title: Voltage-Dependent Inactivation of MscS Occurs Independently of the Positively Charged Residues in the Transmembrane Domain.
abstract: MscS (mechanosensitive channel of small conductance) is ubiquitously found among bacteria and plays a major role in avoiding cell lysis upon rapid osmotic downshock. The gating of MscS is modulated by voltage, but little is known about how MscS senses membrane potential. Three arginine residues (Arg-46, Arg-54, and Arg-74) in the transmembrane (TM) domain are possible to respond to voltage judging from the MscS structure. To examine whether these residues are involved in the voltage dependence of MscS, we neutralized the charge of each residue by substituting with asparagine (R46N, R54N, and R74N). Mechanical threshold for the opening of the expressed wild-type MscS and asparagine mutants did not change with voltage in the range from -40 to +100 mV. By contrast, inactivation process of wild-type MscS was strongly affected by voltage. The wild-type MscS inactivated at +60 to +80 mV but not at -60 to +40 mV. The voltage dependence of the inactivation rate of all mutants tested, that is, R46N, R54N, R74N, and R46N/R74N MscS, was almost indistinguishable from that of the wild-type MscS. These findings indicate that the voltage dependence of the inactivation occurs independently of the positive charges of R46, R54, and R74.
authors.affiliation: International Cooperative Research Project (ICORP)/Solution Oriented Research for Science and Technology (SORST), Cell-Mechanosensing Project, Japan Science and Technology Agency, Nagoya 466-8550, Japan; Department of Rehabilitation, Kyushu Nutrition Welfare University, Kitakyushu 800-029, Japan.
authors.forename: Takeshi
authors.identifier: 0000-0001-8955-3426
authors.initials: T
authors.lastname: Nomura
authors.affiliation: International Cooperative Research Project (ICORP)/Solution Oriented Research for Science and Technology (SORST), Cell-Mechanosensing Project, Japan Science and Technology Agency, Nagoya 466-8550, Japan; Mechanobiology Laboratory, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Sokabe
authors.affiliation: International Cooperative Research Project (ICORP)/Solution Oriented Research for Science and Technology (SORST), Cell-Mechanosensing Project, Japan Science and Technology Agency, Nagoya 466-8550, Japan; Department of Machinery and Control Systems, Shibaura Institute of Technology, Saitama 337-8570, Japan.
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Yoshimura
mesh_terms: D019943:Amino Acid Substitution; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D007473:Ion Channels / Q000235:genetics / Q000378:metabolism*; D008564:Membrane Potentials / Q000502:physiology*; D020125:Mutation, Missense; D000072417:Protein Domains
chemical_list: D029968:Escherichia coli Proteins; D007473:Ion Channels; C469542:MscS protein, E coli