A Novel Peptide Motif for Platelet Fibrinogen Receptor Recognition*

To develop a specific antagonist of platelet αIIbβ3 using small linear peptides, we synthesized a series of hexapeptides that did not have an Arg-Gly-Asp (RGD) sequence and examined their anti-platelet activity and their specificity for αIIbβ3. We found a novel motif sequence, Pro-X1-X2-X3-Asp-X4, where X1 to X4 were all L-form α-amino acids, which specifically inhibited aggregation of human platelets at submicromolar concentrations. The Pro residue at the N terminus was essential to the anti-platelet activity, and the acetylation of the imino group of this residue also resulted in the complete loss of the activity. The results of the binding assay using purified human platelet αIIbβ3 and placental αvβ3 and those of the cell adhesion assay suggest that this motif peptide is highly specific for platelet αIIbβ3 among other integrins. Flow cytometric studies using an fluorescein isothiocyanate-labeled RGD peptide showed that this motif peptide inhibited the binding of an RGD peptide to activated platelets, suggesting that it has the same inhibitory mode as RGD peptides. Conformational analysis of this motif peptide and an RGD-containing peptide suggests that the imino group of the Pro residue may substitute for the role of the guanidino group of the Arg residue of the RGD sequence.

[1]  Y. Hayashi,et al.  Design and synthesis of new antagonist peptides for platelet GPIIb/IIIa receptor as anti-thrombotic agents , 1996 .

[2]  R. Califf,et al.  Effects of platelet glycoprotein IIb/IIIa receptor blockade by a chimeric monoclonal antibody (abciximab) on acute and six-month outcomes after percutaneous transluminal coronary angioplasty for acute myocardial infarction. EPIC investigators. , 1996, The American journal of cardiology.

[3]  Gail Murphy,et al.  Pharmacokinetics and pharmacodynamics of MK‐383, a selective non‐peptide platelet glycoprotein‐IIb/IIIa receptor antagonist, in healthy men , 1994, Clinical pharmacology and therapeutics.

[4]  J. Loftus,et al.  Mutation of a ligand binding domain of beta 3 integrin. Integral role of oxygenated residues in alpha IIb beta 3 (GPIIb-IIIa) receptor function. , 1994, The Journal of biological chemistry.

[5]  T. Connolly,et al.  Disagregin is a fibrinogen receptor antagonist lacking the Arg-Gly-Asp sequence from the tick, Ornithodoros moubata. , 1994, The Journal of biological chemistry.

[6]  J. Tschopp,et al.  Design and synthesis of novel cyclic RGD-containing peptides as highly potent and selective integrin alpha IIb beta 3 antagonists. , 1994, Journal of medicinal chemistry.

[7]  R. Califf,et al.  Profound inhibition of platelet aggregation with monoclonal antibody 7E3 Fab after thrombolytic therapy. Results of the Thrombolysis and Angioplasty in Myocardial Infarction (TAMI) 8 Pilot Study. , 1993, Journal of the American College of Cardiology.

[8]  M. Ishiyama,et al.  A New Sulfonated Tetrazolium Salt That Produces a Highly Water-Soluble Formazan Dye , 1993 .

[9]  D. Phillips,et al.  Design of potent and specific integrin antagonists. Peptide antagonists with high specificity for glycoprotein IIb-IIIa. , 1993, The Journal of biological chemistry.

[10]  C. E. Peishoff,et al.  Investigation of conformational specificity at GPIIb/IIIa: evaluation of conformationally constrained RGD peptides. , 1992, Journal of medicinal chemistry.

[11]  G. Vanhoof,et al.  Kininase activity in human platelets: cleavage of the Arg1-Pro2 bond of bradykinin by aminopeptidase P. , 1992, Biochemical pharmacology.

[12]  J. Cazenave,et al.  A new variant of Glanzmann's thrombasthenia (Strasbourg I). Platelets with functionally defective glycoprotein IIb-IIIa complexes and a glycoprotein IIIa 214Arg----214Trp mutation. , 1992, The Journal of clinical investigation.

[13]  Richard O. Hynes,et al.  Integrins: Versatility, modulation, and signaling in cell adhesion , 1992, Cell.

[14]  R. Timpl,et al.  Dynamic Forcing, a Method for Evaluating Activity and Selectivity Profiles of RGD (Arg‐Gly‐Asp) Peptides , 1992 .

[15]  A. Frelinger,et al.  A spontaneous mutation of integrin alpha IIb beta 3 (platelet glycoprotein IIb-IIIa) helps define a ligand binding site. , 1992, The Journal of biological chemistry.

[16]  R. Calvo,et al.  Development of a small RGD peptide fibrinogen receptor antagonist with potent antiaggregatory activity in vitro. , 1991, Journal of medicinal chemistry.

[17]  M. Humphries,et al.  The minimal essential sequence for a major cell type-specific adhesion site (CS1) within the alternatively spliced type III connecting segment domain of fibronectin is leucine-aspartic acid-valine. , 1991, The Journal of biological chemistry.

[18]  Kenneth M. Yamada,et al.  Adhesive recognition sequences. , 1991, The Journal of biological chemistry.

[19]  V. Fried,et al.  Barbourin. A GPIIb-IIIa-specific integrin antagonist from the venom of Sistrurus m. barbouri. , 1991, The Journal of biological chemistry.

[20]  T. V. D. van de Ven,et al.  Dynamic measurements of the platelet membrane glycoprotein IIb-IIIa receptor for fibrinogen by flow cytometry. I. Methodology, theory and results for two distinct activators. , 1991, Biophysical journal.

[21]  M. Humphries,et al.  The molecular basis and specificity of integrin-ligand interactions. , 1990, Journal of cell science.

[22]  A. Frelinger,et al.  A beta 3 integrin mutation abolishes ligand binding and alters divalent cation-dependent conformation. , 1990, Science.

[23]  D. Cheresh,et al.  Interaction of integrins alpha v beta 3 and glycoprotein IIb-IIIa with fibrinogen. Differential peptide recognition accounts for distinct binding sites. , 1990, The Journal of biological chemistry.

[24]  G. Chang,et al.  Macromodel—an integrated software system for modeling organic and bioorganic molecules using molecular mechanics , 1990 .

[25]  M. Ginsberg,et al.  The ligand binding site of the platelet integrin receptor GPIIb-IIIa is proximal to the second calcium binding domain of its alpha subunit. , 1990, The Journal of biological chemistry.

[26]  G. Chang,et al.  An internal-coordinate Monte Carlo method for searching conformational space , 1989 .

[27]  C. Debouck,et al.  Human immunodeficiency virus 1 protease expressed in Escherichia coli behaves as a dimeric aspartic protease. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[28]  J W Smith,et al.  The Arg-Gly-Asp binding domain of the vitronectin receptor. Photoaffinity cross-linking implicates amino acid residues 61-203 of the beta subunit. , 1988, The Journal of biological chemistry.

[29]  S. Lam,et al.  Localization of an Arg-Gly-Asp recognition site within an integrin adhesion receptor. , 1988, Science.

[30]  M. Poncz,et al.  Comparison of cDNA-derived protein sequences of the human fibronectin and vitronectin receptor alpha-subunits and platelet glycoprotein IIb. , 1987, Biochemistry.

[31]  E Ruoslahti,et al.  New perspectives in cell adhesion: RGD and integrins. , 1987, Science.

[32]  Richard O. Hynes,et al.  Integrins: A family of cell surface receptors , 1987, Cell.

[33]  R. Houghten,et al.  Inhibition of platelet function with synthetic peptides designed to be high-affinity antagonists of fibrinogen binding to platelets. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[34]  M. Horton,et al.  Monoclonal antibodies to osteoclastomas (giant cell bone tumors): definition of osteoclast-specific cellular antigens. , 1985, Cancer research.

[35]  U. Singh,et al.  A NEW FORCE FIELD FOR MOLECULAR MECHANICAL SIMULATION OF NUCLEIC ACIDS AND PROTEINS , 1984 .

[36]  E. Ruoslahti,et al.  [27] Arginine-glycine-aspartic acid adhesion receptors , 1987 .