Protein-protein-interactions in a multiplexed, miniaturized format a functional analysis of Rho GTPase activation and inhibition.

A miniaturized, bead-based protein-protein-interaction assay was developed to study the interaction of Rho GTPases with regulatory proteins. The setup, which uses only minute amounts of sample, was used to analyze small molecules that inhibit the interaction between Rho GTPases and RhoGDI alpha. Prenylcysteine analogues and the replacement of GDP by non-hydrolysable GTP analogues prevented the formation of Rho GTPase-RhoGDI alpha complexes in a concentration-dependent manner.