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Item Structure based mechanistic studies on 2-ketopropyl coenzyme M oxidoreductase / carboxylase from Xanthobacter autotrophicus and [FeFe] hydrogenase from Clostridium pasteurianum(Montana State University - Bozeman, College of Letters & Science, 2007) Pandey, Arti Sharma; Chairperson, Graduate Committee: John W. Peters; Martin Lawrence (co-chair)X-ray crystallography was employed to probe the mechanism of the enzyme 2- ketopropyl coenzymeM oxidoreductase / carboxylase (2-KPCC). We were able to determine the enzyme structure in various catalytically relevant states, providing insights into substrate binding, intermediate stabilization, product formation and release. Structures of 2-KPCC were obtained with the substrate 2-ketopropyl coenzyme M (KCoM), product acetoacetate, 6-oxoheptanoic acid (OHA), 2-oxopropyl phosphonate (OPP), NADP+ and coenzymeM (CoM), the oxidized and reduced states. The binding sites for these ligands in relation to one another have led to important sights into the mechanism. CO2 binds at the base of a hydrophobic channel at the interface of a hydrophobic pocket and the substrate binding site. Acetoacetate binds at an alternate anion binding site, as revealed in the bicarbonate and CoM disulfide bound structures. The enolate intermediate can be stabilized by an Ala430 carbonyl stabilized water molecule as revealed in the OHA bound structure, at a site different from that in KCoM bound structure. Together, the structures reveal a mechanism of concerted attack of a CO2 molecule on the enolate intermediate formed by the nucleophilic attack of Cys82 on the C-S bond of KCoM.