The overall objective of the proposed research is to determine the structural and functional significance of the heterogeneity in the beta and gamma subunits of the GTP-binding (G) proteins involved in transmembrane signaling. The first objective will be to isolate complementary DNA (cDNA) clones for bovine brain gamma subunit mRNAs by oligodeoxyribonucleotide hybridization using probes derived from amino acid sequence unique to each of the subunits. These cDNA clones, or portions thereof, will be used as hybridization probes to identify related gamma unit cDNAs. The second objective will be to express cDNA clones for each of the gamma subunits in combination with cDNA clones for each of the beta subunits in E. coli. The individual beta gamma subunit complexes will then be purified from E. coli and reconstituted into phospholipid vesicles to compare the various functions of these subunits. Functions to be studied include: 1) the relative affinities of the beta gamma subunit complexes for the alpha subunits of the G proteins, 2) the relative abilities of the beta gamma subunit complexes to promote interaction of the alpha subunits of the G proteins with their receptors, and 3) the relative effectiveness of the beta gamma subunit complexes to inhibit hormone-stimulated adenylate cyclase activity. The final objective will be to determine if there is any specific association of particular beta and gamma subunits with the gamma subunits of the G proteins in vivo by immunoprecipitation of different alpha beta gamma complexes with gamma subunit-specific antibodies. This knowledge along with cDNA clones and antibodies for the alpha, beta, and gamma subunits of the G proteins will allow us to pursue our long-term objective to study the regulation of expression, cellular processing, and assembly of these oligomeric proteins.
| Schwindinger, William F; Mirshahi, Uyenlinh L; Baylor, Kelly A et al. (2012) Synergistic roles for G-protein ?3 and ?7 subtypes in seizure susceptibility as revealed in double knock-out mice. J Biol Chem 287:7121-33 |
| Schwindinger, William F; Mihalcik, Lauren J Murphree; Giger, Kathryn E et al. (2010) Adenosine A2A receptor signaling and golf assembly show a specific requirement for the gamma7 subtype in the striatum. J Biol Chem 285:29787-96 |
| Schwindinger, William F; Borrell, Brandon M; Waldman, Lora C et al. (2009) Mice lacking the G protein gamma3-subunit show resistance to opioids and diet induced obesity. Am J Physiol Regul Integr Comp Physiol 297:R1494-502 |
