Enzymes produced by autoactivation of blood factor XII in buffer: A contribution from the Hematology at Biomaterial Interfaces Research Group.
High-resolution electrophoresis of FXII-derived proteins produced by contact activation of FXII in buffer solutions (i.e. in absence of plasma proteins) with hydrophilic and silanized-glass activators spanning the observable range of water wettability (hydrophilic to hydrophobic), shows no evidence of proteolytic cleavage of FXII into αFXIIa or βFXIIa. The autoactivation mixture contains only a single-chain protein with a molecular weight of ∼80 kDa, confirming Oscar Ratnoff's previous finding of a single-chain activated form of FXII that he called 'HFea'. Functional assays have shown that these autoactivation products exhibit procoagulant potential (protease activity inducing clotting of blood) or amidolytic potential (cleaves amino bonds in s-2302 chromogen but do not cause coagulation of plasma) or both amidolytic potential and procoagulant potential. Some of these proteins also have the remarkable potential to 'suppress autoactivation' (i.e. suppress creation of enzymes with procoagulant potential). It is thus hypothesized that autoactivation of FXII in the absence of plasma proteins generates not just a single type of activated conformer, as suggested by previous researchers, but rather an ensemble of conformer products with collective activity that varies with activator surface energy used in contact activation of FXII. Furthermore, reaction of αFXIIa with FXII in buffer solution does not produce additional αFXIIa by the putative autoamplification reaction FXIIa + FXII → 2FXIIa as has been proposed in past literature to account for the discrepancy between chromogenic and plasma-coagulation assays for αFXIIa in buffer solution. Instead, net procoagulant activity measured directly by plasma-coagulation assays, decreases systematically with increasing FXII solution concentration. Under the same reaction conditions, chromogenic assay reveals that net amidolytic activity increases with increasing FXII solution concentration. Thus, although autoamplification does not occur it appears that there is some form of "FXII self reaction" that influences products of αFXIIa reaction with FXII. Electrophoretic measurements indicate that no proteolytic cleavage takes in this reaction leading us to conclude that change in activity is most likely due to change(s) in FXII conformation (with related change in enzyme activity).
['Golas|Avantika|A|', 'Pitakjakpipop|Harit|H|', 'Rahn|Matthew S|MS|', 'Siedlecki|Christopher A|CA|', 'Vogler|Erwin A|EA|']
D001672:Biocompatible Materials D001777:Blood Coagulation D002021:Buffers D004586:Electrophoresis D004789:Enzyme Activation D005174:Factor XII D015956:Factor XIIa D006405:Hematology D006801:Humans D006868:Hydrolysis D019679:Kininogen, High-Molecular-Weight D011288:Prekallikrein D012106:Research D012996:Solutions D013997:Time Factors
Autoactivation FXII Conformers Autoamplification Autohydrolysis Plasma coagulation