Isolation and properties of human neutrophil myeloperoxidase

Abstract

Human leukocyte myeloperoxidase has been purified to homogeneity by a three-step procedure which includes dialysis of a granule extract against low-salt buffer. Sephadex G-75 chromatography, and carboxymethylcellulose chromatography. The final product was homogeneous when examined by acid polyacrylamide gel electrophoresis and sedimentation equilibrium ultracentrifugation. The molecular weight determined by the latter procedure was 118000. With or without reduction of the protein by 2-mercaptoethanol, subunits were formed which migrated as a single band after sodium dodecyl sulfate gel electrophoresis. With reduction, the molecular weight of the apparently identical subunits was 59000, and 42000 without reduction. Other general properties of human leukocyte myeloperoxidase, including amino acid composition, amino terminal sequence analysis, and absorption spectra, are also reported. Myeloperoxidase, in the presence of hydrogen peroxide and chloride ion, and no other substrate, autoinactivates. After completion of the inactivation reaction, several oxidizable amino acids in the enzyme are modified, and the absorption peak at 430 nm disappears. The presence of a substrate of the myeloperoxidase system (alpha-1-proteinase inhibitor), or of high concentration of chloride ion, completely protects the enzyme from autoinactivation.