The cyclooxygenase-2 is a pro-inflammatory and cancer marker, whose mRNA stability and translation is regulated by the CUG-binding protein 2 interacting with AU-rich sequences in the 3' untranslated region. Here, we present the solution NMR structure of CUG-binding protein 2 RRM3 in complex with 5'-UUUAA-3' originating from the COX-2 3'-UTR. We show that RRM3 uses the same binding surface and protein moieties to interact with AU- and UG-rich RNA motifs, binding with low and high affinity, respectively. Using NMR spectroscopy, isothermal titration calorimetry and molecular dynamics simulations, we demonstrate that distinct sub-states characterized by different aromatic side-chain conformations at the RNA-binding surface allow for high- or low-affinity binding with functional implications. This study highlights a mechanism for RNA discrimination possibly common to multiple RRMs as several prominent members display a similar rearrangement of aromatic residues upon binding their targets.The RNA Recognition Motif (RRM) is the most ubiquitous RNA binding domain. Here the authors combined NMR and molecular dynamics simulations and show that the RRM RNA binding surface exists in different states and that a conformational switch of aromatic side-chains fine-tunes sequence specific binding affinities.

Department of Biology ETH Zürich Institute of Molecular Biology and Biophysics HPP L 14 1 Hönggerbergring 64 8093 Zürich Switzerland

Department of Integrative Structural and Computational Biology The Scripps Research Institute MB 33 10550 North Torrey Pines Road La Jolla California 92037 USA

Department of Physical Chemistry Faculty of Science Regional Centre of Advanced Technologies and Materials Palacky University Olomouc 17 listopadu 12 771 46 Olomouc Czech Republic

Department of Structural Biology Stanford University Stanford CA 94305 USA

Institute of Biophysics Academy of Sciences of the Czech Republic Kralovopolska 135 612 65 Brno Czech Republic

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