Regions in the G protein gamma subunit important for interaction with receptors and effectors.
G betagamma dimers containing the gamma11 or gamma1 subunits are often less potent and effective in their ability to regulate effectors compared with dimers containing the gamma2 subunit. To explore the regions of the gamma subunit that affect the activity of the betagamma dimer, we constructed eight chimeric gamma subunits from the gamma1 and gamma2 subunits. Two chimeras were made in which the N-terminal regions of gamma1 and gamma2 were exchanged and two in which the C-terminal regions were transposed. Another set of chimeras was made in which the CAAX motifs of the chimeras were altered to direct modification with different prenyl groups. All eight gamma chimeras were expressed in Sf9 cells with the beta1 subunit, G betagamma dimers were purified, and then they were assayed in vitro for their ability to bind to the G alpha(i1) subunit, to couple G alpha(i1) to the A1 adenosine receptor, to stimulate phospholipase C-beta, and to regulate type I or type II adenyl cyclases. Dimers containing the C-terminal sequence of the gamma2 subunit modified with the geranylgeranyl lipid had the highest affinity for G(i1)alpha (range, 0.5-1.2 nM) and were most effective at coupling the G(i1)alpha subunit to receptor. These dimers were most effective at stimulating the phosphatidylinositol-specific phospholipase C-beta isoform and inhibiting type I adenyl cyclase. In contrast, betagamma dimers containing the N-terminal sequence of the gamma2 subunit and a geranylgeranyl group are most effective at activating type II adenyl cyclase. The results indicate that both the N- and C-terminal regions of the gamma subunit impart specificity to receptor and effector interactions.
Myung
Chang-Seon
CS
Department of Pharmacology, College of Pharmacy, Chungnam National University Daejeon, Korea.
Lim
William K
WK
DeFilippo
Joseph M
JM
Yasuda
Hiroshi
H
Neubig
Richard R
RR
Garrison
James C
JC
D000262:Adenylyl Cyclases / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D019281:Dimerization; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D044388:GTP-Binding Protein gamma Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism; D008969:Molecular Sequence Data; D050506:Mutant Chimeric Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054799:Phospholipase C beta; D043682:Receptor, Adenosine A1 / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism
D044388:GTP-Binding Protein gamma Subunits; D007527:Isoenzymes; D050506:Mutant Chimeric Proteins; D043682:Receptor, Adenosine A1; D010738:Type C Phospholipases; D054799:Phospholipase C beta; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D000262:Adenylyl Cyclases