(a) Signal intensities at varying concentrations of Gαi1 as
activated by aluminum tetrafluoride addition
(sequential application of NaF and AlCl3).
(b) Z’-factors of the assay at varying Gαi1 concentrations.
A non-radioactive method for measuring Gα proteins with a Switch II region is detailed in BMG LABTECH Application Note 196. Heterotrimeric G Proteins are a vast family of proteins made of 3 subunits, alpha, beta and gamma. The Gα subunit, upon activation, undergoes a conformational change whereby it releases GDP and subsequently binds GTP. In the (Gαi/o) subfamily of proteins, there is a switch region that moves upon activation and conformational change. In this Switch II region an intrinsic tryptophan can be monitored, and upon activation its fluorescence intensity changes. This a fairly universal assay that can measure Gαi/o activation as modulated by a variety of inputs (eg. peptides, proteins, small molecules). The assay is a sensitive and high-quality means to measure G-protein activation without the use of radiolabeled nucleotides.
You can find the entire Application Note 196: Using Intrinsic Tryptophan Fluorescence to Measure Heterotrimeric G-Protein Activation here: http://www.bmglabtech.com/application-notes/fluorescence-intensity/intrinsic-tryptophan-fluorescence-196.cfm