Crystallographic Snapshots of the Dunathan and Quinonoid Intermediates provide Insights into the Reaction Mechanism of Group II Decarboxylases

[Display omitted] •MjDC is a PLP-dependent amino acid decarboxylase in Methanocaldococcus jannaschii.•Pre- and post-decarboxylation intermediate complexes of MjDC were determined.•A conserved Arg residue in conjunction with a water dictates reaction specificity.•Protonation of quinonoid intermediate involves an extended hydrogen bond network.•Structural basis of mechanistic features in fold type-I decarboxylases elucidated. PLP-dependent enzymes catalyze a plethora of chemical reactions affecting diverse physiological functions. Here we report the structural determinants of the reaction mechanism in a Group II PLP-dependent decarboxylase by assigning two early intermediates. The in-crystallo complexes of the PLP bound form, and the Dunathan and quinonoid intermediates, allowed direct observation of the active site interactions. The structures reveal that a subtle rearrangement of a conserved Arg residue in concert with a water-mediated interaction with the carboxylate of the Dunathan intermediate, appears to directly stabilize the alignment and facilitate the release of CO2 to yield the quinonoid. Modeling indicates that the conformational change of a dynamic catalytic loop to a closed form controls a conserved network of hydrogen bond interactions between catalytic residues to protonate the quinonoid. Our results provide a structural framework to elucidate mechanistic roles of residues that govern reaction specificity and catalysis in PLP-dependent decarboxylation.