A single amino acid difference between ether-a-go-go- related gene channel subtypes determines differential sensitivity to a small molecule activator.
Activators of human ether-a-go-go-related gene 1 (hERG1) channels, such as (3R,4R)-4-[3-(6-methoxy-quinolin-4-yl)-3-oxo-propyl]-1-[3-(2,3,5-trifluoro-phenyl)-prop-2-ynyl]-piperidine-3-carboxylic acid (RPR260243), reverse the effect of hERG1 blockers and shorten the duration of cardiac action potentials. RPR260243 (RPR) slows the rate of deactivation and shifts the voltage dependence of channel inactivation to more positive potentials. We recently mapped the binding site for RPR to several residues located near the cytoplasmic ends of the S5 and S6 helices of the hERG1 subunit. These residues are conserved in the highly homologous ether-a-go-go-related gene 3 (ERG3) subunit; however, RPR blocks ERG3 channels. Here, we compare hERG1 and rat ERG3 (rERG3) channels to explore the molecular basis for differential channel sensitivity to RPR. Channels were heterologously expressed in Xenopus laevis oocytes, and currents were recorded using the two-electrode voltage-clamp technique. Site-directed mutagenesis was used to swap the two residues within the putative binding domain that differed between hERG1 and rERG3. The differential sensitivity of hERG1 and rERG3 channels to the agonist effect of RPR could be accounted for by a single S5 residue (Thr556 in hERG1, Ile558 in rERG3). A Thr in this position favors agonist activity, whereas an Ile reveals a secondary blocking effect of RPR.
Perry
Matthew
M
Nora Eccles Harrison Cardiovascular Research and Training Institute, Department of Physiology, University of Utah, 95 South 2000 East, Salt Lake City, UT 84112, USA.
Sanguinetti
Michael C
MC
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics*; D000818:Animals; D001665:Binding Sites; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels / Q000378:metabolism*; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D010880:Piperidines / Q000494:pharmacology*; D024642:Potassium Channels, Voltage-Gated / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D011804:Quinolines / Q000494:pharmacology*; D051381:Rats; D016415:Sequence Alignment; D040681:Structural Homology, Protein; D014982:Xenopus laevis
D000596:Amino Acids; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels; C000606913:KCNH2 protein, human; C109501:Kcnh7 protein, rat; D010880:Piperidines; D024642:Potassium Channels, Voltage-Gated; D021122:Protein Subunits; D011804:Quinolines; C499186:RPR260243