X-RAY-ABSORPTION AND RESONANCE RAMAN-SPECTROSCOPY OF HUMAN MYELOPEROXIDASE AT NEUTRAL AND ACID PH

Myeloperoxidase (MPO), an important enzyme in the oxygen-dependent hos t defense system of human polymorphonuclear leukocytes, utilizes hydro gen peroxide to catalyze the production of hypochlorous acid, an oxidi zing bactericidal agent. While MPO shows significant sequence homology with other peroxidases and this homology is particularly striking amo ng the active-site residues, MPO exhibits unusual spectral features an d the unique ability to catalyze the oxidation of chloride ions. We ha ve investigated the MPO active-site with X-ray absorption (XAS) and re sonance Raman (RRS) spectroscopies at neutral pH and also at the physi ological acidic pH (pH similar to 3) and have compared these results w ith these of horseradish peroxidase (HRP). At pH 7.5, XAS results show that the iron heme active site is 6-coordinate where the distal ligan d is likely nitrogen or oxygen, but not sulfur. The heme is distorted compared to HRP, other peroxidases, and heme compounds, but at pH simi lar to 3, the distal ligand is lost and the heme is less distorted. RR S results under identical pH conditions show that the skeletal core-si ze sensitive modes and nu(3) are shifted to higher frequency at pH sim ilar to 3 indicating a 6- to 5-coordination change of high spin ferric heme. In addition, a new band at 270 cm(-1) is observed at pH similar to 3 which is consistent with the loss of the sixth ligand. The highe r symmetry of the heme at pH similar to 3 is reflected by a single nu( 4) mode in the (RRS) spectrum. HRP also loses its loosely associated d istal water at this pH, but little change in heme distortion is observ ed. This change suggests that loss of the distal ligand in MPO release s stress on the heme which may facilitate binding of chloride ion.