QUANTITATIVE-ANALYSIS OF A CYSTEINE (351)GLYCINE MUTATION IN THE G-PROTEIN G(I1)ALPHA - EFFECT ON ALPHA(2A)-ADRENOCEPTOR-G(I1)ALPHA FUSION PROTEIN-ACTIVATION
Ic. Carr et al., QUANTITATIVE-ANALYSIS OF A CYSTEINE (351)GLYCINE MUTATION IN THE G-PROTEIN G(I1)ALPHA - EFFECT ON ALPHA(2A)-ADRENOCEPTOR-G(I1)ALPHA FUSION PROTEIN-ACTIVATION, FEBS letters, 428(1-2), 1998, pp. 17-22
Fusion proteins mere constructed between the porcine alpha(2A)-adrenoc
eptor and either wild-type (Cys(351)) Or a pertussis toxin-resistant (
Gly(351)) form of the G protein G(i1)alpha. Addition of adrenaline to
membranes expressing the fusion proteins resulted in concentration-dep
endent stimulation of their high affinity GTPase activity. The alpha(2
A)-adrenoceptor-wild type G(i1)alpha fusion protein produced substanti
ally higher maximal stimulation of GTPase activity in response to adre
naline than that containing Gly(351) G(i1)alpha. Treatment of the fusi
on proteins as agonist-regulated enzymes allowed measurement of V-max
and turnover number for adrenaline-stimulation of the GTPase activity
of each fusion construct. The turnover number of the alpha(2A)-adrenoc
eptor-Cys(351) Gly G(i1)alpha fusion protein was only 44% of that for
the alpha(2A)-adrenoceptor-wild type G(i1)alpha fusion protein. These
data provide the first direct quantitative evaluation of the effects o
f a mutation of a G protein on the capacity of an agonist-occupied rec
eptor to activate the mutant. (C) 1998 Federation of European Biochemi
cal Societies.