TY - JOUR
T1 - G protein βγ subunits. Simplified purification and properties of novel isoforms
AU - Ueda, Natsuo
AU - Iñiguez-Lluhi, Jorge A.
AU - Lee, Ethan
AU - Smrcka, Alan V.
AU - Robishaw, Janet D.
AU - Gilman, Alfred G.
N1 - Copyright:
Copyright 2004 Elsevier B.V., All rights reserved.
PY - 1994
Y1 - 1994
N2 - The β and γ subunits of heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins) form tightly associated complexes. To examine functional differences among the large number of possible combinations of unique β and γ subunits, we have synthesized and characterized βγ complexes containing γ5 and γ7, two widely distributed γ subunits. When either γ5 or γ7 is expressed concurrently with β1 or β2 subunits in a baculovirus/Sf9 cell system, all four subunit complexes support pertussis toxin-catalyzed ADP-ribosylation of rG(iα1) (where 'r' indicates recombinant), indicating formation of functional complexes. Each of the complexes was purified by subunit exchange chromatography, using the G203A mutant of rG(iα1) as the immobilized ligand. The purified preparations were compared with other recombinant βγ subunits, including β1γ1 and β1γ2, for their ability to modulate type I and II adenylyl cyclase activities; stimulate phosphoinositide-specific phospholipase Cβ; support pertussis toxin-catalyzed ADP-ribosylation of rG(iα1) and G(oα); and inhibit steady-state GTP hydrolysis catalyzed by G(sα), G(oα), and myristoylated rG(iα2). The results emphasize the unique properties of β1γ1. The properties of the complexes containing γ5 or γ7 were similar to each other and to those of β1γ2.
AB - The β and γ subunits of heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins) form tightly associated complexes. To examine functional differences among the large number of possible combinations of unique β and γ subunits, we have synthesized and characterized βγ complexes containing γ5 and γ7, two widely distributed γ subunits. When either γ5 or γ7 is expressed concurrently with β1 or β2 subunits in a baculovirus/Sf9 cell system, all four subunit complexes support pertussis toxin-catalyzed ADP-ribosylation of rG(iα1) (where 'r' indicates recombinant), indicating formation of functional complexes. Each of the complexes was purified by subunit exchange chromatography, using the G203A mutant of rG(iα1) as the immobilized ligand. The purified preparations were compared with other recombinant βγ subunits, including β1γ1 and β1γ2, for their ability to modulate type I and II adenylyl cyclase activities; stimulate phosphoinositide-specific phospholipase Cβ; support pertussis toxin-catalyzed ADP-ribosylation of rG(iα1) and G(oα); and inhibit steady-state GTP hydrolysis catalyzed by G(sα), G(oα), and myristoylated rG(iα2). The results emphasize the unique properties of β1γ1. The properties of the complexes containing γ5 or γ7 were similar to each other and to those of β1γ2.
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M3 - Article
C2 - 8308009
AN - SCOPUS:0028169730
VL - 269
SP - 4388
EP - 4395
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 6
ER -