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 Rg(3) (Rg(3)), 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 Rg(3) regulates Ca2+ channels through specific interaction site(s) for Ca2+ influx inhibition through Ca2+ channels. However, it was not known how Rg(3) 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(alpha(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 Rg(3) action. Mutations of L427R, N428R and L431K in transmembrane domain-I-segment 6 (IS6) of the channel significantly attenuated the Rg(3) action and caused rightward shifts in dose-response curves. Rg(3) 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 Rg(3)-induced negative shift of inactivation voltage. Rg(3) had no effects on inactivation time constant in wild-type and mutant channels. These results indicate that Rg(3) 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 Rg(3) effect on channel properties.