Roles of surface residues of intracellular domains of heag potassium channels

Ether-a-go-go potassium channels have large intracellular regions containing ‘Per-Ant-Sim’ (PAS) and cyclic nucleotide binding (cNBD) domains at the N- and C-termini, respectively. In heag1 and heag2 channels, recent studies have suggested that the N- and C-terminal domains interact, and affect acti...

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Bibliographic Details
Published in:European biophysics journal 2009-04, Vol.38 (4), p.523-532
Main Authors: Stevens, Louisa, Ju, Min, Wray, Dennis
Format: Article
Language:English
Subjects:
Animals
Binding Sites
Biochemistry
Biological and Medical Physics
Biomedical and Life Sciences
Biophysics
Blotting, Western
Cell Biology
Cellular biology
Conserved Sequence
Electrophoresis, Polyacrylamide Gel
Escherichia coli
Ether-A-Go-Go Potassium Channels - chemistry
Ether-A-Go-Go Potassium Channels - genetics
Ether-A-Go-Go Potassium Channels - metabolism
Humans
Ions
Kinetics
Life Sciences
Membrane Biology
Membrane Potentials
Models, Molecular
Mutation
Mutation, Missense
Nanotechnology
Neurobiology
Original Paper
Patch-Clamp Techniques
Potassium
Studies
Time Factors
Xenopus laevis
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Summary:Ether-a-go-go potassium channels have large intracellular regions containing ‘Per-Ant-Sim’ (PAS) and cyclic nucleotide binding (cNBD) domains at the N- and C-termini, respectively. In heag1 and heag2 channels, recent studies have suggested that the N- and C-terminal domains interact, and affect activation properties. Here, we have studied the effect of mutations of residues on the surfaces of PAS and cNBD domains. For this, we introduced alanine and lysine mutations in heag1 channels, and recorded currents by two-electrode voltage clamp. In both the PAS domain and the cNBD domain, contiguous areas of conserved residues on the surfaces of these domains were found which affected the activation kinetics of the channel. Next, we investigated possible effects of mutations on domain interactions of PAS and cNBD proteins in heag2 by co-expressing these domain proteins followed by analysis with native gels and western blotting. We found oligomeric association between these domains. Mutations F30A and A609K (on the surfaces of the PAS and cNBD domains, respectively) affected oligomeric compositions of these domains when proteins for PAS and cNBD domains were expressed together. Taken together, the data suggest that the PAS and cNBD domains form interacting oligomers that have roles in channel function.
ISSN:0175-7571
1432-1017
DOI:10.1007/s00249-009-0402-8