Crude extracts of Desulfovibrio vulgaris reduced sulfite to sulfide. Ammonium sulfate fractionation of crude extracts separated a thiosulfate-forming system from sulfiteand thiosulfate-reductase activities. Further purification by sucrose density centrifugation separated the thiosulfate-forming system into two components, both of which were required for the reaction. In addition to these two components, cytochrome c3, ferredoxin, and hydrogenase were required to form thiosulfate from sulfite. By absorption spectra and from the effect of pH and substrate concentration, the ionic species acting as the substrate for thiosulfate-formation was concluded to be bisulfite.

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Bisulfite reductase was purified from extracts of Desulfovibrio vulgaris. By colorimetric analyses trithionate was found to be the major product, being formed in quantities 5 to 10 times more than two other detectable products, thiosulfate and sulfide. When [35 ]bisulfite was used as the substrate, all three products were radioactively labeled. Degradation of [35Sltrithionate showed that all of its sulfur atoms were equally labeled. In contrast, [35S]thiosulfate contained virtually all of the radioactivity in the sulfonate atom while the sulfane atom was unlabeled. These results, in conjunction with the finding that the sulfide was radioactive, led to the conclusion that bisulfite reductase reduced bisulfite to trithionate as the major product and sulfide as the minor product; the reason for the unusual labeling pattern found in the thiosulfate molecule was not apparent at this time. When bisulfite reductase was incubated with [35S]bisulfite in the presence of another protein fraction, FII, the thiosulfate formed from this reaction contained both sulfur atoms having equal radioactivity. This discovery, plus the fact that trithionate was not reduced to thiosulfate under identical conditions, led to the speculation that bisulfite could be reduced to thiosulfate by another pathway not involving trithionate. 733 on August 6, 2020 by guest

An enzyme that formed thiosulfate from bisulfite and trithionate was purified from extracts ofDesulfovibrio vulgaris. This enzyme, designated as "thiosulfateforming" enzyme, required the presence of both bisulfite and trithionate. Various 35S-labeling studies showed that thiosulfate was formed from bisulfite and the inner sulfur atom of trithionate. This involved a nucleophilic attack by the bisulfite ion, resulting in the displacement of the two outer sulfonate groups of trithionate that recycled to participate as free bisulfite in subsequent reactions.This reaction required a reduction, presumably by a concerted mechanism with thiosulfate formation. The natural electron carrier cytochrome C3 participated in this reductive formation ofthiosulfate. This reaction was coupled to the bisulfite reductase-catalyzed reaction, which resulted in the reconstruction of a thiosulfate-forming pathway from bisulfite.

The fatty acid composition of two thermophilic anaerobes was determined, and the results were compared with those from a mesophilic and a psychrophilic anaerobe. Notable differences were that the thermophiles contained a higher content of saturated straight- and branched-chain fatty acids, and, of the latter, iso C 15 was the predominant type. The mesophile and psychrophile were characterized by having a higher percentage of unsaturated fatty acids. An unidentified fatty acid, present in all of the organisms, was purified from the psychrophile. By physical and chemical analysis the structure of the unknown acid was resolved and found to be the unsaturated cyclopropane fatty acid, 12,13-methylene-9-tetradecenoic acid.