Scanning mutagenesis of the I-II loop of the Cav2.2 calcium channel identifies residues Arginine 376 and Valine 416 as molecular determinants of voltage dependent G protein inhibition

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dc.contributor.authorTedford, Hugo William
dc.contributor.authorKisilevsky, Alexandra E.
dc.contributor.authorVieira, Luciene Bruno
dc.contributor.authorVarela, Diego L.
dc.contributor.authorChen, Lina
dc.contributor.authorZamponi, Gerald W.
dc.date.accessioned2018-05-24T20:26:25Z
dc.date.available2018-05-24T20:26:25Z
dc.date.issued2010-02-03
dc.description.abstractDirect interaction with the beta subunit of the heterotrimeric G protein complex causes voltage-dependent inhibition of N-type calcium channels. To further characterize the molecular determinants of this interaction, we performed scanning mutagenesis of residues 372-387 and 410-428 of the N-type channel alpha1 subunit, in which individual residues were replaced by either alanine or cysteine. We coexpressed wild type Gbeta1gamma2 subunits with either wild type or point mutant N-type calcium channels, and voltage-dependent, G protein-mediated inhibition of the channels (VDI) was assessed using patch clamp recordings. The resulting data indicate that Arg376 and Val416 of the alpha1 subunit, residues which are surface-exposed in the presence of the calcium channel beta subunit, contribute significantly to the functional inhibition by Gbeta1. To further characterize the roles of Arg376 and Val416 in this interaction, we performed secondary mutagenesis of these residues, coexpressing the resulting mutants with wild type Gbeta1gamma2 subunits and with several isoforms of the auxiliary beta subunit of the N-type channel, again assessing VDI using patch clamp recordings. The results confirm the importance of Arg376 for G protein-mediated inhibition and show that a single amino acid substitution to phenylalanine drastically alters the abilities of auxiliary calcium channel subunits to regulate G protein inhibition of the channel.en_US
dc.identifier.citationTedford, H. W., Kisilevsky, A. E., Vieira, L. B., Varela, D., Chen, L., & Zamponi, G. W. (2010). Scanning mutagenesis of the I-II loop of the Cav2.2 calcium channel identifies residues Arginine 376 and Valine 416 as molecular determinants of voltage dependent G protein inhibition. Molecular Brain, 3(1). https://doi.org/10.1186/1756-6606-3-6en_US
dc.identifier.doihttp://dx.doi.org/10.1186/1756-6606-3-6en_US
dc.identifier.urihttp://hdl.handle.net/1880/106668
dc.language.isoenen_US
dc.publisherBioMed Central Ltd.en_US
dc.publisher.departmentPhysiology & Pharmacologyen_US
dc.publisher.facultyCumming School of Medicineen_US
dc.publisher.institutionUniversity of Calgaryen_US
dc.rights.urihttps://creativecommons.org/licenses/by/4.0en_US
dc.titleScanning mutagenesis of the I-II loop of the Cav2.2 calcium channel identifies residues Arginine 376 and Valine 416 as molecular determinants of voltage dependent G protein inhibitionen_US
dc.typeunknown
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