RNA Aptamer-Based Approach to Inhibiting Split-GFP Reconstruction and the Loss of Inhibitory Activity Using Complementary RNA

Abstract

Abstract Although many studies on protein–protein interactions (PPIs) have been conducted and the importance of PPIs in biological processes has been reported, there is still no versatile research approach that enables us to draw a complete picture of PPIs. One orthodox approach to elucidating the mechanism of each PPI would be to inhibit or enhance the PPI of interest and carefully observe its phenotype. However, since the interaction surfaces of PPIs are generally shallow and wide, it is very difficult to design small molecules that can selectively perturb specific PPIs by interaction with these surfaces. In this report, we adopt reconstruction of split green fluorescence protein (splitGFP) as a model of PPI, and obtained RNA aptamers that bind to one of the components. The reconstitution of splitGFP was inhibited by these aptamers, and this inhibition was cancelled by the addition of their complementary sequences. These processes were monitored by the loss and recovery, respectively, of fluorescence from the reconstructed GFP. The successful development of molecules that reversibly regulate specific PPI is expected to make a significant contribution to life science research.