Receptor-coupling properties of the invertebrate visual guanine nucleotide binding protein iGqα

Abstract

Invertebrate visual iGqα is homologous to mammalian mGqα in two of the three domains important for G protein interaction with receptors; the C-terminus and the linker regions that connect the helical and ras-like domains of the α subunit. The third receptor-interacting domain, the N-terminus, contains a six amino acid extension MTLESI in mGqα that is not present in iGqα. In co-expression studies we assessed the promiscuity and efficacy of receptor coupling to phospholipase C (PLC) by iGqα, a non-palmitoylated mutant iGqα(C3,4A), mGqα and G15α. The invertebrate G proteins and mGqα only coupled to Gq-coupled receptors (m1 muscarinic acetylcholine receptor (mChR1), α1A-adrenergic receptor (α1-AR)) and not to the Gi/s-coupled receptors (CCR1 receptor, β2-adrenergic receptor or dopamine D1 receptor) while G15α coupled to all receptors. iGqα and iGqα(C3,4A) both had double the efficacy for PLC activation compared to the mammalian G proteins when co-expressed with mChR1 and α1-AR. The increased efficacy of iGqα compared to mGqα was also seen downstream of PLC with carbachol stimulation of the mitogen-activated protein kinase, ERK1/2. Addition of the MTLESI extension onto the N-terminus of iGqα decreased its efficacy by 35% whereas deletion of this sequence from mGqα increased its efficacy by 60% in the PLC and ERK1/2 assays. iGqα, iGqα(C3,4A) and mGqα all displayed similar receptor-independent AlF4–activation of PLC and guanosine triphosphate hydrolysis (GTPase) activity. iGqα, and iGqα(C3,4A) both had increased receptor-activated guanosine 5′-[γ-[35S]thio]triphosphate ([35S]GTPγS) binding when compared to mGqα when co-expressed with the mChR1. These results demonstrate that Gq protein efficacy is at least partially determined by the presence of the amino-terminal MTLESI extension. Comparison of [35S]GTPγS binding rates helps explain the increased efficacy of the invertebrate G proteins.