Site-specific Profiling of RNA-binding Proteins Enabled by Isotopic Signature Enhanced Mass Spectrometry

We developed an innovative method named “isoRIC” for profiling RNA-binding proteomes with site-specific resolution, which combined the metabolic labeling of isotopic nucleotides for photo-crosslinking of RNA-binding proteins and the real-time targeted LC-MS/MS analysis of RNA-crosslinked peptides. This method showed dramatic improvement of sensitivity in identifying RNA-crosslinked peptides with low abundance as compared to the DDA-based proteomic approaches, which enables the discovery of novel RNA-binding proteins and precise mapping of RNA-protein binding interfaces at single amino acid resolution.