Activation of human plasma prekallikrein: Influence of activators, activation time and temperature and inhibitors

Abstract

Plasma prekallikrein (PKK) was activated with Cephotest and dextran sulphate (DS) under various conditions and the resulting kallikrein (KK) assayed on a chromogenic peptide substrate for this enzyme. Cephotest: plasma ratios of 30:1 and DS concentrations of between 0.001 and 0.01% (equal volumes plasma and DS) gave maximal activities when activation proceeded at 0°C. However, enzyme activity was also dependent on incubation time and temperature and activation times of 3 minutes (Cephotest) and 15 minutes (DS) at 0°C gave highest activities. Incubation at 37°C gave progressively lower KK yields with Cephotest and very little KK activity was detected with DS at this temperature. These results were shown to be due to progressive inhibition of plasma KK at 37°C. With C 1-esterase inhibitor deficient plasma higher KK activities were obtained with both activators at 0°C and the loss of activity on incubation at 37°C was markedly reduced. The addition of pure C 1-esterase inhibitor to the deficient plasma prior to activation resulted in a normal activity and normal inhibition profile. Gel filtration studies on normal and C 1-esterase deficient plasmas after activation at 0°C with Cephotest revealed that both free KK and KK- α 2 macroglobulin complexes were present in the activated plasmas. In the deficient plasma both activities were higher indicating that in normal plasma some KK is immediately bound by α 2-macroglobulin and C 1-esterase inhibitor even when activation takes place at 0°C.