Self-S-sulfonation in a bacterial persulfide dioxygenase mediates thiol persulfide detoxification

A ubiquitous class of non-heme Fe(II) enzymes, the persulfide dioxygenases (PDOs), provide protection against hydrogen sulfide (H2S) poisoning. The PDO in humans is a single-domain enzyme, while bacterial PDOs, such as CstB of Staphylococcus aureus, are often fused to a sulfurtransferase (rhodanese). Canonical PDOs cleave the S-S bond of glutathione persulfide (GSSH) to produce GSH and sulfite. In contrast, CstB converts two thiol persulfides (RSSH) to thiosulfate without releasing sulfite. Crystallographic structures of S. aureus CstB reveal that a Cys-Gly sequence (C201-G202) in a CstB-unique dynamic loop functions as a glutathione mimic, occupying one face of the hemifacial octahedral Fe(II) coordination site. We establish that this loop facilitates three distinct chemical transformations along the reaction pathway: (i) transfer of a persulfide sulfur from and RSSH substrate to C201, (ii) Fe(II)- and O2-dependent oxidation of the C201 persulfide to an S-sulfonate, and (iii) rhodanese-dependent conversion of the S-sulfonate to thiosulfate. The rhodanese active-site cysteine is ≈27 Å away from the Fe(II) active site in the PDO domain, and molecular dynamics simulations suggest a long-range shuttling of the S-sulfonate from one active site to the other. Self-S-sulfonation renders the enzyme promiscuous toward low-molecular-weight persulfides, ensuring rapid clearance of diverse reactive sulfur species, permitting S. aureus to harness their cytoprotective effects while avoiding cellular toxicity. Persulfide dioxygenases (PDOs) protect against hydrogen sulfide and reactive sulfur species (RSS) poisoning. The bacterial PDO from Staphylococcus aureus CstB is unique in its oxidative metabolism of organic thiol persulfides to thiosulfate and the free thiol, without the release of toxic sulfite. Here, the authors reveal that a dynamic loop in CstB facilitates three distinct chemical transformations, enabling efficient clearance of RSS that is independent of the specific thiol persulfide.