title: Effects of hybrid peptide analogs to receptor recognition domains on alpha- and gamma-chains of human fibrinogen on fibrinogen binding to platelets.
abstract: We synthesized a series of hybrid peptides that correspond to the gamma-chain dodecapeptide (400-411), variable numbers of glycine residues, and the RGDS peptide [Y-HHLGGAKQAGDV(G)nRGDS] to investigate the relationship of these receptor recognition domains of fibrinogen to platelet membrane glycoprotein IIb/IIIa. The tetrapeptide RGDS, the GRGDSPA peptide and the dodecapeptide inhibited binding of fibrinogen to GPIIb/IIIa by 50% (IC50) at concentrations of 17 +/- 1.6 microM, 15 +/- 2.1 microM, and 87 +/- 6.8 microM, respectively. The inhibitory effect of hybrid peptides increased as the number of glycine residues increased, plateauing with 9-11 glycine residues in hybrid peptide analogs, which had an IC50 of 0.68 +/- 0.14 microM. These hybrid peptides completely inhibited the binding of fibrinogen to activated platelets when used in sufficient concentrations. The peptide Y-HHLGGAKQAGDV(G)9RGDS blocked ADP-induced aggregation in citrated platelet-rich plasma with IC50 of 3.5 +/- 0.6 microM. When the peptide Y-HHLGGAKQAGDV(G)9RGDS was labeled with 125I to quantify its binding to platelets, maximal binding was observed within 30 min. The binding sites of the hybrid peptide were 43,600 molecules/platelet (Kd = 3.1 +/- 0.5 x 10(-7) M) to stimulated platelets and 12,500 molecules/platelet (Kd = 1.4 +/- 0.2 x 10(-7) M) to nonstimulated platelets. The hybrid peptides had the same binding affinity to platelets as fibrinogen and inhibited platelet function. Moreover, anti-GPIIb/IIIa antibody inhibited the binding of the labeled hybrid peptide to stimulated platelets. These results indicate that in the native fibrinogen molecule the presence of both RGD sequence or gamma-chain domain at optimal distances increased the binding affinity to GPIIb/IIIa.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: First Department of Internal Medicine, Yokohama City University School of Medicine, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Mohri
authors.forename: T
authors.initials: T
authors.lastname: Ohkubo
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D005340:Fibrinogen / Q000378:metabolism*; D005344:Fibrinopeptide A / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects
chemical_list: D009842:Oligopeptides; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; C027539:fibrinopeptides gamma; D005344:Fibrinopeptide A; D005340:Fibrinogen; C046617:arginyl-glycyl-aspartyl-serine