Interaction of T-type calcium channel Ca(V)3.3 with the β-subunit.
The β-subunit of high-voltage-activated (HVA) calcium channels is essential for the regulation of expression and gating. On the other hand, various reports have suggested that β subunits play no role in the regulation of low-voltage-activated T-type channels. In addition there has been no clear demonstration of a physical interaction between the α-subunit of T-type channel with β-subunit. In this study, we systematically investigated the interaction between Ca(V)α and Ca(V)β. The four Ca(V)β isoforms were expressed in a bacterial system and purified into homogeneity, whereas the ten types of Ca(V)α alpha interaction domain (AID) peptides were chemically synthesized. All possible combinations of Ca(V)α and Ca(V)β were then tested for by in vitro immunoassays. We describe here the identification of a new interaction between Ca(V)3.3 and Ca(V)β proteins. This interaction is of low affinity compared to that between the AID of the HVA α-subunit and the alpha-binding pocket (ABP) site of the β-subunit. The AID peptide of HVA channel exerted no effect on the Ca(V)3.3-Ca(V)β interaction, thus demonstrating that another site not in the ABP of Ca(V)β protein played a role in binding with Ca(V)3.3. This is the first demonstration of an α-β subunit interaction in a T-type calcium channel.
Bae
Jinhee
J
Life Sciences Division, Korea Institute of Science and Technology, Seoul, 136-791, Korea.
Suh
Eui Jin
EJ
Lee
Cheolju
C
D000818:Animals; D020747:Calcium Channels, T-Type / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D003001:Cloning, Molecular; D020650:Combinatorial Chemistry Techniques; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000235:genetics; D021122:Protein Subunits / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020349:Surface Plasmon Resonance
C469757:CACNA1I protein, human; D020747:Calcium Channels, T-Type; D010446:Peptide Fragments; D021122:Protein Subunits; D011994:Recombinant Proteins