Characterization of a protein from normal human polymorphonuclear leukocytes with bactericidal activity against Pseudomonas aeruginosa.
Purification of a bactericidal protein (BP) from the cytoplasmic granules of normal human polymorphonuclear leukocytes (PMN) with activity against Pseudomonas aeruginosa is described. Bactericidal activity from acid extracts of a mixed granule population was purified 175-fold by a two-step chromatographic procedure. The first step, dye-ligand affinity chromatography with Matrex-Gel Orange A, was followed by cation-exchange chromatography with Bio-Rex 70 resin, and this combination routinely gave a yield near 80%. Only one peak of bactericidal activity against P. aeruginosa type I was found after each chromatographic step. Characterization of BP showed a single band with an apparent molecular weight of 55,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Purified BP was most active against the six strains of P. aeruginosa tested and against Escherichia coli B (a deep rough mutant). Purified BP killed 5 X 10(6) CFU of P. aeruginosa type I per ml at a concentration of 60 to 80 ng/ml. Proteus mirabilis and Staphylococcus aureus were both resistant to the bactericidal activity of BP. BP was shown to be glycosylated by periodic acid staining after gel electrophoresis and to have an isoelectric point near 7.5 by chromatofocusing. The amino acid composition of BP is presented.
D000596:Amino Acids / Q000032:analysis; D023181:Antimicrobial Cationic Peptides; D001770:Blood Bactericidal Activity; D001798:Blood Proteins / Q000032:analysis*; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D006801:Humans; D008565:Membrane Proteins; D008970:Molecular Weight; D009504:Neutrophils / Q000032:analysis / Q000276:immunology*; D011550:Pseudomonas aeruginosa
D000596:Amino Acids; D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D008565:Membrane Proteins; C021623:bactericidal permeability increasing protein