Molecular Basis of Altered hERG1 Channel Gating Induced by Ginsenoside Rg3

Abstract

Outward current conducted by human -related gene type 1 (hERG1) channels is a major determinant of action potential repolarization in the human ventricle. Ginsenoside 20()-Rg3 [Rg3; (2,3,4,5,6)-2-[(2,3,4,5,6)-4,5-dihydroxy-2-[[(3,5,8,9,10,12,13,14,17)-12-hydroxy-17-[(2)-2-hydroxy-6-methylhept-5-en-2-yl]-4,4,8,10,14-pentamethyl-2,3,5,6,7,9,11,12,13,15,16,17-dodecahydro-1-cyclopenta[]phenanthren-3-yl]oxy]-6-(hydroxymethyl)oxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol], an alkaloid isolated from the root of , slows the rate of hERG1 deactivation, induces channels to open at more negative potentials than normal, and increases current magnitude. The onset of Rg3 action is extremely fast, suggesting that it binds to an extracellular accessible site on the channel to alter its gating. Here we used a scanning mutagenesis approach to identify residues in the extracellular loops and transmembrane segments of hERG1 that might interact with Rg3. Single or multiple residues of hERG1 were mutated to Ala or Cys and the resulting mutant channels were heterologously expressed in oocytes. The effects of Rg3 on the voltage dependence of activation and the deactivation rate of mutant channel currents were characterized using the two-microelectrode voltage clamp technique. Mutation to Ala of specific residues in the S1 (Tyr420), S2 (Leu452, Phe463), and S4 (Ile521, Lys525) segments partially inhibited the effects of Rg3 on hERG1. The double mutant Y420A/L452A nearly eliminated the effects of Rg3 on voltage-dependent channel gating but did not prevent the increase in current magnitude. These findings together with molecular modeling suggest that Rg3 alters the gating of hERG1 channels by interacting with and stabilizing the voltage sensor domain in an activated state.

Overcoming challenges of HERG potassium channel liability through rational design: Eag1 inhibitors for cancer treatment

“…161 The residues in the outer region of VSD that reduce the effect of ginsenoside Rg3 (7) on HERG were identified using alanine scanning mutagenesis and MD simulations. The relevant residues were Y420, L452, HERG in an activated state, 180 compared to deactivated state stabilization by divalent ions that bind to a similar location. 181…”

“…Mutation L417A produced a channel that enhanced the effects of ginsenoside Rg3. It is proposed that ginsenoside Rg3 ( 7 ) stabilizes the VSD of HERG in an activated state, 180 compared to deactivated state stabilization by divalent ions that bind to a similar location 181 …”

Targeting Potassium Channel Trafficking in Cardiac Arrhythmia

“…Activators influence gating kinetics and can, for example, slow down or remove inactivation and/or facilitate activation . Kv11.1 activators normally interact with a region distant from the inner cavity , but they can bind to several distinct sites of the channel (Perry et al, 2007;Guo et al, 2015;Gardner et al, 2017). Negative allosteric modulators decrease the binding affinity of IKr blockers, either by increasing dissociation rates, lowering association rates, or both (Christopoulos et al, 2014).…”