Abstract
We recently identified glycoRNA—a previously undescribed glycoconjugate—which consists of RNAs modified with secretory N-glycans and presented on the cell surface. While previous work supported a covalent linkage between RNA and glycans, the direct chemical nature of the RNA-glycan connection was not described. Here we develop a sensitive and scalable protocol to detect and characterize native glycoRNAs. Leveraging periodate oxidation and aldehyde ligation (rPAL) and Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH-MS), we identified the modified RNA base 3-(3-amino-3-carboxypropyl)uridine (acp3U) as a site of attachment of N-glycans in glycoRNA. The sensitivity and robustness of rPAL provided the first evidence of a direct glycan-RNA linkage, and its flexibility will enable further characterization of glycoRNA biology.
Competing Interest Statement
R.A.F and C.R.B. are cofounders and stockholders of GanNA Bio. R.A.F is a board of directors member and stockholder of Chronus Health. C.R.B. is a cofounder and Scientific Advisory Board member of Lycia Therapeutics, Palleon Pharmaceuticals, Enable Bioscience, Redwood Biosciences (a subsidiary of Catalent), and InterVenn Biosciences. The other authors declare no competing interests.
Footnotes
We found one additional chemical structure in Fig 2 that was miss annotated and have now corrected it.