Role of Active Site Residues and Weak Noncovalent Interactions In Substrate Positioning in N,N-Dimethylformamidase

N,N-Dimethylformamide (DMF) is a pollutant that can be metabolized naturally by DMF-utilizing microorganisms, via the non-heme iron enzyme N,N-dimethylformamidase (DMFase). We investigated the unusual protein fold and Tyr-Tyr-Glu iron coordination sphere of DMFase by performing molecular dynamics simulations and studying DMF binding and substrate positioning. Our docking studies support that the DMF substrate directly coordinates the iron center through its carbonyl group, with Fe-DMF distances consistent with structures of inorganic complexes. DMF binding is predominantly stabilized by weak noncovalent interactions with nearby phenylalanines, which also serve to control access of solvent to the active site based on cavity analysis of crystal structures and MD snapshots. Based on noncovalent interactions sampled in our simulations and sequence conservation, we ascribe roles to active site residues E657, H519, N547, F611, and F693. We perform sequence and structural alignments to identify putative DMFases and active site geometries in protein structures predicted from metagenomic DNA. These analyses suggest common conserved residues amongst putative DMFases and relates them to catalytic function providing guidance for future experimental studies or characterization of new DMFases for DMF bioremediation.