title: Arrangement and mobility of the voltage sensor domain in prokaryotic voltage-gated sodium channels.
abstract: Prokaryotic voltage-gated sodium channels (Na(V)s) form homotetramers with each subunit contributing six transmembrane α-helices (S1-S6). Helices S5 and S6 form the ion-conducting pore, and helices S1-S4 function as the voltage sensor with helix S4 thought to be the essential element for voltage-dependent activation. Although the crystal structures have provided insight into voltage-gated K channels (K(V)s), revealing a characteristic domain arrangement in which the voltage sensor domain of one subunit is close to the pore domain of an adjacent subunit in the tetramer, the structural and functional information on Na(V)s remains limited. Here, we show that the domain arrangement in NaChBac, a firstly cloned prokaryotic Na(V), is similar to that in K(V)s. Cysteine substitutions of three residues in helix S4, Q107C, T110C, and R113C, effectively induced intersubunit disulfide bond formation with a cysteine introduced in helix S5, M164C, of the adjacent subunit. In addition, substituting two acidic residues with lysine, E43K and D60K, shifted the activation of the channel to more positive membrane potentials and consistently shifted the preferentially formed disulfide bond from T110C/M164C to Q107C/M164C. Because Gln-107 is located closer to the extracellular side of helix S4 than Thr-110, this finding suggests that the functional shift in the voltage dependence of activation is related to a restriction of the position of helix S4 in the lipid bilayer. The domain arrangement and vertical mobility of helix S4 in NaChBac indicate that the structure and the mechanism of voltage-dependent activation in prokaryotic Na(V)s are similar to those in canonical K(V)s.
authors.affiliation: Department of Biophysics, Graduate School of Science, Kyoto University, Oiwake, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan.
authors.forename: Takushi
authors.initials: T
authors.lastname: Shimomura
authors.forename: Katsumasa
authors.initials: K
authors.lastname: Irie
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Nagura
authors.forename: Tomoya
authors.initials: T
authors.lastname: Imai
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Fujiyoshi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001407:Bacillus / Q000502:physiology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004220:Disulfides / Q000737:chemistry; D055664:Electrochemical Techniques; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology*; D008564:Membrane Potentials / Q000502:physiology; D008969:Molecular Sequence Data; D016296:Mutagenesis / Q000502:physiology; D017434:Protein Structure, Tertiary; D012964:Sodium / Q000378:metabolism*; D015222:Sodium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004220:Disulfides; C443666:NaChBac protein, bacteria; D015222:Sodium Channels; D012964:Sodium