Open AccessThrombin‐induced fibrinopeptide release from a fibrinogen variant (fibrinogen Sydney I) with an Aα Arg‐16 → His substitution
Author(s) -
SOUTHAN Christopher,
LANE David A.,
BODE Wolfram,
HENSCHEN Agnes
Publication year - 1985
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.0014-2956.1985.00593.x
Subject(s) - fibrinogen , thrombin , trypsin , chemistry , peptide , biochemistry , enzyme kinetics , kinetics , enzyme , chromatography , active site , biology , platelet , immunology , physics , quantum mechanics
Fibrinogen, Purified from a recently identified case of dysfibrinogenaemia, fibrinogen Sydney I, was shown by thrombin digestion, high‐performace liquid chromatography (HPLC) and amino acid analysis to be a heterozygous case of an Aα Arg‐16 → His substitution. Kinetic studies have been carried out on the thrombin‐induced release of fibrinopeptide A (FPA), fibrinopeptide B (FPB) and the variant peptide [His 16 ]FPA. When thrombin was added to fibrinogen Sydney I at a concentration of 0.2 U/ml release of FPA was rapid and there was a 79‐fold reduced rate of release of [His 16 ]FPA, but the rate of release of FPB was not appreciably reduced. In contrast, at lower thrombin concentrations the rate of FPB release was reduced in proportion to the rate of total FPA release, supporting the view that release of fibrinopeptides is a sequential process. The second‐order kinetic constant k cat / K m for hydrolysis of the abnormal Aα chain by thrombin was calculated from Lineweaver‐Burk plots to be 16–30‐fold less than that for the normal Aα chain. Molecular modelling studies, using a refined model of the trypsin‐pancreatic‐trypsin‐inhibitor complex have been used to suggest how the histidine at the P 1 site can be accommodated within the enzyme hydrophobic active‐site pocket.