Xh. He et al., INFLUENCE OF ARGININE-93, ARGININE-97, AND ARGININE-101 OF THROMBIN TO ITS FUNCTIONAL SPECIFICITY, Biochemistry, 36(29), 1997, pp. 8969-8976
Mutation of three Arg residues, 93, 97, and 101, to Ala in thrombin (t
hrombin R93,97,101A) has previously been shown to eliminate most hepar
in acceleration of thrombin inhibition by antithrombin and most of the
ability of chondroitin sulfate (CS) on thrombomodulin (TM) to enhance
affinity for TM and to eliminate the characteristic high-affinity int
eraction with protein C observed with TM lacking CS. In this study we
examined the relative impact of mutation of these Arg residues alone a
nd in combination on the above reactions and, in addition, on the abil
ity of rabbit TM to accelerate thrombin inhibition by antithrombin. Th
e order of importance for heparin acceleration of inhibition by antith
rombin was Arg 101, 93, and 97. In contrast, Arg 97 was the major resi
due required for TM-dependent acceleration of reactivity with antithro
mbin and for CS-dependent enhancement of TM affinity. Arg 101 and 93 w
ere critical for TM-dependent, high-affinity protein C interaction at
low Ca2+ concentrations, while Arg 97, which was critical for the othe
r TM-dependent effects, played no detectable role in this metal depend
ence. These results illustrate that these Arg residues in anion bindin
g exosite 2 contribute very differently to the diverse reactions depen
dent on that domain in thrombin.