-S center of the CO dehydrogenase. We will investigate the mechanism of transfer of the methyl group from the methylated corrinoid protein to CO dehydrogenase with formation of an acetyl intermediate. We hope to detect this methyl-CO dehydrogenase complex using ESR techniques and chemically identify the amino acid sequence at the binding site. Likewise, the sequence near the linkage of CoASH to the enzyme, presumed to be by a disulfide bond, will be investigated. The monomers of the Alpha3Beta3 enzyme will be isolated, their metal content determined and genes for these proteins will be cloned and sequenced. The properties of CO dehydrogenase from certain methanogenic bacteria will be tested for their ability to synthesize acetyl-CoA to determine whether they may use a pathway of acetyl-CoA synthesis similar to that of C. thermoaceticum.

这些调查最终客观地阐明了 某些细菌以CO或CO2和H2为来源的生长途径 碳和能源。这是一条自养生长的途径， 与之前描述的任何一个都不同。它涉及将二氧化碳减少到 甲基四氢叶酸，然后将该甲基与CO和 Coash生成乙酰CoA，然后用作 合成代谢生长。由甲基四氢叶酸合成乙酰辅酶A， CO和Coash是这条途径的独特部分。均一蛋白质 从催化该酶的热醋酸梭菌中分离得到 综合。它们是一氧化碳脱氢酶甲基转移酶，一种皮质激素 酶、铁还蛋白和二硫键还原酶。一氧化碳脱氢酶是 这一途径的主酶。它由CO或CO2形成C1中间体 即乙酰辅酶A的前体是羰基，是受体 从甲基皮质醇酶中获得甲基，并通过 CO脱氢酶还原酶与辅酶A结合。然后，它催化了 这三个基团缩合成乙酰辅酶A。我们的主要目标是 阐明CO脱氢酶是如何催化这种缩合的。我们将使用 电子自旋共振(ESR)，电子-核双共振， 穆斯堡尔谱和磁圆二向色性技术来表征 CO脱氢酶Ni-Fe-S中心的几何构型和氧化还原状态我们 将研究甲基转移的机制 甲基化皮质醇蛋白合成一氧化碳脱氢酶 乙酰中间体。我们希望能检测到这种甲基一氧化碳脱氢酶 用ESR技术和化学方法鉴定氨基酸 在结合位点处测序。同样，在连接的附近的序列 与酶的结合，推测是通过二硫键，将是 调查过了。Alpha3Beta3酶的单体将被分离， 它们的金属含量将被测定，这些蛋白质的基因将被克隆 并进行了测序。某些来源的一氧化碳脱氢酶的性质 产甲烷细菌将被测试其合成能力 乙酰-辅酶A来确定它们是否可能使用乙酰辅酶A途径 与热醋酸菌相似的合成。