Cold-shock domain containing protein C2 (CSD-C2; also known as PIPPin) is an RNA-binding protein conserved in the evolution that interacts with the3’-untranslated region (3’-UTR) of rat H1.0 and H3.3 histone messengers. Biolayer interferometry (BLI) is a technique that measures changes in an interferencepattern generated from visible light, reflected from an optical layer, and a biolayer which contains molecules of interest. In this study, we used the BLI methodology inorder to analyze and describe the binding properties of CSD-C2 and the mRNA encoding the rat brain histone protein H3.3. Recombinant CSD-C2 was incubatedwith in vitro transcribed, and biotinylated H3.3 RNA fragments bound to streptavidin-conjugated Octet optical biosensors. In order to define the RNA regioninvolved in binding, we used RNA probes corresponding to different portions of H3.3 RNA 3’-UTR.In this study, we showed that CSD-C2 binds to the last 199 nucleotides of the H3.3 RNA 3’-UTR, and that the apparent affinity constant of the interaction is in thenanomolar range. In addition, this study confirmed that BLI can be a very efficient and reliable method for studying RNA-protein interactions.
|Number of pages||4|
|Journal||BRAIN AND NERVES|
|Publication status||Published - 2017|