Physical and Functional Interactions Between the KCNQ1 and KCNE1 C-Terminal Domains

KCNE1 is a regulatory subunit that associates with pore-forming subunits of KCNQ1 to form a channel complex that carries the slowly activating delayed rectifier current, IKs. Interactions between the transmembrane regions of KCNE1 and KCNQ1 S6 control activation kinetics of IKs, while interactions between the intracellular C-termini of the two subunits may dictate deactivation kinetics. Numerous Long QT Syndrome mutations occur in the C-termini of KCNQ1 and KCNE1, indicating the functional importance of these regions. We have located the proximal third of the KCNQ1 C-terminus (KCNQ1-CT) as a site of direct interaction with the KCNE1 C-terminus (KCNE1-CT) via co-immunoprecipitation studies, in vitro pull-down assays and Surface Plasmon Resonance analyses of purified recombinant proteins. Electrophysiological studies employing co-expressed soluble KCNE1-CT with full length KCNQ1 now provide functional evidence that support the physical association findings. When KCNE1-CT is co-expressed with KCNQ1 in CHO cells and analyzed by whole-cell patch clamp, deactivation kinetics of the KCNQ1 current are accelerated. Similarly, deactivation of IKs is accelerated by co-expressing KCNE1-CT with KCNQ1 and full-length KCNE1. KCNE1-CT also shifts the voltage dependence of activation of KCNQ1 current but not IKs current, and has no significant effect on activation kinetics of either current. Thus, excess soluble KCNE1-CT is capable of interacting with KCNQ1 and can perturb the KCNQ1/KCNE1-C-terminal interactions determining deactivation rates of IKs. Work in the laboratory now focuses on identifying the interacting residues of the KCNQ1 and KCNE1 C-termini. Preliminary experiments using Hydrogen-Deuterium Exchange coupled to Mass Spectrometry show that deuterium incorporation into purified KCNE1-CT is slowed by the addition of KCNQ1-CT, implying that binding between the two peptides protect certain residues of KCNE1-CT from being deuterated. Further experiments will work towards delineating the precise structural nature of this interaction.