PMID- 21376840
OWN - NLM
STAT- MEDLINE
DCOM- 20120118
LR  - 20161222
IS  - 1556-3871 (Electronic)
IS  - 1547-5271 (Linking)
VI  - 8
IP  - 8
DP  - 2011 Aug
TI  - Functional characterization of the LQT2-causing mutation R582C and the associated 
      voltage-dependent fluorescence signal.
PG  - 1273-80
LID - 10.1016/j.hrthm.2011.02.035 [doi]
AB  - BACKGROUND: The R582C mutation is one of many Long-QT Syndrome type 2 
      (LQT2)-causing mutations localized to the human ether-a-go-go related gene (hERG) 
      channel's S5-P linker subdomain, yet its specific mechanism of dysfunction has 
      not been examined. OBJECTIVE: This study sought to characterize the biophysical 
      properties of the congenital LQT2-causing mutation, R582C, and utilize this 
      mutation to provide the first report of voltage-dependent fluorescence from the 
      S5-P linker. METHODS: Properties of the R582C channels were characterized by 
      heterologous expression in both HEK293 cells and Xenopus oocytes using a 
      combination of patch-clamp, 2-electrode voltage-clamp, immunoblot assay, and 
      voltage-clamp fluorimetry. RESULTS: Expression of hERG R582C was found to be 
      deficient in HEK293 cells, yet was amenable to rescue by incubation at reduced 
      temperature or by treatment with dofetilide. Rescued channels expressed at levels 
      comparable to wild type (WT) channels. Kinetic differences result in decreased 
      outward repolarizing current evoked by an action potential clamp protocol. 
      Voltage-clamp fluorimetry experiments utilized the introduced cysteine to 
      covalently attach a fluorescent probe (tetramethylrhodamine-5-maleimide) to the 
      S5-P linker to directly observe conformational changes occurring due to 
      inactivation. CONCLUSION: The major mechanism underlying pathogenicity of the 
      R582C mutation is a trafficking deficiency, although channels also exhibit 
      kinetic deficiencies, perhaps reflecting the position of the mutation in the pore 
      turret. Voltage clamp fluorescence signals from R582C channels provide evidence 
      that the hERG turret undergoes distinct conformational changes during 
      inactivation.
CI  - Copyright (c) 2011 Heart Rhythm Society. Published by Elsevier Inc. All rights 
      reserved.
FAU - Fougere, Robert R
AU  - Fougere RR
AD  - Department of Anesthesiology, Pharmacology and Therapeutics, University of 
      British Columbia, Canada. fedida@interchange.ubc.ca
FAU - Es-Salah-Lamoureux, Zeineb
AU  - Es-Salah-Lamoureux Z
FAU - Rezazadeh, Saman
AU  - Rezazadeh S
FAU - Eldstrom, Jodene
AU  - Eldstrom J
FAU - Fedida, David
AU  - Fedida D
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20110303
PL  - United States
TA  - Heart Rhythm
JT  - Heart rhythm
JID - 101200317
RN  - 0 (DNA, Complementary)
RN  - 0 (ERG1 Potassium Channel)
RN  - 0 (Ether-A-Go-Go Potassium Channels)
RN  - 0 (KCNH2 protein, human)
RN  - 0 (Phenethylamines)
RN  - 0 (Potassium Channel Blockers)
RN  - 0 (Sulfonamides)
RN  - R4Z9X1N2ND (dofetilide)
SB  - IM
MH  - DNA, Complementary/genetics
MH  - ERG1 Potassium Channel
MH  - Ether-A-Go-Go Potassium Channels/*genetics
MH  - Fluorometry
MH  - HEK293 Cells
MH  - Humans
MH  - Long QT Syndrome/congenital/*genetics
MH  - Mutation
MH  - Patch-Clamp Techniques/methods
MH  - Phenethylamines/pharmacology
MH  - Potassium Channel Blockers/pharmacology
MH  - Protein Transport
MH  - Sulfonamides/pharmacology
MH  - Transfection
EDAT- 2011/03/08 06:00
MHDA- 2012/01/19 06:00
CRDT- 2011/03/08 06:00
PHST- 2010/08/20 00:00 [received]
PHST- 2011/02/24 00:00 [accepted]
PHST- 2011/03/08 06:00 [entrez]
PHST- 2011/03/08 06:00 [pubmed]
PHST- 2012/01/19 06:00 [medline]
AID - S1547-5271(11)00256-6 [pii]
AID - 10.1016/j.hrthm.2011.02.035 [doi]
PST - ppublish
SO  - Heart Rhythm. 2011 Aug;8(8):1273-80. doi: 10.1016/j.hrthm.2011.02.035. Epub 2011 
      Mar 3.