A Specific, UV-Induced RNA-Protein Cross-Link Using 5-Bromouridine-Substituted RNA

Jonatha M. Gott, Michael C. Willis, Tad H. Koch, Olke C. Uhlenbeck*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

53 Scopus citations

Abstract

The well-characterized RNA binding site of the bacteriophage R17 coat protein has been used to investigate the cross-linking of protein to 5-bromouridine (BrU)-substituted RNA using medium-wavelength UV light. We have demonstrated a specific RNA-protein cross-link and identified the site on the RNA of protein attachment. Formation of the covalent complex is dependent upon the presence of BrU at position -5 of the RNA and specific binding of the RNA by coat protein. The amount of cross-linking increases with time and depends on the light source and conditions used. Irradiations using a broad-spectrum UV transilluminator (peak at 312 nm) or monochromatic XeCl excimer laser (308 nm) gave levels of cross-linking exceeding 20 and 50%, respectively. The quantum yield of photo-cross-linking, determined with 308-nm excitation, was 0.003. While little strand breakage or debromination of the RNA occurred, significant protein photodamage was observed.

Original languageEnglish (US)
Pages (from-to)6290-6295
Number of pages6
JournalBiochemistry
Volume30
Issue number25
DOIs
StatePublished - Jun 1 1991

ASJC Scopus subject areas

  • Biochemistry

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