Dose dependence of thrombin cleavage of recombinant human FVIII.

The dose dependence of WT and mutant thrombin (0.1-2.0 nM) cleavage of human factor VIII (100 nM) was investigated using the 2-step chromogenic assay, as described in “Material and methods.” The diagram shows the dose dependence for WT thrombin (▪), the ABE-I mutants Lys65Ala (▴), Arg68Ala (■), and Tyr71Ala (♦), the ABE-II mutant Arg98Ala (●), the 50-insertion loop mutant Trp50Ala (○), and the Na⁺ loop mutant Glu229Ala (▵). All points had standard deviations of 15% or less from at least 2 separate experiments performed in duplicate.