Guo, Jiqing; Cheng, Yen May; Lees-Miller, James P.; Perissinotti, Laura L.; Claydon, Tom W.; Hull, Christina M.; Thouta, Samrat; Roach, Daniel E.; Durdagi, Serdar; Noskov, Sergei Y.; Duff, Henry J.
NS1643 Interacts around L529 of hERG to Alter Voltage Sensor Movement on the Path to Activation
BIOPHYSICAL JOURNAL, 108:1400-1413, MAR 24 2015

Activators of hERG1 such as NS1643 are being developed for congenital/acquired long QT syndrome. Previous studies identify the neighborhood of L529 around the voltage-sensor as a putative interacting site for NS1643. With NS1643, the V-1/2 of activation of L529I (-34 +/- 4 mV) is similar to wild-type (WT) (-37 +/- 3 mV; P > 0.05). WT and L529I showed no difference in the slope factor in the absence of NS1643 (8 +/- 0 vs. 9 +/- 0) but showed a difference in the presence of NS1643 (9 +/- 0.3 vs. 22 +/- 1; P < 0.01). Voltage-clamp-fluorimetry studies also indicated that in L529I, NS1643 reduces the voltage-sensitivity of S4 movement. To further assess mechanism of NS1643 action, mutations were made in this neighborhood. NS1643 shifts the V-1/2 of activation of both K525C and K525C/L529I to hyperpolarized potentials (-131 +/- 4 mV for K525C and -120 +/- 21 mV for K525C/L529I). Both K525C and K525C/K529I had similar slope factors in the absence of NS1643 (18 +/- 2 vs. 34 +/- 5, respectively) but with NS1643, the slope factor of K525C/L529I increased from 34 +/- 5 to 71 +/- 10 (P < 0.01) whereas for K525C the slope factor did not change (18 +/- 2 at baseline and 16 +/- 2 for NS1643). At baseline, K525R had a slope factor similar to WT (9 vs. 8) but in the presence of NS1643, the slope factor of K525R was increased to 24 +/- 4 vs. 9 +/- 0 mV for WT (P < 0.01). Molecular modeling indicates that L529I induces a kink in the S4 voltage-sensor helix, altering a salt-bridge involving K525. Moreover, docking studies indicate that NS1643 binds to the kinked structure induced by the mutation with a higher affinity. Combining biophysical, computational, and electrophysiological evidence, a mechanistic principle governing the action of some activators of hERG1 channels is proposed.


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