title: Human high molecular weight kininogen binds to human umbilical vein endothelial cells via its heavy and light chains.
abstract: High molecular weight kininogen (HK) is a multifunctional plasma glycoprotein that occupies a critical position in pathways that link inflammation and coagulation. Excision of the vasoactive peptide bradykinin by plasma kallikrein results in kinin-free HK that consists of a 65-Kd N-terminal heavy chain (HK-HC) linked to the C-terminal 45-Kd light chain (HK-LC) by a disulfide bridge. HK-HC is an inhibitor of SH-proteases and HK-LC contains the binding sites for coagulation cofactors prekallikrein and factor XI. HK has previously been shown to bind specifically to human umbilical vein endothelial cells (HUVEC) in a zinc(2+)-dependent manner by a single class of high-affinity binding sites. We have further characterized that interaction in order to determine the cell-binding regions of HK. Competition binding experiments have indicated that either HK-LC or HK-HC was able to inhibit the binding of labeled HK with a 50% inhibitory concentration (IC50) of 77 nmol/L and 89 nmol/L, respectively. Cleaved two-chain HK (HKa) had an IC50 of 73 nmol/L, whereas uncleaved HK had an IC50 of 335 nmol/L. Direct binding experiments have indicated that HUVEC bind both purified [125I]HK-HC and [125I]HK-LC in a zinc(2+)-dependent manner and that HK-LC did not displace bound HK-HC. The light chain of low molecular weight kininogen or prekallikrein-binding region of HK did not inhibit the binding of HK to HUVEC. Our results, therefore, indicate that (1) HK is capable of binding to endothelial cells via both heavy and light chain moieties, (2) HKa has a higher affinity to HUVEC, and (3) purified heavy and light chains are capable of directly binding to HUVEC. The data are consistent with the presence of a single high-affinity site for HK on endothelial cells within which are subsites that bind to heavy and light chains.
authors.affiliation: Department of Medicine, State University of New York, Stony Brook 11794.
authors.forename: S R
authors.initials: SR
authors.lastname: Reddigari
authors.forename: P
authors.initials: P
authors.lastname: Kuna
authors.forename: G
authors.initials: G
authors.lastname: Miragliotta
authors.forename: Y
authors.initials: Y
authors.lastname: Shibayama
authors.forename: K
authors.initials: K
authors.lastname: Nishikawa
authors.forename: A P
authors.initials: AP
authors.lastname: Kaplan
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D006801:Humans; D007704:Kininogens / Q000737:chemistry / Q000378:metabolism*; D008970:Molecular Weight; D014471:Umbilical Veins; D015032:Zinc / Q000494:pharmacology
chemical_list: D007704:Kininogens; D015032:Zinc