IDENTIFICATION AND CHARACTERIZATION OF A BINDING-SITE FOR PLATELETS IN THE APPLE-3 DOMAIN OF COAGULATION-FACTOR-XI

Activated platelets expose a specific, reversible high affinity (K-dapp similar or equal to 10 nM) binding site (n similar or equal to 1500 sites/platelet platelet) for factor XI that requires the presence of high molecular weight kininogen (HK) and ZnCl2 (Greengard, J. S., Heeb, M. J., Ersdal, E., Walsh, P. N., and Griffin, J. H. (1986) Biochemistry 25, 3884-3890), Synthetic, conformationally constrained peptides from four tandem repeat (Apple) domains were tested for their capacity to inhibit I-125-factor XI binding to platelets, A peptide from the Apple 3 (A3) domain (Asn(235)-Arg(266)) inhibits factor XI binding to platelets in the presence of HK (42 nM), CaCl2, (2 mM), and ZnCl2 (25 mu M), with a K-i similar or equal to 10 nM which is identical to the K-d for factor XI binding to platelets, A peptide from the A1 domain (Phe(56)-Ser(86)) partially inhibits factor XI binding to platelets (Ki = 6 mu M) by inhibiting factor M binding to HK, whereas peptides from the A2 and A4 domains have no effect, Using computer modeling for rational design, conformationally constrained peptides were synthesized (Pro(229) Gln(233), Thr(241)-Leu(246), and Ser(248)-Ser(261)) each of which acted alone and synergistically when added together to inhibit factor XI: binding to platelets, Finally, the I-125-labeled A3 domain peptide (Asn(235)-Arg(266)) was found to bind to thrombin activated platelets in a specific, reversible, and saturable manner, Thus, the sequence of amino acids Asn(235)-Arg(266) Of the A3 domain of factor XI comprises a contact surface for interaction with a platelet receptor.