Studies on the pathway for acetate conversion to methane by the methanoarchaeon Methanosarcina thermophila have identified enzymes commonly found in physiologically and metabolically diverse anaerobic microbes from the Bacteria domain. Three of these enzymes are virtually ubiquitous in anaerobes: acetate kinase, phosphotransacetylase, and carbonic anhydrases of the (and (classes. Acetate kinase and phosphotransacetylase play key roles in energy-yielding pathways, and the widespread occurrence of carbonic anhydrases suggests equally important functions for this enzyme. The sequences of these enzymes from M. thermophila have high identity to the enzymes from the Bacteria domain indicating common ancestors and mechanisms of catalysis. Despite their broad occurrence and importance in anaerobic procaryotes, very few mechanistic studies had been reported until the enzymes from M. thermophila were recently overproduced in Escherichia coli (the first for any of these enzymes) opening the way for modern methods of investigation. This application proposes to continue the investigation of these enzymes to understand the biochemical properties, focusing on the mechanisms of catalysis. The results are expected to contribute not only to a deeper understanding of the acetate fermentation pathway, but more importantly to the broader field of anaerobic microbiology and beyond.
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