IDENTIFICATION OF THE ZINC-DEPENDENT ENDOTHELIAL-CELL BINDING-PROTEINFOR HIGH-MOLECULAR-WEIGHT KININOGEN AND FACTOR-XII - IDENTITY WITH THE RECEPTOR THAT BINDS TO THE GLOBULAR HEADS OF C1Q (GC1Q-R)

High molecular weight kininogen (HK) and factor XII are known to bind to human umbilical vein endothelial cells (HUVEC) in a zinc-dependent and saturable manner indicating that HUVEC express specific binding si te(s) for those proteins. However, identification and immunochemical c haracterization of the putative receptor site(s) has not been previous ly accomplished. In this report, we have identified a cell surface gly coprotein that is a likely candidate for the HK binding site on HUVECs . When solubilized HUVEC membranes were subjected to an HK-affinity co lumn in the presence or absence of 50 mu M ZnCl2 and the bound membran e proteins eluted, a single major protein peak was obtained only in th e presence of zinc. SDS/PAGE analysis and silver staining of the prote in peak revealed this protein to be 33 kDa and partial sequence analys is matched the NH2 terminus of gC1q-R, a membrane glycoprotein that bi nds to the globular ''heads'' of C1q. Two other minor proteins of appr oximate to 70 kDa and 45 kDa were also obtained. Upon analysis by West ern blotting, the 33-kDa band was found to react with several monoclon al antibodies (mAbs) recognizing different epitopes on gC1q-R. Ligand and dot blot analyses revealed zinc-dependent binding of biotinylated HK as well as biotinylated factor XII to the isolated 33-kDa HUVEC mol ecule as well as recombinant gC1q-R. In addition, binding of I-125-HK to HUVEC cells was inhibited by selected monoclonal anti-gC1q-R antibo dies. C1q, however, did not inhibit I-125-HK binding to HUVEC nor did those monoclonals known to inhibit C1q binding to gC1q-R. Taken togeth er, the data suggest that HK (and factor XII) bind to HUVECs via a 33- kDa cell surface glycoprotein that appears to be identical to gC1q-R b ut interact with a site on gC1q-R distinct from that which binds C1q.