Structural and mechanistic basis of sulfolytic C–S bond cleavage by an Fe(II)/α-ketoglutarate-dependent sulfoquinovose dioxygenase

Abstract

Sulfoquinovose dioxygenase (SqoD) enables bacterial carbon assimilation from the abundant sulfosugar sulfoquinovose (SQ) by Fe(II)/α-ketoglutarate (αKG)–dependent C–S bond cleavage. Here we report crystal structures of the Marinobacterium aestuarii enzyme (MaSqoD) in multiple states with inert Mn²⁺ in place of Fe²⁺ (SQ-bound; Mn²⁺•αKG; Mn²⁺•αKG•SQ; Mn²⁺•succinate), together with steady-state and pre–steady-state kinetics that link the structures with kinetically-inferred intermediates. X-ray crystal structures of SQ-bound, Mn²⁺•αKG, Mn²⁺•αKG•SQ, and Mn²⁺•succinate states show a canonical 2-His-1-carboxylate core metal center with SQ recognition via a mainly neutral network (Gln120, Trp253, backbone carbonyl of Ala185, and backbone amides of Ala89/Met118). Substrate binding triggers a hexacoordinate octahedral-to-pentacoordinate change at the metal center, unveiling a vacant site for O₂ in the fully assembled Mn²⁺•αKG•SQ complex and thereby curbing uncoupled reactions. Pre-steady-state stopped-flow data support the canonical Fe(IV)=O chemistry of the dioxygenase and reveal an additional intermediate consistent with an enzyme-bound α-hydroxysulfonate or 6-dehydroglucose species. Together, these findings define the structural and mechanistic basis of sulfolytic SQ catabolism and expand the functional repertoire of Fe(II)/αKG-dependent dioxygenases into carbon assimilation.