RT Journal Article SR Electronic T1 Functional Role of a C-Terminal Gβγ-Binding Domain of Cav2.2 Channels JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP 761 OP 769 VO 66 IS 3 A1 Li, Bin A1 Zhong, Huijun A1 Scheuer, Todd A1 Catterall, William A. YR 2004 UL http://molpharm.aspetjournals.org/content/66/3/761.abstract AB Presynaptic Ca2+ channels are inhibited by neurotransmitters acting through G protein-coupled receptors via a membrane-delimited pathway. Inhibition is reversed by strong depolarization, resulting in prepulse facilitation. Activated G protein βγ subunits (Gβγ) are required for maximal prepulse facilitation. Gβγ binds to multiple sites on Cav2.1, Cav2.2, and Cav2.3 α1 subunits. Here we examine the functional relevance of a C-terminal binding site for Gβγ on Cav2.2b channels, which mediate N-type Ca2+ currents. In vitro binding studies showed that Gβγ subunits bind to the intracellular loop connecting domains I and II and the C-terminal domain of Cav2.2b but not the intracellular loops connecting domains II and III or III and IV. Deletion analysis revealed that the binding site is located near the C terminus, within amino acid residues 2257 to 2336. Directed yeast two-hybrid analysis confirmed this specific binding interaction in vivo in yeast cells. Cav2.2b channels with this site deleted had normal function properties, and they were inhibited essentially normally by strong activation of G proteins with guanosine 5′-3-O-(thio)triphosphate (GTPγS) and were facilitated nearly normally by depolarizing prepulses. Similarly deletion of this site had small, statistically insignificant effects on inhibition of Ca2+ current and on prepulse facilitation in the presence of somatostatin to stimulate receptor-mediated activation of G proteins. In contrast, deletion of the C-terminal Gβγ site substantially reduced the low level of intrinsic prepulse facilitation present at the basal level of G protein activation in tsA-201 cells. Thus, this C-terminal Gβγ binding site contributes to the affinity or efficacy of Gβγ regulation at basal levels of G protein activation. The simplest interpretation of our results is that the C-terminal binding site increases the affinity of Gβγ for the channel but is not required for Gβγ action. C-terminal binding of Gβγ may influence the physiological responsiveness of Ca2+ channels to low-level G protein activation.