Linkage at Steady State: Allosteric Transitions of Thrombin

This chapter illustrates the importance of exploring linkage at steady state and the limitations of the equilibrium description. The exact analytical solution for the kinetic linkage scheme describing allosteric effects in serine proteases of the blood coagulation cascade and use this scheme to characterize energetically the allosteric properties of thrombin. The control of thrombin activity by allosteric effectors such as Na⁺ and the hirudin tail binding to the fibrinogen recognition site demonstrates that a great deal of information can be obtained from linkage studies under nonequilibrium conditions. In the case of thrombin, the linkage between important structural domains of the enzyme is dominated by the kinetic, rather than the equilibrium, components. The exact solution of the linkage scheme for serine proteases in the presence of an allosteric effector, as an extension of the Botts-Morales treatment of the action of a modifier is presented. The solution reveals the substantial complexity of linked functions at steady state and, at the same time, provides a convincing example of how macromolecules can exploit more complicated pathways of communication to accomplish biological function. The treatment sets the stage for a quantitative analysis of allosteric effects that dominate the blood coagulation cascade. It also provides the necessary framework for casting protein-protein interactions in this biologically relevant system.