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Single‐chain factor XII exhibits activity when complexed to polyphosphate
Author(s) -
Engel R.,
Brain C. M.,
Paget J.,
Lionikiene A. S.,
Mutch N. J.
Publication year - 2014
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/jth.12663
Subject(s) - factor xii , chemistry , zymogen , cleave , prekallikrein , active site , biochemistry , polyphosphate , kallikrein , stereochemistry , biophysics , enzyme , coagulation , phosphate , biology , psychology , psychiatry
Summary Background The mechanism underpinning factor XII autoactivation was originally characterized with non‐physiological surfaces, such as dextran sulfate ( DS ), ellagic acid, and kaolin. Several ‘natural’ anionic activating surfaces, such as platelet polyphosphate (polyP), have now been identified. Objective To analyze the autoactivation of FXII by polyP of a similar length to that found in platelets (polyP 70 ). Methods and results PolyP 70 showed similar efficacy to DS in stimulating autoactivation of FXII , as detected with amidolytic substrate. Western blotting revealed different forms of FXII with the two activating surfaces: two‐chain α FXII a was formed with DS , whereas single‐chain FXII (sc FXII ; 80 kD a) was formed with polyP 70 . Dissociation of sc FXII from polyP 70 abrogated amidolytic activity, suggesting reversible exposure of the active site. Activity of sc FXII –polyP 70 was enhanced by Zn 2+ and was sensitive to NaCl concentration. A bell‐shaped concentration response to polyP 70 was evident, as is typical of surface‐mediated reactions. Reaction of sc FXII –polyP 70 with various concentrations of S2302 generated a sigmoidal curve, in contrast to a hyperbolic curve for α FXII a, from which a Hill coefficient of 3.67 was derived, indicative of positive cooperative binding. sc FXII –polyP 70 was more sensitive to inhibition by H‐ d ‐Pro‐Phe‐Arg‐chloromethylketone and corn trypsin inhibitor than α FXII a, but inhibition profiles for C1‐inhibitor were similar. Active sc FXII –polyP 70 was also able to cleave its physiological targets FXI and prekallikrein to their active forms. Conclusions Autoactivation of FXII by polyP, of the size found in platelets, proceeds via an active single‐chain intermediate. sc FXII –polyP 70 shows activity towards physiological substrates, and may represent the primary event in initiating contact activation in vivo .