Mutations Leu427, Asn428, and Leu431 Residues within Transmembrane Domain-I-Segment 6 Attenuate Ginsenoside-Mediated L-Type Ca²⁺ Channel Current Inhibitions

Abstract

Many lines of evidences have shown that Panax ginseng exhibits beneficial effects on cardiovascular systems. We previously demonstrated that ginsenoside Rg₃ (Rg₃), one of active ingredients of Panax ginseng, inhibits Ca²⁺ channel currents in a stereospecific manner and affects the steady-state activation but not inactivation. This points a possibility that Rg₃ regulates Ca²⁺ channels through specific interaction site(s) for Ca²⁺ influx inhibition through Ca²⁺ channels. However, it was not known how Rg₃ interacts with Ca²⁺ channel proteins. In the current study, we sought to identify these site(s) in Xenopus oocytes expressing cardiac wild-type and mutant L(α_1C)-type Ca²⁺ channels using the two-microelectrode voltage-clamp technique. To this end, we assessed how various point mutations of the L-type Ca²⁺ channel affected the Rg₃ action. Mutations of L427R, N428R and L431K in transmembrane domain-I-segment 6 (IS6) of the channel significantly attenuated the Rg₃ action and caused rightward shifts in dose–response curves. Rg₃ treatment produced a negative shift in the inactivation voltage but did not alter the steady-state activation voltage, and none of the mutant channels affected the Rg₃-induced negative shift of inactivation voltage. Rg₃ had no effects on inactivation time constant in wild-type and mutant channels. These results indicate that Rg₃ inhibition of L-type Ca²⁺ channel currents is attenuated by mutations of Leu427, Asn428 and Leu431 in transmembrane IS6 residues. Leu427, Asn428 and Leu431 residues of the L-type Ca²⁺ channel play important roles in the Rg₃ effect on channel properties.