Talk abstract:

Voltage Sensitivity of the Sodium-Potassium Pump:
Structural Inferences from Kinetic Observations

Paul De Weer
Department of Physiology
School of Medicine
University of Pennsylvania
Philadelphia, PA 19104-6085
deweer@mail.med.upenn.edu

The sodium pump produces electric current and must therefore be sensitive to the voltage across the membrane (Vm), and have a reversal Vm where current changes direction. This Vm sensitivity, which reflects a charge moving across a field or vice versa holds mechanistic and structural information about the enzyme. In the forward mode, the sodium pump's current-voltage relationship has --at high [Na+]o-- a positive slope at negative Vm and --at low [K+]o-- a negative slope at positive Vm. In the backward mode, turnover is enhanced by negative Vm and reaches a plateau.

Vm sensitivity could reside in the Na-transporting hemicycle or the K hemicycle or both. We focused on the former because electroneutral (ADP-requiring) Na+/Na+ exchange is conveniently measured in squid axons and because charge translocation in the Na hemicycle had been demonstrated on bilayer-adsorbed Na/K-ATPase. This exchange mode represents the reversible operation of the Na hemicycle via internal ion binding, occlusion, and external release. For kinetic analysis, this scheme can be reduced to two steps with four rate constants, any or all possibly Vm sensitive. We found that ADP-requiring Na+/Na+ exchange, though electroneutral, is Vm sensitive, being enhanced by negative Vm along a saturating sigmoid whose midpoint moves a fixed distance leftward for each 2-fold reduction in [Na+]o. Such kinetic equivalence between [Na+]o and Vm suggests that only the pseudo-first-order rate constant for external Na+ rebinding is Vm dependent. The most economical model for such kinetics is one in which Na+ are released from the pump to, or regain access to the pump from, the external medium via a narrow high-field channel. A Boltzmann partition effect enhances effective [Na+] at the bottom of the well at negative Vm, more so for deeper wells. Our model explains Vm dependence of Na+/Na+ exchange and backward pumping as well as the effect of [Na+]o on the forward mode's Vm dependence. Steady-state analysis cannot resolve the Vm/[Na+]o equivalence of individual steps in a reaction involving sequential release/rebinding of several ions. By subjecting the squid axon sodium pump (in the Na+/Na+ exchange mode) to Vm jumps and analyzing the resulting transient currents we identified three components.

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1998-1999 Mathematics in Biology