One single hydrogen bond guarantees conformational stability and activity in coproheme decarboxylase from Corynebacterium diphtheriae

Active site architectures of enzymes are defined by many interactions between substrate and amino acid residues and are optimized for specific and efficient substrate turnover. In the case of coproheme decarboxylase (ChdC) the active site architecture is well described by structural and thermodynamic means. Coproheme decarboxylases transform iron coproporphyrin III (coproheme) into iron protoporphyrin IX (heme b) by oxidatively decarboxylating two propionate groups to vinyls. In this study we have investigated an arginine residue (R208, ChdC from Corynebacterium diphtheriae) in close proximity to propionate at position 2 (p2) that has been indicated to have an important steric role within the active site architecture. Here we focus on the molecular basis of its steric role and the catalytic consequences by investigating several R208 variants of coproheme decarboxylase from the Actinobacterium Corynebacterium diphtheriae. Analyses of the exchange of R208 into His, Lys, Glu, Asp, and Ser (serine mimics the situation of ChdCs in Firmicutes) help to deepen our understanding of this enzyme and its reaction mechanism. By employing experimental biochemical studies and molecular dynamics simulations we identify one single hydrogen bond of particular importance, proving that the protonation state matters and that R208 is an essential residue without having a direct mechanistic role during catalytic turnover.