Institute of Multidisciprlinary Research for Advanced Materials, Tohoku University
○Masaki Unno Masao Ikeda-Saito
Heme oxygenase (HO) catalyzes the regiospecific oxidation of heme to biliverdin with concomitant liberation of CO and iron. In its catalytic cycle, HO first binds one equivalent of heme to form a ferric heme-HO complex. The first electron donated from the reducing substrate converts the heme iron to the ferrous state. Then O2 binds to reduced 5-coordinate heme to form a meta-stable oxy complex. One-electron reduction of the oxy ferrous form generates a ferric hydroperoxo complex, which self-hydroxylates the alpha meso-carbon of the porphyrin ring. The latter reaction is different from what occurs in P450 enzymes, in which the O-O bond of the hydroperoxo complex is heterolytically cleaved to generate an actively hydroxylating, ferryl (Fe4+=O) intermediate. Ferric alpha meso-hydroxyheme is then converted to biliverdin by multiple oxidoreductive steps involving a verdoheme intermediate.
To understand the unique mechanism of HO catalysis at atomic level, we have engaged in crystallographic analysis of its reaction intermediates. Recently, we solved the structure of the hydroxyheme-bound HO from Corynebacterium diphtheriae at 1.5 Å resolution. The occupancy of hydroxyl group bound to alpha meso-carbon is approximately 0.9. This high reactive intermediate was trapped by X-ray induced photo reduction and crystal annealing techniques. The binding distance between alpha meso-carbon and O atom of hydroxyl group is 1.25 Å, indicative that the structure dominantly exists in a keto form.
In the conference, we report the detailed structure of the hydroxyheme-bound HO and how to trap such a high reactive intermediate in crystals with high occupancy. We also discuss the HO reaction mechanism.