title: Role of hydrophobic and ionic forces in the movement of S4 of the Shaker potassium channel.
abstract: Voltage-gated ion (K(+), Na(+), Ca(2+)) channels contain a pore domain (PD) surrounded by four voltage sensing domains (VSD). Each VSD is made up of four transmembrane helices, S1-S4. S4 contains 6-7 positively charged residues (arginine/lysine) separated two hydrophobic residues, whereas S1-S3 contribute to two negatively charged clusters. These structures are conserved among all members of the voltage-gated ion channel family and play essential roles in voltage gating. The role of S4 charged residues in voltage gating is well established: During depolarization, they move out of the membrane electric field, exerting a mechanical force on channel gates, causing them to open. However, the role of the intervening hydrophobic residues in voltage sensing is unclear. Here we studied the role of these residues in the prototypical Shaker potassium channel. We have altered the physicochemical properties of both charged and hydrophobic positions of S4 and examined the effect of these modifications on the gating properties of the channel. For this, we have introduced cysteines at each of these positions, expressed the mutants in Xenopus oocytes, and examined the effect of in situ addition of charge, via Cd(2+), on channel gating by two-electrode voltage clamp. Our results reveal a face of the S4 helix (comprising residues L358, L361, R365 and R368) where introduction of charge at hydrophobic positions destabilises the closed state and removal of charges from charged positions has an opposite effect. We propose that hydrophobic residues play a crucial role in limiting gating to a physiological voltage range.
authors.affiliation: Multidisciplinary Cardiovascular Research Centre, Institute of Membrane and Systems Biology, Faculty of Biological Sciences, University of Leeds, Leeds, UK.
authors.forename: David J S
authors.initials: DJ
authors.lastname: Elliott
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Neale
authors.forename: Tim S
authors.initials: TS
authors.lastname: Munsey
authors.forename: John P
authors.initials: JP
authors.lastname: Bannister
authors.forename: Asipu
authors.initials: A
authors.lastname: Sivaprasadarao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002104:Cadmium / Q000494:pharmacology; D003545:Cysteine / Q000235:genetics; D057927:Hydrophobic and Hydrophilic Interactions / Q000187:drug effects; D015640:Ion Channel Gating / Q000187:drug effects; D007477:Ions; D007700:Kinetics; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry; D021381:Protein Transport / Q000187:drug effects; D051637:Shaker Superfamily of Potassium Channels / Q000737:chemistry* / Q000378:metabolism*; D014981:Xenopus
chemical_list: D007477:Ions; D050505:Mutant Proteins; D021122:Protein Subunits; D051637:Shaker Superfamily of Potassium Channels; D002104:Cadmium; D003545:Cysteine