Crystal Structure of the Dioxygen-bound Heme Oxygenase from Corynebacterium diphtheriae.

HmuO, a heine oxygenase of Corynebacterium diphtheriae, catalyzes degradation of heme using the same mechanism as the mammalian enzyme. The oxy form of HmuO, the precursor of the catalytically active ferric hydroperoxo species, has been characterized by ligand binding kinetics, resonance Raman spectroscopy, and x-ray crystallography. The oxygen association and dissociation rate constants are 5 μM-1 s-1 and 0.22 s-1, respectively, yielding an 02 affinity of 21 μM-l, which is -20 times greater than that of mammalian myoglobins. However, the affinity of HmuO for CO is only 3–4-fold greater than that for mammalian myoglobins, implying the presence of strong hydrogen bonding interactions in the distal pocket of HmuO that preferentially favor 02 binding. Resonance Raman spectra show that the Fe—O2 vibrations are tightly coupled to porphyrin vibrations, indicating the highly bent Fe—O&mdsh;O geometry that is characteristic of the oxy forms of heme oxygenases. In the crystal structure of the oxy form the Fe—O—O angle is 110°, the O—O bond is pointed toward the heme α-meso-carbon by direct steric interactions with Gly-135 and Gly-139, and hydrogen bonds occur between the bound 02 and the amide nitro. gen of Gly-139 and a distal pocket water molecule, which is a part of an extended hydrogen bonding network that provides the solvent protons required for oxygen activation. In addition, the O—O bond is orthogonal to the plane [ABSTRACT FROM AUTHOR]