The conserved active site histidine-glutamate pair of ferrochelatase coordinately catalyzes porphyrin metalation

Ferrochelatase catalyzes the insertion of ferrous iron into protoporphyrin IX to generate heme. Despite recent research on the reaction mechanism of ferrochelatase, the precise roles and localization of individual active site residues in catalysis, particularly those involved in the insertion of the ferrous iron into the protoporphyrin IX substrate, remain controversial. One outstanding question is from which side of the macrocycle of the bound porphyin substrate is the ferrous iron substrate inserted. Pre-steady state kinetic experiments done under single-turnover conditions conclusively demonstrate that metal ion insertion is pH-dependent, and that the conserved active site His-Glu pair coordinately catalyzes the metal ion insertion reaction. Further, pKa calculations and molecular dynamic simulations indicate that the active site His is deprotonated and the protonation state of the Glu relates to the conformational state of ferrochelatase. Specifically, the conserved Glu in the open conformation of fer...