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MJ Rabiet, M Jandrot-Perrus, JP Boissel, J Elion and F Josso
Thrombin Metz and normal thrombin, resulting from activation of the
respective prothrombins by factor Xa in the presence of calcium,
phospholipid, and factor Va, were purified by chromatography on sulfopropyl
Sephadex. By physicochemical criteria, thrombin Metz is identical to normal
thrombin. Its functional properties were investigated in some reactions in
which thrombin is classically involved. Thrombin Metz exhibits less than 4%
of fibrinogen clotting activity. Both Km and Kcat, determined on S2238, are
abnormal. Titration with the high-affinity competitive inhibitor of
thrombin, DAPA, shows that fluorescence enhancement of the probe is only
34% in binding to thrombin Metz when compared to that observed in binding
to normal thrombin. High-performance liquid chromatography has been used to
measure the simultaneous rate of release of fibrinopeptides A and B. A
decreased release rate for both fibrinopeptides, more marked for
fibrinopeptide B, results in a slow fibrin polymerization, as followed by
absorbance at 450 nm. Thrombin Metz is less than 5% as effective as normal
thrombin in inducing platelet aggregation. Interaction with antithrombin
III is slower than normal when followed by SDS gel electrophoresis and
inhibition of the amidolytic activity of thrombin on S2238. This
abnormality is not observed in the presence of heparin. However, thrombin
Metz binds less tightly to a heparin-Sepharose column, and the direct
inhibition of heparin on its activity on S2238 is weaker. From these
results, we can predict that the defect in thrombin Metz affects the
catalytic site or its vicinity and, jointly or consequently, the region of
interaction of thrombin with antithrombin III and heparin.
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| Copyright © 1984 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
