Abstract
Heterotrophic bacteria and human mitochondria often use sulfide: quinone oxidoreductase (SQR) and persulfide dioxygenase (PDO) to oxidize sulfide to sulfite and thiosulfate. Bioinformatic analysis showed that the genes encoding RHOD domains were widely presented in annotated sqr-pdo operons and grouped into three types: fused with an SQR domain, fused with a PDO domain, and dissociated proteins. Biochemical evidence suggests that RHODs facilitate the formation of thiosulfate and promote the reaction between inorganic polysulfide and glutathione to produce glutathione polysulfide. However, the physiological roles of RHODs during sulfide oxidation by SQR and PDO could only be tested in an RHOD-free host. To test this, 8 genes encoding RHOD domains in Escherichia coli MG1655 were deleted to produce E. coli RHOD-8K. The sqr(Cp) and pdo(Cp) genes from Cupriavidus pinatubonensis JMP134 were cloned into E. coli RHOD-8K. SQR(Cp) contains a fused RHOD domain at the N-terminus. When the fused RHOD domain of SQR(Cp) was inactivated, the cells oxidized sulfide into increased thiosulfate with the accumulation of cellular sulfane sulfur in comparison with cells containing the intact sqr(Cp) and pdo(Cp). The complementation of dissociated DUF442 minimized the accumulation of cellular sulfane sulfur and reduced the production of thiosulfate. Further analysis showed that the fused DUF442 domain modulated the activity of SQR(Cp) and prevented it from directly passing the produced sulfane sulfur to GSH. Whereas, the dissociated DUF442 enhanced the PDO(Cp) activity by several folds. Both DUF442 forms minimized the accumulation of cellular sulfane sulfur, which spontaneously reacted with GSH to produce GSSG, causing disulfide stress during sulfide oxidation. Thus, RHODs may play multiple roles during sulfide oxidation.