title: Rat and human neutrophil N-formyl-peptide chemotactic receptors. Species difference in the glycosylation of similar 35-38 kDa polypeptide cores.
abstract: Rat and human neutrophil N-formyl-peptide chemotactic receptors were subjected to glycosidase and proteinase treatments to determine the extent and species differences of glycosylation and the carbohydrate requirement in the high-affinity ligand binding. N-Formyl-Nle-Leu-Phe-Nle-125I-Tyr-Lys was attached to rat and human neutrophils either before or after glycosidase and proteinase treatments, and the labelled receptors were solubilized after glutaraldehyde cross-linking and analysed by SDS/PAGE and autoradiography. Both the rat and human N-formyl-peptide chemotactic receptors contain only N-linked oligosaccharides, as demonstrated by their sensitivity to peptide N-glycosidase F (PNGase F) and resistance to O-glycanase treatment. The N-linked oligosaccharides seem to be of the complex type rather than the high-mannose or hybrid type and lack terminal sialic acid, as demonstrated by their resistance to endoglycosidases D and H and neuraminidase treatments. This sensitivity pattern was similar in both species, and the shift in the molecular size of the receptors to 35-38 kDa after PNGase F treatment occurred through one intermediate product, suggesting that both receptors contain a similar 35-38 kDa polypeptide core with two N-linked complex-type oligosaccharides, the heterogeneity of which is responsible for the species difference in receptor size. Papain treatment alone or followed by PNGase F produced in both species a 33-36 kDa membrane-bound fragment that was still able to bind the ligand, suggesting that the oligosaccharides are located on the approx. 2 kDa papain-cleavable polypeptide fragment of the receptors. The cleavage sites for both papain and PNGase F were hidden in occupied receptors, suggesting a conformational or topographical change in these upon ligand binding. Scatchard analyses and cross-linking experiments demonstrated that carbohydrates are not required for high-affinity ligand binding and that the 33-36 kDa membrane-bound papain fragment of both receptors contains the ligand-binding site.
authors.affiliation: Biocenter, University of Oulu, Finland.
authors.forename: J J
authors.initials: JJ
authors.lastname: Remes
authors.forename: U E
authors.initials: UE
authors.lastname: Petäjä-Repo
authors.forename: H J
authors.initials: HJ
authors.lastname: Rajaniemi
mesh_terms: D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases; D006801:Humans; D008970:Molecular Weight; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000378:metabolism*; D009504:Neutrophils / Q000378:metabolism*; D009844:Oligosaccharides / Q000032:analysis; D010206:Papain / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D013045:Species Specificity
chemical_list: D009844:Oligosaccharides; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic; D009240:N-Formylmethionine Leucyl-Phenylalanine; D006026:Glycoside Hydrolases; D010206:Papain