Receptor-mediated changes at the myristoylated amino terminus of Galpha(il) proteins.
G protein-coupled receptors (GPCRs) catalyze nucleotide release in heterotrimeric G proteins, the slow step in G protein activation. G i/o family proteins are permanently, cotranslationally myristoylated at the extreme amino terminus. While myristoylation of the amino terminus has long been known to aid in anchoring G i proteins to the membrane, the role of myristoylation with regard to interaction with activated receptors is not known. Previous studies have characterized activation-dependent changes in the amino terminus of Galpha proteins in solution [Medkova, M. (2002) Biochemistry 41, 9963-9972; Preininger, A. M. (2003) Biochemistry 42, 7931-7941], but changes in the environment of specific residues within the Galpha i1 amino terminus during receptor-mediated G i activation have not been reported. Using site-specific fluorescence labeling of individual residues along a stretch of the Galpha il amino terminus, we found specific changes in the environment of these residues upon interaction with the activated receptor and following GTPgammaS binding. These changes map to a distinct surface of the amino-terminal helix opposite the Gbetagamma binding interface. The receptor-dependent fluorescence changes are consistent with a myristoylated amino terminus in the proximity of the membrane and/or receptor. Myristoylation affects both the rate and intensity of receptor activation-dependent changes detected at several residues along the amino terminus (with no significant effect on the rate of receptor-mediated GTPgammaS binding). This work demonstrates that the myristoylated amino terminus of Galpha il proteins undergoes receptor-mediated changes during the dynamic process of G protein signaling.
Preininger
Anita M
AM
Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232-6600, USA.
Parello
Joseph
J
Meier
Scott M
SM
Liao
Guihua
G
Hamm
Heidi E
HE
D003902:Detergents; D044385:GTP-Binding Protein alpha Subunits / Q000737:chemistry* / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D008958:Models, Molecular; D019814:Myristic Acid / Q000378:metabolism; D011487:Protein Conformation; D017382:Reactive Oxygen Species / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000378:metabolism; D012995:Solubility; D013499:Surface Properties; D014364:Tryptophan / Q000032:analysis
D003902:Detergents; D044385:GTP-Binding Protein alpha Subunits; D017382:Reactive Oxygen Species; D043562:Receptors, G-Protein-Coupled; D019814:Myristic Acid; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D014364:Tryptophan
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