The N-terminal coiled-coil domain of beta is essential for gamma association: a model for G-protein beta gamma subunit interaction.
We have identified the N terminus of the beta subunit as an essential domain for G-protein beta gamma assembly. A C-terminal fragment, beta 1-(130-340), fails to bind gamma unless coexpressed with the complementary N-terminal fragment, beta 1-(1-129). Deletion of the N-terminal 33 residues of beta 1, a region identified by computer algorithm to favor coiled-coil formation, abolishes gamma 2 association. On the basis of these findings, we propose a coiled-coil model of beta gamma interaction and refine this by computer-assisted molecular modeling. The model is tested by further mutagenesis: reversing the charge of residues in beta 1 that are hypothesized to be involved in interhelical salt bridges precludes gamma association. Insertions in the coiled-coil region, which disrupt the proposed hydrophobic interface, prevent gamma association. This structural basis for beta gamma dimerization provides a starting point for the design of beta and gamma mutants that can be used to map regions in beta gamma critical for interactions with the alpha subunit, receptors, and effectors.
Garritsen
A
A
Molecular Pathophysiology Branch, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
van Galen
P J
PJ
Simonds
W F
WF
D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D002460:Cell Line; D004268:DNA-Binding Proteins; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D046911:Macromolecular Substances; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000302:isolation & purification; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism; D017433:Protein Structure, Secondary; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D014162:Transfection
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D046911:Macromolecular Substances; D010446:Peptide Fragments; D029701:Saccharomyces cerevisiae Proteins; D001120:Arginine; D011494:Protein Kinases; D019204:GTP-Binding Proteins