Dissecting the influence of Mg2+ on 3D architecture and ligand-binding of the guanine-sensing riboswitch aptamer domain.
Long-range tertiary interactions determine the three-dimensional structure of a number of metabolite-binding riboswitch RNA elements and were found to be important for their regulatory function. For the guanine-sensing riboswitch of the Bacillus subtilis xpt-pbuX operon, our previous NMR-spectroscopic studies indicated pre-formation of long-range tertiary contacts in the ligand-free state of its aptamer domain. Loss of the structural pre-organization in a mutant of this RNA (G37A/C61U) resulted in the requirement of Mg(2+) for ligand binding. Here, we investigate structural and stability aspects of the wild-type aptamer domain (Gsw) and the G37A/C61U-mutant (Gsw(loop)) of the guanine-sensing riboswitch and their Mg(2+)-induced folding characteristics to dissect the role of long-range tertiary interactions, the link between pre-formation of structural elements and ligand-binding properties and the functional stability. Destabilization of the long-range interactions as a result of the introduced mutations for Gsw(loop) or the increase in temperature for both Gsw and Gsw(loop) involves pronounced alterations of the conformational ensemble characteristics of the ligand-free state of the riboswitch. The increased flexibility of the conformational ensemble can, however, be compensated by Mg(2+). We propose that reduction of conformational dynamics in remote regions of the riboswitch aptamer domain is the minimal pre-requisite to pre-organize the core region for specific ligand binding.
Buck
Janina
J
Institute for Organic Chemistry and Chemical Biology, Johann Wolfgang Goethe-University, Max von Laue-Strasse 7 & 9, 60438 Frankfurt am Main, Germany.
Noeske
Jonas
J
Wöhnert
Jens
J
Schwalbe
Harald
H
D052157:Aptamers, Nucleotide / Q000737:chemistry; D001412:Bacillus subtilis / Q000235:genetics; D002413:Cations, Divalent; D006147:Guanine / Q000378:metabolism*; D008024:Ligands; D008274:Magnesium / Q000737:chemistry*; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000737:chemistry* / Q000378:metabolism; D038621:Regulatory Sequences, Ribonucleic Acid; D013696:Temperature
D052157:Aptamers, Nucleotide; D002413:Cations, Divalent; D008024:Ligands; D012329:RNA, Bacterial; D038621:Regulatory Sequences, Ribonucleic Acid; D006147:Guanine; D008274:Magnesium