Structural, biophysical, and biochemical insights into C-S bond cleavage by dimethylsulfone monooxygenase.

Abstract

Sulfur is an essential element for life. Bacteria can obtain sulfur from inorganic sulfate; but in the sulfur starvation-induced response, <i>Pseudomonads</i> employ two-component flavin-dependent monooxygenases (TC-FMOs) from the <i>msu</i> and <i>sfn</i> operons to assimilate sulfur from environmental compounds including alkanesulfonates and dialkylsulfones. Here, we report binding studies of oxidized FMN to enzymes involved within the <i>P. fluorescens</i> enzymatic pathway responsible for converting dimethylsulfone (DMSO₂) to sulfite. In this catabolic pathway, SfnG serves as the initial TC-FMO for sulfur assimilation, which is investigated in detail by solving the 2.6-Å resolution crystal structure of unliganded SfnG and the 1.75-Å resolution crystal structure of the SfnG ternary complex containing FMN and DMSO₂. We find that SfnG adopts a (β/α)₈ barrel fold with a distinct quaternary configuration from other tetrameric class C TC-FMOs. To probe the unexpected tetramer arrangement, structural heterogeneity is assessed by chromatography and light scattering to confirm ligand binding correlates with a tetramer. Binding of FMN and DMSO₂ accompanies ordering of the active site, with DMSO₂ bound on the <i>si</i>-face of the flavin. A previously unobserved protein backbone conformation is found within the oxygen-binding site on the <i>re</i>-face of the flavin. Functional assays and the positioning of ligands with respect to the oxygen-binding site are consistent with use of an N5-(hydro)peroxyflavin pathway. Biochemical endpoint assays and docking studies reveal SfnG breaks the C-S bond of a range of dialkylsulfones.