title: Charged Residues at the First Transmembrane Region Contribute to the Voltage Dependence of the Slow Gate of Connexins.
abstract: Connexins (Cxs) are a family of membrane-spanning proteins that form gap junction channels and hemichannels. Connexin-based channels exhibit two distinct voltage-dependent gating mechanisms termed slow and fast gating. Residues located at the C terminus of the first transmembrane segment (TM-1) are important structural components of the slow gate. Here, we determined the role of the charged residues at the end of TM-1 in voltage sensing in Cx26, Cx46, and Cx50. Conductance/voltage curves obtained from tail currents together with kinetics analysis reveal that the fast and slow gates of Cx26 involves the movement of two and four charges across the electric field, respectively. Primary sequence alignment of different Cxs shows the presence of well conserved glutamate residues in the C terminus of TM-1; only Cx26 contains a lysine in that position (lysine 41). Neutralization of lysine 41 in Cx26 increases the voltage dependence of the slow gate. Swapping of lysine 41 with glutamate 42 maintains the voltage dependence. In Cx46, neutralization of negative charges or addition of a positive charge in the Cx26 equivalent region reduced the slow gate voltage dependence. In Cx50, the addition of a glutamate in the same region decreased the voltage dependence, and the neutralization of a negative charge increased it. These results indicate that the charges at the end of TM-1 are part of the slow gate voltage sensor in Cxs. The fact that Cx42, which has no charge in this region, still presents voltage-dependent slow gating suggests that charges still unidentified also contribute to the slow gate voltage sensitivity.
authors.affiliation: From the Centro Interdisciplinario de Neurociencias de Valparaíso, Universidad de Valparaíso, Valparaíso 2360102 and.
authors.forename: Bernardo I
authors.initials: BI
authors.lastname: Pinto
authors.affiliation: From the Centro Interdisciplinario de Neurociencias de Valparaíso, Universidad de Valparaíso, Valparaíso 2360102 and.
authors.forename: Isaac E
authors.initials: IE
authors.lastname: García
authors.affiliation: From the Centro Interdisciplinario de Neurociencias de Valparaíso, Universidad de Valparaíso, Valparaíso 2360102 and.
authors.forename: Amaury
authors.initials: A
authors.lastname: Pupo
authors.affiliation: the Centro de Fisiología Celular e Integrativa, Facultad de Medicina, Clínica Alemana Universidad del Desarrollo, Santiago 7710162, Chile.
authors.forename: Mauricio A
authors.initials: MA
authors.lastname: Retamal
authors.affiliation: From the Centro Interdisciplinario de Neurociencias de Valparaíso, Universidad de Valparaíso, Valparaíso 2360102 and.
authors.forename: Agustín D
authors.initials: AD
authors.lastname: Martínez
authors.affiliation: From the Centro Interdisciplinario de Neurociencias de Valparaíso, Universidad de Valparaíso, Valparaíso 2360102 and ramon.latorre@uv.cl.
authors.forename: Ramón
authors.initials: R
authors.lastname: Latorre
authors.affiliation: From the Centro Interdisciplinario de Neurociencias de Valparaíso, Universidad de Valparaíso, Valparaíso 2360102 and carlos.gonzalezl@uv.cl.
authors.forename: Carlos
authors.initials: C
authors.lastname: González
mesh_terms: D000818:Animals; D030161:Avian Proteins / Q000235:genetics / Q000378:metabolism*; D002645:Chickens; D017630:Connexins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology*; D008564:Membrane Potentials / Q000502:physiology*; D000072417:Protein Domains; D051381:Rats; D014982:Xenopus laevis
chemical_list: D030161:Avian Proteins; D017630:Connexins
keywords: biophysics; connexin; connexon (hemichannel); electrophysiology; ion channel; kinetics; voltage dependency