title: Extracellular conserved cysteine forms an intersubunit disulphide bridge in the KCNK5 (TASK-2) K+ channel without having an essential effect upon activity.
abstract: The functional channel unit of K(+) channels with two pore regions in tandem is thought to be a homodimer and it has been suggested that this dimeric structure occurs by interaction of an extracellular domain, the self-interacting domain. Interaction and functional assembly have been studied in some detail for KCNK1. It is proposed that a disulphide bond between highly conserved C69 residues of the self-interacting domain is formed which is essential for channel activity. We mutated C51, the equivalent residue in the pH-dependent KCNK5, to study its effect on channel function. Western analysis of proteins from cells expressing epitope-tagged KCNK5 and KCNK5-C51S was consistent with reduction-sensitive self-association of monomers dependent upon the presence of C51. Patch-clamp analysis of heterologously expressed KCNK5-C51S, however, revealed it was functional and indistinguishable in rectification properties and pH dependence from the non-mutated channel. The same result was found with KCNK5-C115S. It is concluded that the proposed disulphide bond between cysteine 51 residues of KCNK5 subunits does occur and preserves a dimeric structure in the detergent solubilized complex. Functional assays, on the other hand, suggest that such a disulphide bridge is not essential for correct functional expression.
authors.affiliation: Centro de Estudios Científicos (CECS), Valdivia, Chile. miniemeyer@cecs.cl
authors.forename: Maria Isabel
authors.initials: MI
authors.lastname: Niemeyer
authors.forename: L Pablo
authors.initials: LP
authors.lastname: Cid
authors.forename: Ximena
authors.initials: X
authors.lastname: Valenzuela
authors.forename: Verónica
authors.initials: V
authors.lastname: Paeile
authors.forename: Francisco V
authors.initials: FV
authors.lastname: Sepúlveda
mesh_terms: D017124:Conserved Sequence; D003545:Cysteine / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism*; D007700:Kinetics; D008564:Membrane Potentials; D009154:Mutation; D015221:Potassium Channels / Q000737:chemistry*; D024683:Potassium Channels, Tandem Pore Domain; D013329:Structure-Activity Relationship
chemical_list: D004220:Disulfides; C410295:KCNK5 protein, human; D015221:Potassium Channels; D024683:Potassium Channels, Tandem Pore Domain; D003545:Cysteine