Oxidation of Sulfur and Inorganic Sulfur Compounds in the Thermoacidophilic Archaeon Acidianus ambivalens

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The thermophilic and acidophilic Archaeon Acidianus ambivalens oxidizes elemental sulfur to sulfuric acid as electron donor and energy source when growing aerobically. We purified several enzymes of the oxidation pathway recently including a soluble sulfur oxygenase reductase (SOR), a membrane-bound thiosulfate:quinone oxidoreductase (TQO) and a soluble tetrathionate hydrolase (TTH). In addition, we determined the 3D structure of the SOR. The goal of this proposal is to elucidate the reaction mechanisms of each of the three enzymes in molecular detail. With the SOR we plan to use a combination of site-directed mutagenesis, spectroscopy, rapid reaction kinetics of reaction intermediates, and X-ray crystallography of mutant proteins. With the TQO we want to establish gene expression, to do a systematic mutagenesis survey, and try to solve the 3D structure. With the TTH we want to identify the gene, establish gene expression and do a thorough biochemical characterization. With all three enzymes we will try to identify yet unknown inhibitors as probes for the active site structure.