Sulfatide-dependent autoactivation of human blood coagulation Factor XII (Hageman Factor).

G Tans,J Rosing,J H Griffin

doi:10.1016/s0021-9258(20)82051-1

Abstract

When purified human blood coagulation Factor XII (Hageman factor) is incubated with sulfatides at 37 degrees C, activation of Factor XII occurs as judged by the appearance of amidolytic activity towards the chromogenic substrate H-D-Pro-Phe-Arg-p-nitroanilide. Polyacrylamide gel electrophoresis studies using 125I-Factor XII as a marker show that the appearance of amidolytic activity correlates with Factor XII cleavage, that activation goes to completion and that virtually all Factor XIIa formed is present as the two-chain, 80,000 Mr form, alpha-Factor XIIa. Rigorous analysis of kinetic data establishes that, between 0.02 and greater than 90% of the reaction, the activation of Factor XII is described by a mechanism of autoactivation of Factor XII by Factor XIIa. The rate of autoactivation increases with increasing Factor XII concentrations at constant sulfatide levels but decreases with increasing sulfatide concentrations at constant levels of Factor XII. These findings suggest that the concentrations of Factor XII and Factor XIIa bound to the sulfatide surface determine the rate of autoactivation. Soybean trypsin inhibitor, Trasylol, and anti-prekallikrein antibodies have no influence on the rate of sulfatide-dependent autoactivation of Factor XII. Benzamidine inhibits autoactivation with an inhibitor constant, Ki, of 1.9 mM which is similar to the Ki of 1.5 mM for the enzyme, alpha-Factor XIIa. Thus, sulfatide-dependent activation of purified Factor XII is not due to contaminating proteases and is described by a second order mechanism of autoactivation due to the action of surface-bound Factor XIIa on surface-bound Factor XII.

Highlights

From the Department of Immunology, Scripps Clinic and Research Foundation, La Jolla, California92037 and the $l)epartment of Biochemistry, University of Limburg, Maastricht, The Netherlands
Genic substrate H-D-Pro-Phe-Arg-p-nitroanilideP.ol- Factor XII, has been suggested to be capable of activating yacrylamide gel electrophoresis studies usin'g251-Fa~- its own zymogen, Factor XII, in thepresence of glass, kaolin, tor XI1 as a marker show that the appearanceof ami- or dextran sulfate [3,4,5,6,7], enhancing the rate at which dolytic activity correlates with Factor XI1 cleavage, Factor XII, will be formed during the early stagesof contact that activation goes to completion and that virtually allFactor XII, formedispresent as the two-chain, 80,000 M, form, a-Factor XII
The rateof autoactivation increases with Cerebroside sulfates(sulfatides) provide a very effective increasing Factor XI1 concentrations a t constant sul- surface for the reactions taking place during contact activafatide levels but decreases with increasing sulfatide tion [8, 9]

Summary

Introduction

It was not possible to establishif the reaction from 90% up to 100% was fit according to this mechanism since the experiment, a constant amountof Factor XI1 (14 pg/ml) underwent activation in the presence of varying amounts of sulfatides. T h e K , of inhibition of autoactivation by benzamidinewasdetermined.Fig. 7 A showstheeffect of The experiments presented in this paper show that purified benzamidine on sulfatide-dependent autoactivationof Factor human Factor XI1 is rapidly activated upon incubation with

Results

Conclusion