The use of acetylated factor X to prevent feedback activation of factor VIII during factor X activation: a tool for kinetic studies

Analytical Biochemistry
P Neuenschwander, J Jesty

Abstract

The modification of human factor X by 2-sulfo-N-succinimidyl acetate was investigated and shown to produce a factor X species which, when activated, has no activity toward factor VIII. Acylation of factor X (0.9 microM) was carried out in the presence of 1 mM calcium at different reagent concentrations and pH values at 22 degrees C for time courses up to 1 h. Optimal modification was achieved using 0.3 mM reagent at pH 8.0 for 30 min. The modified zymogen, acetylated factor X, is activated at full rates by factor IXa/VIIIa and by the factor X-activating protein of Russell's viper venom. The activated product, acetylated Xa, has an enhanced amidolytic activity (110%) but has almost no detectable clotting activity (0.1%). More importantly, we have shown that acetylated Xa, in contrast to native Xa, does not activate factor VIII. This allows accurate quantitation of factor VIII activation without complications due to positive feedback reactions. We have demonstrated this in an examination of the activation of factor VIII by factor IXa.

References

Jan 1, 1988·Annual Review of Biochemistry·K G MannS Krishnaswamy
Jan 29, 1988·Biochimica Et Biophysica Acta·P J Fay
Apr 1, 1988·Proceedings of the National Academy of Sciences of the United States of America·D D Pittman, R J Kaufman
Dec 10, 1986·Biochimica Et Biophysica Acta·S A MorrisonJ Jesty
Dec 31, 1985·Biochemistry·P LollarD N Fass
Apr 1, 1982·The Journal of Clinical Investigation·M B Hultin

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