RESUMO
Ca(V) channels are multi-subunit protein complexes that enable inward cellular Ca(2+) currents in response to membrane depolarization. We recently described structure-function studies of the intracellular α1 subunit domain I-II linker, directly downstream of domain IS6. The results show the extent of the linker's helical structure to be subfamily dependent, as dictated by highly conserved primary sequence differences. Moreover, the difference in structure confers different biophysical properties, particularly the extent and kinetics of voltage and calcium-dependent inactivation. Timothy syndrome is a human genetic disorder due to mutations in the Ca(V)1.2 gene. Here, we explored whether perturbation of the I-II linker helical structure might provide a mechanistic explanation for a Timothy syndrome mutant's (human Ca(V)1.2 G406R equivalent) biophysical effects on inactivation and activation. The results are equivocal, suggesting that a full mechanistic explanation for this Timothy syndrome mutation requires further investigation.
Assuntos
Canais de Cálcio Tipo L/química , Canais de Cálcio Tipo L/metabolismo , Síndrome do QT Longo/metabolismo , Sindactilia/metabolismo , Animais , Transtorno Autístico , Canais de Cálcio Tipo L/genética , Humanos , Ativação do Canal Iônico , Síndrome do QT Longo/genética , Síndrome do QT Longo/fisiopatologia , Mutação/genética , Estrutura Secundária de Proteína , Relação Estrutura-Atividade , Sindactilia/genética , Sindactilia/fisiopatologia , XenopusRESUMO
Voltage-dependent calcium channels (VDCCs) allow the passage of Ca(2+) ions through cellular membranes in response to membrane depolarization. The channel pore-forming subunit, α1, and a regulatory subunit (Ca(V)ß) form a high affinity complex where Ca(V)ß binds to a α1 interacting domain in the intracellular linker between α1 membrane domains I and II (I-II linker). We determined crystal structures of Ca(V)ß2 functional core in complex with the Ca(V)1.2 and Ca(V)2.2 I-II linkers to a resolution of 1.95 and 2.0 Å, respectively. Structural differences between the highly conserved linkers, important for coupling Ca(V)ß to the channel pore, guided mechanistic functional studies. Electrophysiological measurements point to the importance of differing linker structure in both Ca(V)1 and 2 subtypes with mutations affecting both voltage- and calcium-dependent inactivation and voltage dependence of activation. These linker effects persist in the absence of Ca(V)ß, pointing to the intrinsic role of the linker in VDCC function and suggesting that I-II linker structure can serve as a brake during inactivation.
