Pinpointing RNA-Protein Cross-Links with Site-Specific Stable Isotope-Labeled Oligonucleotides

A1 Journal article (refereed)


Internal Authors/Editors


Publication Details

List of Authors: Lelyveld VS, Bjorkbom A, Ransey EM, Sliz P, Szostak JW
Publisher: AMER CHEMICAL SOC
Publication year: 2015
Journal: Journal of the American Chemical Society
Journal acronym: J AM CHEM SOC
Volume number: 137
Issue number: 49
Start page: 15378
End page: 15381
Number of pages: 4
ISSN: 0002-7863
eISSN: 1520-5126


Abstract

High affinity RNA-protein interactions are critical to cellular function, but directly identifying the determinants of binding within these complexes is often difficult. Here, we introduce a stable isotope mass labeling technique to assign specific interacting nucleotides in an oligonudeotide-protein complex by photo-cross-linking. The method relies on generating site-specific oxygen-18-labeled phosphodiester linkages in oligonucleotides, such that covalent peptide-oligonucleotide cross-link sites arising from ultraviolet irradiation can be assigned to specific sequence positions in both RNA and protein simultaneously by mass spectrometry. Using Lin28A and a let-7 pre-element RNA, we demonstrate that mass labeling permits unambiguous identification of the cross-linked sequence positions in the RNA-protein complex.

Last updated on 2019-15-10 at 02:51