title: In vitro effect of a thrombin inhibition peptide selected by phage display technology.
abstract: A repeated selection of phages from a cyclic heptapeptide phage display library resulted in the enrichment of phages that bind to human alpha-thrombin. One clone of the binding phages that competed with PPACK for binding to thrombin and that had the best binding characteristics was chosen. The amino acid sequence of the peptide displayed on this phage was determined and a peptide with the sequence, Cys-Asn-Arg-Pro-Phe-Ile-Pro-Thr-Cys was synthesised. This peptide, thrombin-inhibiting peptide (TIP), is a full competitive inhibitor of thrombin with an inhibition constant (K(i)) of 0.4974 mM. It lengthened the thrombin time and inhibited thrombin-induced platelet activation and the platelet release reaction, both in a dose-dependent manner. It also reduced platelet adhesion onto a human microvascular endothelial matrix in the parallel plate flow chamber under both arterial and venous shear conditions. Thus, we have selected and synthesised a cyclic heptapeptide that competes with PPACK to bind to thrombin and that can be developed as a direct antithrombin.
authors.affiliation: Department of Haematology and Cell Biology, University of the Free State, PO Box 339(G2), Bloemfontein 9300, South Africa. GNHMSMM@med.uovs.ac.za
authors.forename: Muriel S
authors.initials: MS
authors.lastname: Meiring
authors.forename: Derek
authors.initials: D
authors.lastname: Litthauer
authors.forename: Jolan
authors.initials: J
authors.lastname: Hársfalvi
authors.forename: Veronica
authors.initials: V
authors.lastname: van Wyk
authors.forename: Philip N
authors.initials: PN
authors.lastname: Badenhorst
authors.forename: Harry F
authors.initials: HF
authors.lastname: Kotzé
mesh_terms: D001435:Bacteriophages; D001483:Base Sequence; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D007118:Immunoassay; D019151:Peptide Library; D010455:Peptides / Q000494:pharmacology*; D010974:Platelet Aggregation / Q000187:drug effects; D018179:Receptors, Thrombin / Q000037:antagonists & inhibitors*; D013918:Thrombin Time
chemical_list: D019151:Peptide Library; D010455:Peptides; D018179:Receptors, Thrombin