Studies on the Structure and Function of Myeloperoxidase from Normal Human Leukocytes

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1. Myeloperoxidase was purified form normal human leukocytes, and it was first crystallized. The crystalline enzyme contained three components, which were isolated homogeneously by cation-exchange chromatograbhy.2. These three components were investigated on their molecular weight, molecular shape, subunit structure, light absorption, circular dichroism, magnetic circular dichroism, and amino acid composition. The enzyme consisted of two large subunits and two small subunits, and the three components were different in their molecular weight of the large subunits.3. The hemienzyme of myeloperoxidase was prepared by alkylation after reduction. The activity of the hemienzyme was not changed. The sedimentation-diffusion and small-angle X-ray scattering experiments showed that the molecular shape of the holo- and hemi-enzymes were more spherical than the values reported before.4 Two kinds of subunits were isolated by chromatography after the reduction of the enzyme in the presence of guanidine hydrochloride, and the amino acid sequences around their N- and C-termini. By comparing these sequences with those deduced from the cDNA base sequences for the precursor of myeloperoxidase, the processing part of the precursor by cellular proteinase was determined. Moreover, the green heme was found to be bound on the large subunit protein by covalent bonding.5. Two intermediate compounds of myeloperoxidase formed by the addition of hydrogen peroxide, and their life times and stoichiometry were investigated. In the course of the reaction, true catalase activity of the enzyme was confirmed. The hemienzyme has the same reaction characteristics of the holo-enzyme.