Human Hageman factor (factor XII) and high molecular weight kininogen compete for the same binding site on human umbilical vein endothelial cells.
Factor XII (FXII, plasma concentration 375 nM) is a critical member of the plasma contact activation system and is the zymogen form of FXIIa, a serine protease involved in intrinsic coagulation, complement activation, activation of factor VII, and generation of the vasoactive peptide bradykinin. As such its interaction with cells involved in inflammatory pathways can be of physiologic and pathologic significance. We have studied the binding of FXII to cultured human umbilical vein endothelial cells (HUVEC). HUVEC were incubated with 125I-FXII, and cell-bound factor FXII was measured. FXII bound to HUVEC saturably in a zinc-dependent manner. The optimal zinc concentration was 50-60 microM. Binding of labeled FXII was drastically reduced when a 200-fold molar excess of unlabeled FXII was included in the incubation mixture at time zero or when added at 60 min during a 150-min time course experiment. Quantitative binding experiments indicated a dissociation constant of 144 nM with 10-12 million binding sites/endothelial cell. Unlabeled high molecular weight kininogen (HK) inhibited the binding of labeled FXII with a Ki of 98 nM, whereas unlabeled FXII inhibited the binding of labeled HK to HUVEC with a Ki of 152 nM. SDS-polyacrylamide gel electrophoresis and autoradiography of cell-bound 125I-FXII showed that factor XII underwent limited proteolysis and the molecular weights of the fragments were similar in size to activated FXII. The cell-bound activated factor XII was also able to activate prekallikrein. These data suggest that (i) FXII binds to HUVEC specifically, saturably, and reversibly in a zinc-dependent manner, (ii) HK and FXII may compete with each other for the same cell-surface receptor/s, and (iii) cell-bound FXII is capable of undergoing activation to FXIIa.
Reddigari
S R
SR
Department of Medicine, State University of New York, Stony Brook 11794-8161.
Shibayama
Y
Y
Brunnée
T
T
Kaplan
A P
AP
D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000494:pharmacology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D004789:Enzyme Activation; D005174:Factor XII / Q000378:metabolism*; D006801:Humans; D007457:Iodine Radioisotopes; D007700:Kinetics; D007704:Kininogens / Q000378:metabolism* / Q000494:pharmacology; D008970:Molecular Weight; D011288:Prekallikrein / Q000378:metabolism; D014471:Umbilical Veins
D007457:Iodine Radioisotopes; D007704:Kininogens; D005174:Factor XII; D011288:Prekallikrein; D002118:Calcium