Mg2+-Responsive riboswitches represent a fascinating example of bifunctional RNAs that sense Mg2+ ions with high selectivity and autonomously regulate the expression of Mg2+-transporter proteins. The mechanism of the mgtA riboswitch is scarcely understood, and a detailed structural analysis is called for to study how this RNA can selectively recognize Mg2+ and respond by switching between two alternative stem loop structures. In this work, we investigated the structure and Mg2+-binding properties of the lower part of the antiterminator loop C from the mgtA riboswitch of Yersinia enterocolitica by solution NMR and report a discrete Mg2+-binding site embedded in the AU-rich sequence. At the position of Mg2+ binding, the helical axis exhibits a distinct kink accompanied by a widening of the major groove, which accommodates the Mg2+-binding pocket. An unusually large overlap between two adenine residues on the opposite strands suggests that the bending may be sequence-induced by strong stacking interactions, enabling Mg2+ to bind at this so-far not described metal-ion binding site.
Metal ions play a crucial role in charge compensation, folding and stabilization of tertiary structures of large nucleic acids. In addition, they may be directly involved in the catalytic mechanism of ribozymes. Most metal ions applied in the context of nucleic acids in vivo and in vitro bind in a kinetically labile fashion. Hence, the detection of metal ion binding sites, not to mention the elucidation of the specific coordination sphere, still poses largely unresolved problems. Here we describe the different strategies applied and the progress made over the last years to characterize metal ion coordination to large nucleic acids by NMR. Metal ions play a crucial role in charge compensation, folding and stabilization of tertiary structures of large nucleic acids. In addition, they may be directly involved in the catalytic mechanism of ribozymes. Most metal ions applied in the context of nucleic acids in vivo and in vitro bind in a kinetically labile fashion. Hence, the detection of metal ion binding sites, not to mention the elucidation of the specific coordination sphere, still poses largely unresolved problems. Here we describe the different strategies applied and the progress made over the last years to characterize metal ion coordination to large nucleic acids by NMR.
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