Preparation, functional characterization, and NMR studies of human KCNE1, a voltage-gated potassium channel accessory subunit associated with deafness and long QT syndrome.
KCNE1, also known as minK, is a member of the KCNE family of membrane proteins that modulate the function of KCNQ1 and certain other voltage-gated potassium channels (KV). Mutations in human KCNE1 cause congenital deafness and congenital long QT syndrome, an inherited predisposition to potentially life-threatening cardiac arrhythmias. Although its modulation of KCNQ1 function has been extensively characterized, many questions remain regarding KCNE1''s structure and location within the channel complex. In this study, KCNE1 was overexpressed in Escherichia coli and purified. Micellar solutions of the protein were then microinjected into Xenopus oocytes expressing KCNQ1 channels, followed by electrophysiological recordings aimed at testing whether recombinant KCNE1 can co-assemble with the channel. Nativelike modulation of channel properties was observed following injection of KCNE1 in lyso-myristoylphosphatidylglycerol (LMPG) micelles, indicating that KCNE1 is not irreversibly misfolded and that LMPG is able to act as a vehicle for delivering membrane proteins into the membranes of viable cells. 1H-15N TROSY NMR experiments indicated that LMPG micelles are well-suited for structural studies of KCNE1, leading to assignment of its backbone resonances and to relaxation studies. The chemical shift data confirmed that KCNE1''s secondary structure includes several alpha-helices and demonstrated that its distal C-terminus is disordered. Surprisingly, for KCNE1 in LMPG micelles, there appears to be a break in alpha-helicity at sites 59-61, near the middle of the transmembrane segment, a feature that is accompanied by increased local backbone mobility. Given that this segment overlaps with sites 57-59, which are known to play a critical role in modulating KCNQ1 channel activation kinetics, this unusual structural feature likely has considerable functional relevance.
PubMed ID: 17892302
PMC ID: PMC2565491
Article link: Biochemistry.
Grant support: HL077188 NHLBI NIH HHS , P41 GM066340 NIGMS NIH HHS , R01 DC007416 NIDCD NIH HHS , R01 DC007416-01A1 NIDCD NIH HHS , R01 HL077188-01A1 NHLBI NIH HHS , R01 DC007416-07 NIDCD NIH HHS , R01 HL077188 NHLBI NIH HHS
Genes referenced: kcne1 kcnq1