Differential interactions of the C terminus and the cytoplasmic I-II loop of neuronal Ca2+ channels with G-protein α and βγ subunits II. Evidence for direct binding

Abstract

The present study was designed to obtain evidence for direct interactions of G-protein α (Gα) and βγ subunits (Gβγ) with N- (α(1B)) and P/Q-type (α(1A)) Ca2+ channels, using synthetic peptides and fusion proteins derived from loop 1 (cytoplasmic loop between repeat I and II) and the C terminus of these channels. For N-type, prepulse facilitation as mediated by Gβγ was impaired when a synthetic loop 1 peptide was applied intracellularly. Receptor agonist-induced inhibition of N-type as mediated by Gα was also impaired by the loop 1 peptide but only when applied in combination with a C-terminal peptide. For P/Q-type channels, by contrast, the Gα-mediated inhibition was diminished by application of a C-terminal peptide alone. Moreover, in vitro binding analysis for N- and P/Q-type channels revealed direct interaction of Gα with C-terminal fusion proteins as well as direct interaction of Gβγ with loop 1 fusion proteins. These findings define loop I of N- and P/Q-type Ca2+ channels as an interaction site for Gβγ and the C termini for Gα.