Extracellular interaction of the voltage-dependent Ca2+ channel α2δ and α1 subunits

Abstract

The role of the extracellular domain of the voltage-dependent Ca2+ channel α2δ subunit in assembly with the α(1C) subunit was investigated. Transiently transfected tsA201 cells processed the α2δ subunit properly as disulfide linkages and cleavage sites between the α2 and subunits were shown to be similar to native channel protein. Coimmunoprecipitation experiments demonstrated that in the absence of δ subunits, α2 subunits do not assemble with α1 subunits. Furthermore, the transmembrane and cytoplasmic sequences in δ can be exchanged with those of an unrelated protein without any effect on the association between the α2δ and α1 proteins. Extracellular domains of the α2δ subunit are also shown to be responsible for increasing the binding affinity of [3H]PN200-110 (isopropyl- 4-(2,1,3-benzoxadiazol-4-yl)-1,4-dihydro-2,6-dimethyl-5- ([3H)methoxycarbonyl)-pyridine-3-carboxylate) for the α(1C) subunit. Investigation of the corresponding interaction site on the α1 subunit revealed that although tryptic peptides containing repeat III of native α(1S) subunit remain in association with the α2δ subunit during wheat germ agglutinin chromatography, repeat III by itself is not sufficient for assembly with the α2δ subunit. Our results suggest that the α2δ subunit likely interacts with more than one extracellular loop of the α1 subunit.