title: Structural characteristics of the redox-sensing coiled coil in the voltage-gated H+ channel.
abstract: Oxidation is an important biochemical defense mechanism, but it also elicits toxicity; therefore, oxidation must be under strict control. In phagocytotic events in neutrophils, the voltage-gated H(+) (Hv) channel is a key regulator of the production of reactive oxygen species against invading bacteria. The cytoplasmic domain of the Hv channel forms a dimeric coiled coil underpinning a dimerized functional unit. Importantly, in the alignment of the coiled-coil core, a conserved cysteine residue forms a potential intersubunit disulfide bond. In this study, we solved the crystal structures of the coiled-coil domain in reduced, oxidized, and mutated (Cys → Ser) states. The crystal structures indicate that a pair of Cys residues forms an intersubunit disulfide bond dependent on the redox conditions. CD spectroscopy revealed that the disulfide bond increases the thermal stability of the coiled-coil protein. We also reveal that two thiol modifier molecules are able to bind to Cys in a redox-dependent manner without disruption of the dimeric coiled-coil assembly. Thus, the biochemical properties of the cytoplasmic coiled-coil domain in the Hv channel depend on the redox condition, which may play a role in redox sensing in the phagosome.
authors.affiliation: Department of Physiology, Graduate School of Medicine, Osaka University, Osaka 565-0871, Japan. fujiwara@phys2.med.osaka-u.ac.jp
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Fujiwara
authors.forename: Kohei
authors.initials: K
authors.lastname: Takeshita
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nakagawa
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Okamura
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007473:Ion Channels / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D008264:Macrophages / Q000378:metabolism; D008564:Membrane Potentials; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010084:Oxidation-Reduction; D018408:Patch-Clamp Techniques; D010588:Phagosomes / Q000378:metabolism; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000378:metabolism
chemical_list: D004220:Disulfides; C540876:Hv1 proton channel, mouse; D007473:Ion Channels; D013438:Sulfhydryl Compounds
keywords: Biosensors; Coiled Coil; Ion Channel Biophysics; Ion Channel Structure-Function; Ion Channels; Oxidation-Reduction; Protein Structure; Proton Channel; Proton Transport; Redox