A simple rate law that describes the kinetics of the heparin-catalyzed reaction between antithrombin III and thrombin.
The kinetics of the reaction between human thrombin and antithrombin III were studied in the presence of heparin fractionated according to its molecular size and affinity for antithrombin III. Reactions were monitored continuously through the loss of fluorescence intensity accompanying the displacement of the reversible inhibitor dansylarginine-N-(3-ethyl-1,5-pentanediyl)amide from thrombin upon its interaction with antithrombin III. As the concentration of heparin was increased in the presence of fixed concentrations of the two proteins, initial rates of the reaction increased up to a maximum and then decreased. Both the maximum rate and the concentration of heparin required to obtain a maximum increased with increased concentration of the proteins. The reaction appeared second order overall and first order with respect to each protein when heparin was in excess and zero order overall when the proteins were in excess. The existence of zero order kinetics at excess concentration of proteins indicates that the reaction is saturable in both proteins. The kinetics of the heparin-catalyzed reaction measured over an appreciable range of protein and heparin concentrations could be rationalized well by the simple rate equation, v = k X fa X fb X [H]t, where k is an intrinsic rate constant for the reaction between bound antithrombin III and thrombin, fa and fb are, respectively, the fractional saturation of available sites on heparin by antithrombin III and thrombin, and [H]t is the total concentration of heparin. This rate equation implies that interactions of both proteins with heparin are involved in the expression of catalytic activity through a template mechanism as proposed by M. W. Pomeranz and W. G. Owen ((1978) Biochim. Biophys. Acta 535, 66-77) and M. J. Griffith ((1982) J. Biol. Chem. 257, 7360-7365).
D000990:Antithrombin III / Q000378:metabolism*; D006493:Heparin / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D008433:Mathematics; D008956:Models, Chemical; D013917:Thrombin / Q000378:metabolism*
D000990:Antithrombin III; D006493:Heparin; D013917:Thrombin