Ligand-induced folding of the guanine-sensing riboswitch is controlled by a combined predetermined induced fit mechanism.
All known guanine-sensing riboswitches regulate gene expression by specifically binding to guanine (G) or related analogs with high affinity to switch off transcription. The aptamers of this class of riboswitches are characterized by three helices (P1-P3), surrounding a central core of phylogenetically conserved nucleotides and a long-range loop-loop interaction. To gain more insight into the switching mechanism, we present here a comparison between the solution-state structures of the G-free and G-bound forms of the guanine aptamer from the xpt-pbuX operon of Bacillus subtilis, as derived from NMR chemical shifts and magnetic-field-induced residual dipolar couplings. The high-resolution NMR analysis shows the G-free aptamer is highly structured with parallel P2 and P3 helices and the long-range loop-loop interaction already present, implying that the structure is largely preformed to bind the ligand. Structural changes upon guanine binding are found to be localized to the central core. In the free state, the G-quadruple interaction and two base pairs of the P1 stem flanking the central core appear to be largely disordered. The ligand thus binds via a combined predetermined-induced fit mechanism, involving a previously unstructured five-residue loop of the J2-3 junction that folds over the ligand. These limited additional interactions within a preorganized setting possibly explain how the aptamer rapidly responds to ligand binding, which is necessary to switch the structural state of the expression platform within a narrow time frame before the RNA polymerase escapes the 5'-UTR.
Ottink
Otmar M
OM
Institute for Molecules and Materials and Department of Biophysical Chemistry, University of Nijmegen, 6525 ED Nijmegen, The Netherlands.
Rampersad
Sumientra M
SM
Tessari
Marco
M
Zaman
Guido J R
GJ
Heus
Hans A
HA
Wijmenga
Sybren S
SS
D000225:Adenine / Q000737:chemistry / Q000378:metabolism; D052157:Aptamers, Nucleotide / Q000737:chemistry; D001412:Bacillus subtilis / Q000187:drug effects / Q000235:genetics; D020029:Base Pairing; D001483:Base Sequence; D002413:Cations, Divalent / Q000494:pharmacology; D006147:Guanine / Q000737:chemistry / Q000378:metabolism; D008024:Ligands; D008274:Magnesium / Q000494:pharmacology; D009682:Magnetic Resonance Spectroscopy; D009690:Nucleic Acid Conformation; D009876:Operon; D012329:RNA, Bacterial / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics; D038621:Regulatory Sequences, Ribonucleic Acid
D052157:Aptamers, Nucleotide; D002413:Cations, Divalent; D008024:Ligands; D012329:RNA, Bacterial; D038621:Regulatory Sequences, Ribonucleic Acid; D006147:Guanine; D008274:Magnesium; D000225:Adenine