title: Mutations of nonconserved residues within the calcium channel alpha1-interaction domain inhibit beta-subunit potentiation.
abstract: Voltage-dependent calcium channels consist of a pore-forming subunit (Ca(V)alpha(1)) that includes all the molecular determinants of a voltage-gated channel, and several accessory subunits. The ancillary beta-subunit (Ca(V)beta) is a potent activator of voltage-dependent calcium channels, but the mechanisms and structural bases of this regulation remain elusive. Ca(V)beta binds reversibly to a conserved consensus sequence in Ca(V)alpha(1), the alpha(1)-interaction domain (AID), which forms an alpha-helix when complexed with Ca(V)beta. Conserved aromatic residues face to one side of the helix and strongly interact with a hydrophobic pocket on Ca(V)beta. Here, we studied the effect of mutating residues located opposite to the AID-Ca(V)beta contact surface in Ca(V)1.2. Substitution of AID-exposed residues by the corresponding amino acids present in other Ca(V)alpha(1) subunits (E462R, K465N, D469S, and Q473K) hinders Ca(V)beta's ability to increase ionic-current to charge-movement ratio (I/Q) without changing the apparent affinity for Ca(V)beta. At the single channel level, these Ca(V)1.2 mutants coexpressed with Ca(V)beta(2a) visit high open probability mode less frequently than wild-type channels. On the other hand, Ca(V)1.2 carrying either a mutation in the conserved tryptophan residue (W470S, which impairs Ca(V)beta binding), or a deletion of the whole AID sequence, does not exhibit Ca(V)beta-induced increase in I/Q. In addition, we observed a shift in the voltage dependence of activation by +12 mV in the AID-deleted channel in the absence of Ca(V)beta, suggesting a direct participation of these residues in the modulation of channel activation. Our results show that Ca(V)beta-dependent potentiation arises primarily from changes in the modal gating behavior. We envision that Ca(V)beta spatially reorients AID residues that influence the channel gate. These findings provide a new framework for understanding modulation of VDCC gating by Ca(V)beta.
authors: Centro de Neurociencia de Valparaíso, Universidad deValparaíso, 2349400 Valparaíso, Chile
authors: Giovanni
authors: G
authors: Gonzalez-Gutierrez
authors: Erick
authors: E
authors: Miranda-Laferte
authors: David
authors: D
authors: Naranjo
authors: Patricia
authors: P
authors: Hidalgo
authors: Alan
authors: A
authors: Neely
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D015220:Calcium Channels / Q000235:genetics* / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D017124:Conserved Sequence; D015640:Ion Channel Gating / Q000235:genetics*; D008564:Membrane Potentials / Q000502:physiology*; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology; D021122:Protein Subunits; D014982:Xenopus laevis
chemical_list: D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D021122:Protein Subunits