In Vitro Analysis of G-Protein Functions

This chapter focuses on the methods used for biochemical studies that are unique to G proteins. It describes several methods commonly used to detect the presence of G proteins in cell extracts or cell fractions, and discusses techniques to determine whether a G protein is involved in a particular cellular process. Examples of use of these procedures in plants are presented. Guanosine triphosphate (GTP) analogs and the cholera and pertussis bacterial toxins are extremely useful tools to detect G proteins in different cellular fractions and to study the involvement of G proteins in various cellular functions. GTP analogs, particularly GTPγS, are used frequently because of the relative ease with which they can detect G proteins and probe G protein functions. However GTP analogs are not specific, therefore, conclusions from studies using GTP analogs are not definitive. The bacterial toxins, particularly pertussis toxin, are more specific, and they are used for analyses of heterotrimeric G proteins. Although there are limitations, these biochemical methods can provide valuable information, which complements information gained from molecular and genetic studies.