Structure-Guided Topographic Mapping and Mutagenesis to Elucidate Binding Sites for the Human Ether-a-Go-Go-Related Gene 1 Potassium Channel (KCNH2) Activator NS1643

Abstract

Loss-of -function mutations in human ether-a-go-go-related gene 1 (hERG1) is associated with life-threatening arrhythmias. hERG1 activators are being developed as treatments for acquired or genetic forms of long QT syndrome. The locations of the putative binding pockets for activators are still being elucidated. In silico docking of the activator 1,3-bis-(2-hydroxy-5-trifluoromethylphenyl)-urea (NS1643) to an S1-S6 transmembrane homology model of hERG1 predicted putative binding sites. The predictions of the in silico docking guided subsequent in vitro mutagenesis and electrophysiological measurements. The novel interacting site for NS1643 is predicted around Asn629 at the outer mouth of the channel. The applied N629H mutation is the sole amino acid replacement in the literature that abrogates the NS1643-induced left shift of the V 1/2 of activation. In contrast, both N629T and N629D showed pharmacologic responses similar to wild type. Another important interacting pocket is predicted at the intracellular surface in the S4 -S5 linker. Mutagenesis of the residues critical to interactions in this pocket had major effects on the pharmacologic response to NS1643. The inward conductance elicited by hyperpolarization of D540K hERG1 was abrogated by NS1643 treatment, suggesting that it alters the inward movement of the S4 segment. The neighboring E544L mutation markedly exaggerated tail-current responses to NS1643. However, an L564A substitution inhibited drug response. Structure-guided mutagenesis identified widespread clusters of amino acids modulating drug-induced shifts in inactivation; such modulation may reflect allosteric changes in tertiary structure. Modelguided mutagenesis led to the discovery of a range of novel interacting residues that modify NS1643-induced pharmacologic responses.

The Human Ether-A-Go-Go–Related Gene Activator NS1643 Enhances Epilepsy-Associated KCNQ Channels

“…Identification of F137, critical for NS1643 potentiation activity on the KCNQ2 channel, further raises the speculation that NS1643 may act on the voltage-sensing domain as ztz240 does . In the previous study by Durdagi et al (2012), some critical residues for NS1643 potentiating hERG channels have been identified. One putative binding site is N629 at the outer mouth of the channel.…”

“…Noticeable differences in the effects of NS1643 on hERG channels were observed depending on expression systems (Casis et al, 2006;Hansen et al, 2006;Schuster et al, 2011;Durdagi et al, 2012). Thus, potentiation effects of NS1643 on hERG channels expressed in CHO-K1 cells, the transient expression system we used in this study, were characterized first.…”

In silicoinvestigation of PARP-1 catalytic domains inholoandapostates for the design of high-affinity PARP-1 inhibitors

“…They were docked into the central cavities of previously generated and validated human ether-a go-go related gene (hERG1) K-channel models by our group [12][13][14][15][16][17][18][19] . Table 2 shows the Glide/XP docking scores of these drugs in hERG1.…”

“…It is well established that human ether-à-go-go related gene 1 (hERG1) K channel is overexpressed in many tumors and, therefore, pro-arrhythmic potential of all novel inhibitors is an important concern in structure driven drug development. To circumvent potential risks associated with druginduced QT prolongation, we report on an in silico cardiotoxicity screening against previously developed hERG1 models [12][13][14][15][16][17][18][19] by our group.…”

Investigation of inhibition of human glucose 6-phosphate dehydrogenase by some 99mTc chelators byin silicoandin vitromethods

“…Docking and atomistic MD simulations approved as informative approaches, profoundly provided to track and pursue the treatments of protein and small molecules [35][36][37][38][39][40][41][42][43][44][45] . Conformational stabilities and transitions, protein-ligand interactions (hydrogen bonding, hydrophobic interactions, salt bridges, etc.)…”