The proposed study will be conducted with the acetogenic bacteria Clostridium thermoaceticum, Clostridium thermoautotrophicum and Clostridium formicoaceticum. The important feature of these bacteria is their capacity to synthesize acetate from carbon dioxide. This occurs via a new autotrophic pathway used by a number of anaerobic bacteria present in the intestine of man, many animals and insects. It is a major pathway in anaerobic environments and it is ecologically important. Bacteria using the pathway may in the intestine of man produce 13,000 metric tons of acetate per day from C02 in the total human population. The pathway is known as the acetyl-CoA pathway, because acetyl-CoA is the first 2-carbon compound formed from C02. The initial C02 fixations are catalyzed by formate dehydrogenase and an enzyme with several functions called carbon monoxide dehydrogenase/acetyl-CoA synthtase (CODH). C02 destined to form the methyl group of acetate is first reduced to formate, which is converted to 5-methyltetrahydrofolate and subsequently to methyl-B12. This involves several enzymes that are, tetrahydrofolate- and vitamin B12 dependent. Sequencing and active site studies will be performed. These experiments will give knowledge about enzymes converting 1-carbon compounds, which are needed for DNA, RNA and protein synthesis. C02 to form from the carboxyl group of acetate is first reduced to form carbon monoxide by CODH. This enzyme synthesizes from Co, methyl-B12 and coenzyme A acetyl-CoA. Formate dehydrogenase and CODH contains tungsten, selenium and iron and nickel, zinc and iron, respectively. The role of these elements in the catalytic processes will be studied using isotopes and spectrometric methods such as EPR, XAF, Raman, variable temperature magnetic circular dichroism (VTMCD) . These studies will increase our understanding on how trace elements function in biological reaction catalyzed by enzymes and increase our understanding of a new pathway of medical and ecological importance.
