Structural determinants for ligand capture by a class II preQ₁ riboswitch

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 111; no. 6; pp. E663 - E671
• Main Authors: Kang, Mijeong, Eichhorn, Catherine D, Feigon, Juli
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 11-02-2014; National Acad Sciences
• Series: PNAS Plus
• Subjects: Base Sequence; Binding Sites; Biological Sciences; Calcium - metabolism; Lactobacillus rhamnosus - genetics; Ligands; Models, Molecular; Molecular Sequence Data; Nuclear Magnetic Resonance, Biomolecular; Nucleic Acid Conformation; PNAS Plus; Riboswitch; Streptococcus pneumoniae - genetics
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1400126111

Prequeuosine (preQ ₁) riboswitches are RNA regulatory elements located in the 5′ UTR of genes involved in the biosynthesis and transport of preQ ₁, a precursor of the modified base queuosine universally found in four tRNAs. The preQ ₁ class II (preQ ₁-II) riboswitch regulates preQ ₁ biosynthesis at the translational level. We present the solution NMR structure and conformational dynamics of the 59 nucleotide Streptococcus pneumoniae preQ ₁-II riboswitch bound to preQ ₁. Unlike in the preQ ₁ class I (preQ ₁-I) riboswitch, divalent cations are required for high-affinity binding. The solution structure is an unusual H-type pseudoknot featuring a P4 hairpin embedded in loop 3, which forms a three-way junction with the other two stems. ¹³C relaxation and residual dipolar coupling experiments revealed interhelical flexibility of P4. We found that the P4 helix and flanking adenine residues play crucial and unexpected roles in controlling pseudoknot formation and, in turn, sequestering the Shine–Dalgarno sequence. Aided by divalent cations, P4 is poised to act as a “screw cap” on preQ ₁ recognition to block ligand exit and stabilize the binding pocket. Comparison of preQ ₁-I and preQ ₁-II riboswitch structures reveals that whereas both form H-type pseudoknots and recognize preQ ₁ using one A, C, or U nucleotide from each of three loops, these nucleotides interact with preQ ₁ differently, with preQ ₁ inserting into different grooves. Our studies show that the preQ ₁-II riboswitch uses an unusual mechanism to harness exquisite control over queuosine metabolism.