Roles of surface residues of intracellular domains of heag potassium channels

• 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
• ISSN: 0175-7571; 1432-1017
• DOI: 10.1007/s00249-009-0402-8

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.