A highly conserved glycine within linker I and the extreme C terminus of G protein alpha subunits interact cooperatively in switching G protein-coupled receptor-to-effector specificity

TY - JOUR

T1 - A highly conserved glycine within linker I and the extreme C terminus of G protein alpha subunits interact cooperatively in switching G protein-coupled receptor-to-effector specificity

AU - Kostenis, Evi

AU - Martini, Lene

AU - Ellis, James

AU - Waldhoer, Maria

AU - Heydorn, Arne

AU - Rosenkilde, Mette M

AU - Norregaard, Pia K

AU - Jorgensen, Rasmus

AU - Whistler, Jennifer L

AU - Milligan, Graeme

N1 - Keywords: Amino Acid Sequence; Animals; Blotting, Western; COS Cells; Calcium; Cell Membrane; Cercopithecus aethiops; Conserved Sequence; DNA; Enzyme-Linked Immunosorbent Assay; GTP-Binding Protein alpha Subunits; Glycine; Guanosine 5'-O-(3-Thiotriphosphate); Inositol Phosphates; Ligands; Molecular Sequence Data; Receptors, G-Protein-Coupled; Signal Transduction; Transfection

PY - 2004

Y1 - 2004

N2 - Numerous studies have attested to the importance of the extreme C terminus of G protein alpha subunits in determining their selectivity of receptor recognition. We have previously reported that a highly conserved glycine residue within linker I is important for constraining the fidelity of receptor recognition by Galpha(q) proteins. Herein, we explored whether both modules (linker I and extreme C terminus) interact cooperatively in switching G protein-coupled receptor (GPCR)-to-effector specificity and created as models mutant Galpha(q) proteins in which glycine was replaced with various amino acids and the C-terminal five Galpha(q) residues with the corresponding Galpha(i) or Galpha(s) sequence. Coupling properties of the mutated Galpha(q) proteins were determined after coexpression with a panel of 13 G(i)-and G(s) -selective receptors and compared with those of Galpha proteins modified in only one module. Galpha proteins modified in both modules are significantly more efficacious in channeling non-G(q) -selective receptors to G(q)-mediated signaling events compare with those containing each module alone. Additive effects of both modules were observed even if individual modules lacked an effect on GPCR-to-effector specificity. Dually modified Galpha proteins were also superior in conferring high-affinity agonist sites onto a coexpressed GPCR in the absence, but not in the presence, of guanine nucleotides. Together, our data suggest that receptor-G protein coupling selectivity involves cooperative interactions between the extreme C terminus and linker I of Galpha proteins and that distinct determinants of selectivity exist for individual receptors.

AB - Numerous studies have attested to the importance of the extreme C terminus of G protein alpha subunits in determining their selectivity of receptor recognition. We have previously reported that a highly conserved glycine residue within linker I is important for constraining the fidelity of receptor recognition by Galpha(q) proteins. Herein, we explored whether both modules (linker I and extreme C terminus) interact cooperatively in switching G protein-coupled receptor (GPCR)-to-effector specificity and created as models mutant Galpha(q) proteins in which glycine was replaced with various amino acids and the C-terminal five Galpha(q) residues with the corresponding Galpha(i) or Galpha(s) sequence. Coupling properties of the mutated Galpha(q) proteins were determined after coexpression with a panel of 13 G(i)-and G(s) -selective receptors and compared with those of Galpha proteins modified in only one module. Galpha proteins modified in both modules are significantly more efficacious in channeling non-G(q) -selective receptors to G(q)-mediated signaling events compare with those containing each module alone. Additive effects of both modules were observed even if individual modules lacked an effect on GPCR-to-effector specificity. Dually modified Galpha proteins were also superior in conferring high-affinity agonist sites onto a coexpressed GPCR in the absence, but not in the presence, of guanine nucleotides. Together, our data suggest that receptor-G protein coupling selectivity involves cooperative interactions between the extreme C terminus and linker I of Galpha proteins and that distinct determinants of selectivity exist for individual receptors.

U2 - 10.1124/jpet.104.080424

DO - 10.1124/jpet.104.080424

M3 - Journal article

C2 - 15615862

SN - 0022-3565

VL - 313

SP - 78

EP - 87

JO - Journal of Pharmacology and Experimental Therapeutics

JF - Journal of Pharmacology and Experimental Therapeutics

IS - 1

ER -