Involvement of methionine residues in the fast inactivation mechanism of the sodium current from toad skeletal muscle fibers

The role of methionine residues on the fast inactivation of the sodium chan nel from toad skeletal muscle fibers was studied with the mild oxidant chlo ramine-T (CT). Isolated segments of fibers were voltage clamped in a triple Vaseline(R) gap chamber. Sodium current was isolated by replacing potassiu m ions by tetramethylammonium ions in the external and internal solutions. Externally applied chloramine-CT was found to render noninactivating a larg e fraction of sodium channels and to slow down the fast inactivation mechan ism of the remainder fraction of inactivatable channels. The action of CT a ppeared to proceed first by slowing and then removing the fast inactivation mechanism. The voltage dependence of the steady-state inactivation of the inactivatable CT-treated currents was shifted +10 mV. CT also had a blockin g effect on the sodium current, but was without effect on the activation me chanism. The effects of CT were time and concentration dependent and irreve rsible. The use of high CT concentrations and/or long exposure times was fo und to be deleterious to the fiber. This side effect precluded the complete removal of fast inactivation. The effects of CT on the fast inactivation o f the sodium current can be explained assuming that at least two methionine residues are critically involved in the mechanism underlying this process.