Bruce A. Benjamin
Oklahoma State University
College of Osteopathic Medicine
Tulsa, OK 74107
Joint work with E. A. Johnson, El Laboratorio de Fisiologia Cuantitativo, 04638 Mojacar Almeria, Spain.
Computer simulation of the classical Post-Albers scheme for the reaction cycle of the sodium pump (characterized by single translocational steps (E1 leftrightarow E2) for Na+ & K+) gave a poor fit to the non-vectorial activation of the enzyme by Na+ & K+ and the binding of K+ to the Na+-free enzyme. However, the inclusion of an intermediate ("occluded") translocational state for both Na+ & K+ abolished this limitation. The resulting best, simultaneous, fits to the Na+ & K+ activation and K+-binding data showed that the optimized rate constants for the additional steps for translocation of Na+ and the MgATP-free-K+-bound forms of the enzyme assumed values consistent with these steps being "de-occlusion" steps. The equilibrium constants of these steps were of the order Of 10-6 and the on-rate for binding of the ions to the E2 form of the enzyme approached 1010 M-1 sec-1 -the on-rate for binding of small ions (free of H2O). The second translocational step for the K+-MgATP-bound form of the enzyme effectively disappeared in that the forward and reverse rates of the step became equally large, as though the regulatory effect of MgATP binding to the E2 form was to effectively remove the "occluded" state. In addition, with minor changes in parameter values, this enhanced Post-Albers scheme gave excellent fits to vectorial pump activity in single, isolated cardiac muscle cells. Such fits included the activation of membrane pump-generated current by intracellular [Na+], extracellular [K+], and the relationship between pump current and transmembrane potential at various extracellular [Na+]. Given this working mechanism for the sodium pump and the corresponding mechanism for the Na,K,2Cl cotransporter (Benjamin, B.A. & E.A. Johnson, Am. J. Physiol., 273: 473-482, 1997), it will now be possible to study their cooperative role in the regulation of intracellular Na+, K+ and C1-.