Specific binding, internalization, and degradation of human neutrophil activating factor by human polymorphonuclear leukocytes.
The interaction of 125I-labeled recombinant human neutrophil activating factor (NAF) with polymorphonuclear leukocytes (PMN) was studied by means of a radioreceptor assay. The binding was characterized by a rapid transition (t1/2 less than or equal to 1 min) from a pH 3-sensitive state at 4 degrees C to pH 3 resistance at 37 degrees C. This was not caused by internalization of NAF since pH 3-resistant bound iodinated NAF could still be exchanged by an excess of nonlabeled NAF, i.e. was dissociable. Internalized iodinated NAF was processed into trichloroacetic acid-soluble forms. Scatchard transformation of binding isotherms at 4 and 37 degrees C led to nonlinear curves, a finding which is consistent with the expression of two receptor populations, one with high (KD = 11-35 pM) and the other with lower affinity (KD = 640-830 pM) at 4 degrees C. Numbers of the low affinity binding sites were approximately 34,000, and those with high affinity were 5,200/PMN when estimated at 4 degrees C. Binding of iodinated NAF to PMN was specific since it could be competed by an excess of nonlabeled NAF but not by two other activators of PMN function, formylmethionyl-leucyl-phenylalanine or human recombinant granulocyte-macrophage colony-stimulating factor. In addition to human PMN, NAF also bound specifically to two human monocytic cell lines; however, only the low affinity binding site could be detected on these cells.
Besemer
J
J
Sandoz Forschungsinstitut, Vienna, Austria.
Hujber
A
A
Kuhn
B
B
D001665:Binding Sites; D001667:Binding, Competitive; D002630:Chemotactic Factors; D006801:Humans; D006863:Hydrogen-Ion Concentration; D016209:Interleukin-8; D007457:Iodine Radioisotopes; D007700:Kinetics; D009000:Monocytes / Q000378:metabolism; D009504:Neutrophils / Q000378:metabolism*; D010455:Peptides / Q000097:blood*; D011869:Radioligand Assay; D011994:Recombinant Proteins / Q000097:blood; D013696:Temperature; D014407:Tumor Cells, Cultured
D002630:Chemotactic Factors; D016209:Interleukin-8; D007457:Iodine Radioisotopes; D010455:Peptides; D011994:Recombinant Proteins