Diffusion-ordered spectroscopy and saturation transfer difference NMR spectroscopy studies of selective interactions between ELAV protein fragments and an mRNA target

Recently, the crucial role of post-transcriptional processes in the control of gene expression opened a new fascinating route to the translation of basic mechanisms into clinical medicine. In this contest, Embryonic Lethal Abnormal Vision (ELAV) proteins represent a family of well-characterized proteins that bind mRNA molecules and affect their fate and, thus, the amount of corresponding proteins. Interestingly, preliminary data bode well that the ELAV-mediated mRNA stabilization is a mechanism relevant to Alzheimer's disease. Herein, we report the analysis of four ELAV-derived peptides (P1-P4) belonging to the RNA recognition motif of ELAV proteins in the presence of an mRNA fragment from tumor necrosis factor a. Advanced NMR techniques were used to gain insights into the interaction mechanism. Saturation transfer difference and diffusion-ordered NMR spectroscopy experiments showed that appropriate mixtures of peptides (and not each peptide individually) bind effectively to the biological target. The diffusion-ordered spectroscopy (DOSY) technique was applied here for the first time to the study of mRNA-peptides complexes. For each peptide, the 3D structure was also determined by NOE analysis supported by molecular dynamics simulations.