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

VS Lelyveld, Anders Björkbom, EM Ransey, P Sliz, JW Szostak

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    10 Citeringar (Scopus)

    Sammanfattning

    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.
    OriginalspråkOdefinierat/okänt
    Sidor (från-till)15378–15381
    Antal sidor4
    TidskriftJournal of the American Chemical Society
    Volym137
    Utgåva49
    DOI
    StatusPublicerad - 2015
    MoE-publikationstypA1 Tidskriftsartikel-refererad

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