The Role of Putative Intramembraneous Glutamic Acid Residues of the α-Subunit of the Sodium Pump in External Cation Binding

Abstract

During the cation translocation of the Na⁺/K⁺ transport cycle two positive charges have been suggested to be compensated by negative charges (3) It was proposed that like in other cation binding proteins the cations are ligated with six to eight oxygens of carboxyl groups and that glutamic acid residues in intramembranous domains are involved (1,2,4). Based on their measurements with the carboxyl reagent DCCD Karlish and coworkers (4) suggested for possible candidates the glutamic acid residues E-334 and E-959 (in Torpedo numbering). We addressed this question in voltage-clamp experiments on Xenopus oocytes with expressed mutated pumps of Torpedo electroplax by analysing the influence of external K⁺ on transport activity. In absence of extracellular Na⁺ an apparent K_m value for pump stimulation by external K⁺ can be determined that shows exponential dependence on membrane potential. This has been interpreted by a voltage-dependent access of the K⁺ ions to their occulsion site (5). For the Torpedo pump the voltage dependence of K_m had to be described by the sum of two exponentials

$$K_{m}=K_{ml}\left ( 0mV \right)e^{z_{1}VF\ /\ RT}+K_{m2}\ e^{z_{1}VF\ /\ RT}$$

and has been explained by a sequential binding of the two K⁺ ions (6) where the effective valencies Z _i reflect the dielectric length of the access for the two cations.