Identification of a disulfide bridge essential for structure and function of the voltage-gated Ca(2+) channel alpha(2)delta-1 auxiliary subunit.

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Calderon-Rivera A, Andrade A, Hernandez-Hernandez O, Gonzalez-Ramirez R, Sandoval A, Rivera M, Gomora JC, Felix R

Identification of a disulfide bridge essential for structure and function of the voltage-gated Ca(2+) channel alpha(2)delta-1 auxiliary subunit.

Cell Calcium. 2012 Jan;51(1):22-30. doi: 10.1016/j.ceca.2011.10.002. Epub 2011 Nov 3.

PubMed ID
22054663 [ View in PubMed
]
Abstract

Voltage-gated calcium (Ca(V)) channels are transmembrane proteins that form Ca(2+)-selective pores gated by depolarization and are essential regulators of the intracellular Ca(2+) concentration. By providing a pathway for rapid Ca(2+) influx, Ca(V) channels couple membrane depolarization to a wide array of cellular responses including neurotransmission, muscle contraction and gene expression. Ca(V) channels fall into two major classes, low voltage-activated (LVA) and high voltage-activated (HVA). The ion-conducting pathway of HVA channels is the alpha(1) subunit, which typically contains associated beta and alpha(2)delta ancillary subunits that regulate the properties of the channel. Although it is widely acknowledged that alpha(2)delta-1 is post-translationally cleaved into an extracellular alpha(2) polypeptide and a membrane-anchored delta protein that remain covalently linked by disulfide bonds, to date the contribution of different cysteine (Cys) residues to the formation of disulfide bridges between these proteins has not been investigated. In the present report, by predicting disulfide connectivity with bioinformatics, molecular modeling and protein biochemistry experiments we have identified two Cys residues involved in the formation of an intermolecular disulfide bond of critical importance for the structure and function of the alpha(2)delta-1 subunit. Site directed-mutagenesis of Cys404 (located in the von Willebrand factor-A region of alpha(2)) and Cys1047 (in the extracellular domain of delta) prevented the association of the alpha(2) and delta peptides upon proteolysis, suggesting that the mature protein is linked by a single intermolecular disulfide bridge. Furthermore, co-expression of mutant forms of alpha(2)delta-1 Cys404Ser and Cys1047Ser with recombinant neuronal N-type (Ca(V)2.2alpha(1)/beta(3)) channels, showed decreased whole-cell patch-clamp currents indicating that the disulfide bond between these residues is required for alpha(2)delta-1 function.

DrugBank Data that Cites this Article

Polypeptides
NameUniProt ID
Voltage-dependent calcium channel subunit alpha-2/delta-1P54289Details