a IIb b 3 , a member of the integrin family of adhesive protein receptors, is the most abundant glycoprotein on platelet plasma-membranes and binds to adhesive proteins via the recognition of short amino acid sequences, for example the ubiquitous RGD motif. However, elucidation of the ligandbinding domains of the receptor remains controversial, mainly owing to the fact that integrins are conformationally labile during purification and storage. In this study, a detailed mapping of the extracellular region of the a IIb subunit is presented, using overlapping 20-peptides, in order to identify the binding sites of a IIb potentially involved in the platelet-aggregation event. Regions a IIb 313-332, a IIb 265-284 and a IIb 57-64 of a IIb b 3 were identified as putative fibrinogen-binding domains because the corresponding peptides inhibited platelet aggregation and antagonized fibrinogen association, possibly by interacting with this ligand. The latter is further supported by the finding that the above peptides did not interfere with the binding of PAC-1 to the activated form of a IIb b 3 . Furthermore, a IIb 313-332 was found to bind to fibrinogen in a solid-phase binding assay. It should be emphasized that all the experiments in this study were carried out on activated platelets and consequently on the activated form of this integrin receptor. We hypothesize that RAD and RAE adhesive motifs, encompassed in a IIb 313-332, 265-284 and 57-64, are capable of recognizing complementary domains of fibrinogen, thus inhibiting the binding of this ligand to platelets.Keywords: a IIb -binding domains; a IIb mapping; plateletaggregation inhibitors; a IIb b 3 receptor; integrin inhibitors.The integrin family of adhesive protein receptors, composed of noncovalently associated a and b subunits, participates in a number of diverse functions ranging from embryogenesis to cellular aggregation, and differentiation to tumor cell growth and metastasis [1][2][3][4][5]. Integrin receptors consist of at least 20 members composed of different combinations of a and b subunits with distinct ligand-recognition specificity [6].The integrin receptor a IIb b 3 is the most abundant glycoprotein on platelet plasma-membranes. This receptor binds to adhesive proteins, such as fibrinogen, von Willebrand factor, fibronectin, and vitronectin, via the recognition of short amino acid sequences, including the ubiquitous motif RGD, as well as the HHLGGAKQAGDV sequence of the fibrinogen c-chain [7,8]. Binding studies suggest that platelet activation (e.g. by ADP) induces conformational changes of a IIb b 3 , which result in higher affinity to fibrinogen, an event essential for platelet aggregation and thrombus formation [9,10]. mAbs recognizing specific epitopes on the extracellular domains of both subunits are also able to induce/stabilize conformational changes of a IIb b 3 , which increase the affinity of the receptor for its ligands [11][12][13].The discovery that the RGD sequence is present in a surprisingly large number of adhesive proteins, serving div...
