Deletion Strains Reveal Metabolic Roles for Key Elemental Sulfur-Responsive Proteins in Pyrococcus furiosus

Transcriptional and enzymatic analyses of Pyrococcus furiosus previously indicated that three proteins play key roles in the metabolism of elemental sulfur (S0): a membrane-bound oxidoreductase complex (MBX), a cytoplasmic coenzyme A-dependent NADPH sulfur oxidoreductase (NSR), and sulfur-induced protein A (SipA). Deletion strains, referred to as MBX1, NSR1, and SIP1, respectively, have now been constructed by homologous recombination utilizing the uracil auxotrophic COM1 parent strain (ΔpyrF). The growth of all three mutants on maltose was comparable without S0, but in its presence, the growth of MBX1 was greatly impaired while the growth of NSR1 and SIP1 was largely unaffected. In the presence of S0, MBX1 produced little, if any, sulfide but much more acetate (per unit of protein) than the parent strain, demonstrating that MBX plays a critical role in S0 reduction and energy conservation. In contrast, comparable amounts of sulfide and acetate were produced by NSR1 and the parent strain, indicating that NSR is not essential for energy conservation during S0 reduction. Differences in transcriptional responses to S0 in NSR1 suggest that two sulfide dehydrogenase isoenzymes provide a compensatory NADPH-dependent S0 reduction system. Genes controlled by the S0-responsive regulator SurR were not as highly regulated in MBX1 and NSR1. SIP1 produced the same amount of acetate but more sulfide than the parent strain. That SipA is not essential for growth on S0 indicates that it is not required for detoxification of metal sulfides, as previously suggested. A model is proposed for S0 reduction by P. furiosus with roles for MBX and NSR in bioenergetics and for SipA in iron-sulfur metabolism.