EPR Spectroscopic Studies on the Binding of the Full Length Human KCNE1 Protein with the Voltage Gated Potassium Channel KCNQ1

Abstract

KCNE1 is a single transmembrane protein that modulates the activity of voltage gated potassium channels. KCNE1 interacts with KCNQ1 and decreases the rate of channel activation, increases conductance, and generates a slowly activating K+ current critical for cardiac repolarization. Mutations on either KCNE1 or KCNQ1 genes in E1/Q1 complex can lead to cardiac long QT syndrome. Despite the biological significance of the Q1/E1 interaction, its exact nature is not fully understood yet. In this study, we have used different electron paramagnetic resonance (EPR) spectroscopic techniques to provide the direct evidence of binding activities of KCNQ1 with KCNE1. A CW-EPR titration experiment was conducted on different sites in the C-terminus of KCNQ1 with the addition of WT-KCNE1 to determine the binding mechanism of Q1 and E1. The CW EPR line shape analysis indicates that the motion of the spin label on Q1 decreases with the addition of WT-KCNE1 and saturates at molar ratio of 1:3 (Q1:E1). Double electron-electron resonance (DEER) Spectroscopy was used to measure distances between the spin labels attached on E1 and Q1 separately in Q1/E1 complex. A DEER distance of 33 A was obtained between the spin labels at 318th position on Q1 and 106th position on E1 that provides the direct evidence of formation of a Q1 and E1 complex. These EPR techniques are very useful for determining the structural model of the Q1/E1 complex.