Extracellular interaction of the voltage-dependent Ca2+ channel alpha2delta and alpha1 subunits.
The role of the extracellular domain of the voltage-dependent Ca2+ channel alpha2delta subunit in assembly with the alpha1C subunit was investigated. Transiently transfected tsA201 cells processed the alpha2delta subunit properly as disulfide linkages and cleavage sites between the alpha2 and delta subunits were shown to be similar to native channel protein. Coimmunoprecipitation experiments demonstrated that in the absence of delta subunits, alpha2 subunits do not assemble with alpha1 subunits. Furthermore, the transmembrane and cytoplasmic sequences in delta can be exchanged with those of an unrelated protein without any effect on the association between the alpha2delta and alpha1 proteins. Extracellular domains of the alpha2delta 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 alpha1C subunit. Investigation of the corresponding interaction site on the alpha1 subunit revealed that although tryptic peptides containing repeat III of native alpha1S subunit remain in association with the alpha2delta subunit during wheat germ agglutinin chromatography, repeat III by itself is not sufficient for assembly with the alpha2delta subunit. Our results suggest that the alpha2delta subunit likely interacts with more than one extracellular loop of the alpha1 subunit.
Gurnett
C A
CA
Department of Physiology and Biophysics, Howard Hughes Medical Institute, University of Iowa College of Medicine, Iowa City, Iowa 52242, USA.
Felix
R
R
Campbell
K P
KP
D000818:Animals; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D015220:Calcium Channels / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D002460:Cell Line; D000939:Epitopes / Q000032:analysis; D006801:Humans; D017275:Isradipine / Q000378:metabolism; D007668:Kidney; D046911:Macromolecular Substances; D008322:Mammals; D018482:Muscle, Skeletal / Q000378:metabolism*; D010446:Peptide Fragments / Q000302:isolation & purification; D010449:Peptide Mapping; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D014162:Transfection; D014357:Trypsin
D000911:Antibodies, Monoclonal; D015220:Calcium Channels; D000939:Epitopes; D046911:Macromolecular Substances; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014357:Trypsin; D017275:Isradipine