Using phosphorothioate-substituted RNA to investigate the thermodynamic role of phosphates in a sequence specific RNA-protein complex

Dagmar Dertinger, Linda S. Behlen, Olke C. Uhlenbeck*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

35 Scopus citations

Abstract

Part of the binding affinity and specificity in RNA-protein complexes is often contributed by contacts between the protein and backbone phosphates that are held in position by the RNA structure. This study focuses on the well-characterized interaction between a dimer of the MS2 coat protein and a small RNA hairpin. Using a short oligoribonucleotide which contains all the necessary sequence elements required for tight protein binding, a single phosphorothioate linkage was introduced at 13 different positions. In each case, the R(p) and S(p) stereoisomers were separated and their affinities to the MS2 coat protein were determined. Comparison of these biochemical data with the crystal structure of the protein-hairpin complex indicates that introduction of a phosphorothioate only affects binding at sites where a protein-phosphate contact is observed in the crystal structure. This means that phosphorothioate-containing oligoribonucleotides should also be useful for mapping phosphate contacts in RNA-protein complexes for which no crystal structure is available.

Original languageEnglish (US)
Pages (from-to)55-63
Number of pages9
JournalBiochemistry
Volume39
Issue number1
DOIs
StatePublished - Jan 11 2000

ASJC Scopus subject areas

  • Biochemistry

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