MOL Manuscript #108886 1 Supplemental Materials Article title: Molecular basis of altered hERG1 channel gating induced by ginsenoside Rg3 Authors: Alison Gardner, Wei Wu, Steven Thomson, Eva-Maria Zangerl-Plessl, Anna Stary- Weinzinger and Michael C. Sanguinetti Journal: Molecular Pharmacology Supplemental Fig. 1. Scanning mutagenesis of select residues located in the S5-pore region of hERG1. The shift in V 0.5 for activation (black bars) and fold-change in deact (grey bars) induced by 3 μM ginsenoside Rg3 are plotted for wild-type (WT) and select mutant hERG1 channels as indicated on x-axis (n indicated below V 0.5 bars).
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MOL Manuscript #108886
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Supplemental Materials
Article title: Molecular basis of altered hERG1 channel gating induced by ginsenoside Rg3
Authors: Alison Gardner, Wei Wu, Steven Thomson, Eva-Maria Zangerl-Plessl, Anna Stary-
Weinzinger and Michael C. Sanguinetti
Journal: Molecular Pharmacology
Supplemental Fig. 1. Scanning mutagenesis of select residues located in the S5-pore region of
hERG1. The shift in V0.5 for activation (black bars) and fold-change in deact (grey bars) induced
by 3 µM ginsenoside Rg3 are plotted for wild-type (WT) and select mutant hERG1 channels as
indicated on x-axis (n indicated below V0.5 bars).
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Supplemental Fig. 2. Plot of the fold change in deact (logarithmic scale) as a function of deact (at
a Vret of −70 mV) under control conditions for all mutant channels examined in this study. The 4
mutant channels with a control deact > 1.4 s (see Suppl. Table 2) are not included. There was no
correlation between the control deact and the fold change in this parameter induced by Rg3
(linear regression adjusted R2 = 0.099).
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Supplemental Fig. 3. Plot of the fold change in Itail-peak induced by ginsenoside Rg3 as a function
of deact (at a Vret of −70 mV) under control conditions for all the mutant channels examined in
this study. Itail was fitted to an exponential function, extrapolated to the time point where Vt was
pulsed to −70 mV to estimate the initial value of Itail-peak. Most of the faster deactivating mutants
had a greater than average fold change in Itail-peak. Specifically, 9 of the 11 mutant channels with a
deact < 130 ms had a fold change in Itail-peak > 1.3 (indicated by vertical dashed line). WT channel
is indicated by red circle. Y420A/L452A and the two other outlying mutant channels are labeled.
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Supplemental Fig. 4. Surface representations of the top view (upper) and side view (lower) of
the VSD with the ginsenoside Rg3 shown as sticks in the activated state (A) and the activated1
state (B).
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Supplemental Fig. 5. Root mean square deviation (RMSD) plots of ginsenoside Rg3 over
simulation time. Rmsd of the two 50 ns runs in the activated model (black and blue) of the
activated state (A) and the activated1 model (B).
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Supplemental Fig. 6. Root mean square fluctuations (RMSF) of two 50 ns runs (black and blue)
for all residues of the hERG1 voltage sensor. Both states reveal relatively similar RMSF profiles,
with the loop regions being most flexible.
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Supplemental Fig. 7. Comparison of the VSDs from cryo-EM structures of hERG1 and rat
EAG1. Cartoon representation of the VS module of hERG1 (blue) and EAG1 (light orange).
The missing loops in the hERG1 structure are indicated by red dots. The overall structure of the
VSD is similar between EAG1 and hERG1, with an overall RMSD value of all atoms of 1.25 Å.
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Supplemental Table 1. Voltage dependence of activation for WT and mutant hERG1
channels. Normalized g/gmax-Vt plots were fitted with a Boltzmann function to determine the
half-point of activation (V0.5) and slope factor (k) for the relationships. Data represent the average
± S.D. determined before (control) and after treatment of oocytes with 3 µM ginsenoside Rg3
unless indicated otherwise (n = number of oocytes).
V0.5 (mV) k
Mutation Control S.D. 3µM Rg3 S.D. Control S.D. 3µM Rg3 S.D. n