ISOLATION OF 2 POLYPEPTIDES COMPRISING THE NEUTROPHIL-IMMOBILIZING FACTOR OF HUMAN-LEUKOCYTES

  • 1 January 1983
    • journal article
    • research article
    • Vol. 48  (1) , 79-86
Abstract
Human leukocyte lysosomal polypeptides of MW 4000-5000, which constitute the neutrophil-immobilizing factor (NIF), were isolated from the 22,000 g supernate of sonicates of human neutrophils by filtration on Sephadex G-75. The larger (NIF-1) and smaller (NIF-2) of the polypeptides were resolved by filtration on Bio-Gel P6 and purified to homogeneity by sequential reverse-phase high performance liquid chromatography and paper electrophoresis. The results of analyses of amino acid composition indicated that NIF-1 and NIF-2 are distinct polypeptides composed of an apparent total of 41 and 38 amino acid, respectively. Both NIF polypeptides contain 1 cysteine and 1 methionine, lack isoleucine, tyrosine and phenylalanine and are rich in histidine and proline. The sequence of 20 of the amino-terminal amino acids of both NIF polypeptides is identical, but NIF-2 possesses an additional alanine at the amino-terminus. Highly purified NIF-1 and NIF-2 inhibited human neutrophil random migration and chemotaxis to diverse stimuli in a concentration-dependent manner, with 50% inhibition of chemotaxis by 0.3-1 .times. 10-8 M NIF-1 and 1-3 .times. 10-7 M NIF-2. Neither NIF polypeptide was cytotoxic for neutrophils, altered neutrophil phagocytosis or release of lysosomal enzymes or inhibited mononuclear leukocyte chemotaxis. The leukocyte and functional specificity of the NIF polypeptides and the quantities released upon stimulation of human leukocytes suggest that the transition to a mononuclear leukocyte population in chronic inflammation may be attributable in part to the NIF derived from the leukocyte infiltrates of acute responses.