Multiple structural domains contribute to voltage-dependent inactivation of rat brain α(1E) calcium channels

Abstract

We have investigated the molecular determinants that mediate the differences in voltage-dependent inactivation properties between rapidly inactivating (R-type) α(1E) and noninactivating (L-type) α(1C) calcium channels. When coexpressed in human embryonic kidney cells with ancillary β(1b) and α2-δ subunits, the wild type channels exhibit dramatically different inactivation properties; the half-inactivation potential of α(1E) is 45 mV more negative than that observed with α(1C), and during a 150-ms test depolarization, α(1E) undergoes 65% inactivation compared with only about 15% for α(1C). To define the structural determinants that govern these intrinsic differences, we have created a series of chimeric calcium channel α1 subunits that combine the major structural domains of the two wild type channels, and we investigated their voltage-dependent inactivation properties. Each of the four transmembrane domains significantly affected the half-inactivation potential, with domains II and III being most critical. In particular, substitution of α(1C) sequence in domains II or III with that of α(1E) resulted in 25-mV negative shifts in half-inactivation potential. Similarly, the differences in inactivation rate were predominantly governed by transmembrane domains II and III and to some extent by domain IV. Thus, voltage-dependent inactivation of α(1E) channels is a complex process that involves multiple structural domains and possibly a global conformational change in the channel protein.