HERG eag/PAS (ether-á-go-go/Per-Arnt-Sim) Domain: Elucidating its Role in Regulating Channel Gating in Physiology and Disease.
|dc.contributor.author||Gianulis, Elena Constance|
|dc.description||University of Maryland, Baltimore. Molecular Physiology and Pharmacology. Ph.D. 2012||en_US|
|dc.description.abstract||The human ether-á-go-go related gene (hERG) voltage-gated potassium channel plays a critical role in cardiac repolarization. hERG exhibits unique kinetic properties, including fast inactivation and slow deactivation, which make it well suited for its role in the heart. Long QT Syndrome Type 2 (LQT2) is caused by loss of hERG function, which can degenerate into ventricular arrhythmias and sudden death. It is well established that the N-terminal ether-á-go-go/Per-Arnt-Sim (eag/PAS) domain is an essential regulator of hERG's characteristic slow deactivation kinetics. However, the precise mechanisms by which the eag/PAS domain regulates gating, as well as the functional effects of LQT2 mutations located in the domain, remained unclear. In this work, I showed that hERG channels containing LQT2 mutations within a common region of the eag/PAS domain structure exhibited altered kinetic properties, revealing a putative "gating face" that is important in eag/PAS domain regulation of gating. Co-expression with a wild-type eag/PAS domain rescued the gating-deficient phenotype in hERG PAS-LQT2 channels, presenting a potential biological therapeutic for LQT2. Previous groups have proposed a role for the S4-S5 linker in eag/PAS domain regulation of deactivation. Using alanine scanning mutagenesis, including complete replacement of the S4-S5 linker sequence with alanine residues, I showed that the S4-S5 linker was not required for eag/PAS domain association with the channel - as measured by FRET spectroscopy - but was involved in eag/PAS domain regulation of slow deactivation. I tested the hypothesis that the C-terminal CNBHD is the primary direct site of interaction for the eag/PAS domain. hERG channels with deletion of the CNBHD resulted in a loss of regulation of deactivation by an eag/PAS domain gene fragment, as well as a lack of a FRET signal between the channel and the eag/PAS domain fragment. Further, a FRET-based peptide hybridization assay between isolated eag/PAS domains and CNBHDs revealed a specific and direct interaction between the two domains, providing strong evidence that the CNBHD is the primary site of interaction for the eag/PAS domain. Altogether, this work provides a greater understanding of the mechanisms underlying the hERG channel's unique kinetic properties, as well as insights into the disease phenotype of LQT2.||en_US|
|dc.title||HERG eag/PAS (ether-á-go-go/Per-Arnt-Sim) Domain: Elucidating its Role in Regulating Channel Gating in Physiology and Disease.||en_US|
|dc.title.alternative||HERG eag/PAS (ether-a-go-go/Per-Arnt-Sim) Domain: Elucidating its Role in Regulating Channel Gating in Physiology and Disease|
|dc.contributor.advisor||Trudeau, Matthew C.|