Domain 3 of kininogens contains a cell-binding site and a site that modifies thrombin activation of platelets.
High and low molecular weight kininogens (HK and LK) are able to bind to platelets to inhibit thrombin binding to and activation of platelets. The heavy chain domain on the kininogens that contains these functions has been determined. Domain 3 (D3) but not domains 1 or 2, completely inhibited 125I-HK binding to platelets (Ki = 24 +/- 7 nM, n = 4). 125I-D3 specifically bound to unstimulated platelets and human umbilical vein endothelial cells. On platelets, it was blocked by unlabeled D3 and HK but not prekallikrein, factor XII, C1s, or C1 inhibitor. Further, one monoclonal antibody (HKH13) directed to kininogens' D3 blocked 125I-HK and 125I-D3 binding to platelets. The binding of 125I-D3 to platelets was fully reversible by addition of 35 molar excess of unlabeled D3. D3 binding to platelets was saturable with an apparent Kd of 39 +/- 8 nM (n = 4) and 1227 +/- 404 binding sites/platelet. D3, like HK and LK, inhibited thrombin-induced platelet activation by preventing thrombin binding to platelets. Another monoclonal antibody (HKH12), directed to D3, which did not block HK binding to platelets, reduced HK's ability to inhibit 125I-alpha-thrombin binding. This result suggests that the region on D3 that inhibits 125I-alpha-thrombin binding to platelets is different from that which directly binds to platelets. These studies indicate that D3 of the kininogens contains both a binding region for platelets and endothelial cells and another region that inhibits thrombin-induced platelet activation.
Jiang
Y P
YP
Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.
Muller-Esterl
W
W
Schmaier
A H
AH
D000818:Animals; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D002417:Cattle; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000166:cytology; D006801:Humans; D007457:Iodine Radioisotopes; D007704:Kininogens / Q000378:metabolism*; D008970:Molecular Weight; D015539:Platelet Activation / Q000187:drug effects*; D010974:Platelet Aggregation; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D007457:Iodine Radioisotopes; D007704:Kininogens; D013917:Thrombin