NMR solution structure of the N-terminal domain of hERG and its interaction with the S4–S5 linker

► The N-terminal domain (NTD, eag domain) containing 135 residues of hERG was expressed and purified from E. coli cells. ► Solution structure of NTD was determined with NMR spectroscopy. ► The alpha-helical region (residues 13–23) was demonstrated to possess the characteristics of an amphipathic hel...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 2010-12, Vol.403 (1), p.126-132
Main Authors: Li, Qingxin, Gayen, Shovanlal, Chen, Angela Shuyi, Huang, Qiwei, Raida, Manfred, Kang, CongBao
Format: Article
Language:English
Subjects:
60 APPLIED LIFE SCIENCES
Amphipathic helix
CHROMATOGRAPHY
DICHROISM
EDTA
ELECTROPHORESIS
ESCHERICHIA COLI
Ether-A-Go-Go Potassium Channels - chemistry
ETHERS
GELS
GENES
hERG
Humans
MONOMERS
MUTATIONS
NMR spectroscopy
NUCLEAR MAGNETIC RESONANCE
Nuclear Magnetic Resonance, Biomolecular
PAS domain
PEPTIDES
Peptides - chemistry
POTASSIUM
Potassium channel
Protein Structure, Secondary
Protein Structure, Tertiary
PURIFICATION
RECTIFIERS
SODIUM
SPECTROSCOPY
SULFATES
TITRATION
TRANSLOCATION
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Summary:► The N-terminal domain (NTD, eag domain) containing 135 residues of hERG was expressed and purified from E. coli cells. ► Solution structure of NTD was determined with NMR spectroscopy. ► The alpha-helical region (residues 13–23) was demonstrated to possess the characteristics of an amphipathic helix. ► NMR titration confirmed the interaction between NTD and the peptide from the S4–S5 linker. The human Ether-à-go-go Related Gene (hERG) potassium channel mediates the rapid delayed rectifier current (IKr) in the cardiac action potential. Mutations in the 135 amino acid residue N-terminal domain (NTD) cause channel dysfunction or mis-translocation. To study the structure of NTD, it was overexpressed and purified from Escherichia coli cells using affinity purification and gel filtration chromatography. The purified protein behaved as a monomer under purification conditions. Far- and near-UV, circular dichroism (CD) and solution nuclear magnetic resonance (NMR) studies showed that the purified protein was well-folded. The solution structure of NTD was obtained and the N-terminal residues 13–23 forming an amphipathic helix which may be important for the protein–protein or protein-membrane interactions. NMR titration experiment also demonstrated that residues from 88 to 94 in NTD are important for the molecular interaction with the peptide derived from the S4–S5 linker.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2010.10.132