G protein β₂ subunit-derived peptides for inhibition and induction of G protein pathways: Examination of voltage-gated Ca²⁺ and G protein inwardly rectifying K⁺ channels

Voltage-gated Ca²⁺ channels of the N-, P/Q-, and R-type and G protein inwardly rectifying K⁺ channels (GIRK) are modulated via direct binding of G proteins. The modulation is mediated by G protein βγ subunits. By using electrophysiological recordings and fluorescence resonance energy transfer, we characterized the modulatory domains of the G protein β subunit on the recombinant P/Q-type channel and GIRK channel expressed in HEK293 cells and on native non-L-type Ca²⁺ currents of cultured hippocampal neurons. We found that Gβ₂ subunit-derived deletion constructs and synthesized peptides can either induce or inhibit G protein modulation of the examined ion channels. In particular, the 25-amino acid peptide derived from the Gβ₂ N terminus inhibits G protein modulation, whereas a 35-amino acid peptide derived from the Gβ₂ C terminus induced modulation of voltage-gated Ca ²⁺ channels and GIRK channels. Fluorescence resonance energy transfer (FRET) analysis of the live action of these peptides revealed that the 25-amino acid peptide diminished the FRET signal between G protein β ₂γ₃ subunits, indicating a reorientation between G protein β₂γ₃ subunits in the presence of the peptide. In contrast, the 35-amino acid peptide increased the FRET signal between GIRK1,2 channel subunits, similarly to the Gβγ-mediated FRET increase observed for this GIRK subunit combination. Circular dichroism spectra of the synthesized peptides suggest that the 25-amino acid peptide is structured. These results indicate that individual G protein β subunit domains can act as independent, separate modulatory domains to either induce or inhibit G protein modulation for several effector proteins.