Identification of Spermidine Binding Site in T-box Riboswitch Antiterminator RNA

• Published in: Chemical biology & drug design Vol. 87; no. 2; pp. 182 - 189
• Main Authors: Liu, Jia, Zeng, Chunxi, Hogan, Vivian, Zhou, Shu, Monwar, Md Masud, Hines, Jennifer V.
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01-02-2016
• Subjects: antiterminator; Base Pairing; binding; Binding Sites; Codon, Terminator; Molecular Docking Simulation; Nuclear Magnetic Resonance, Biomolecular; Nucleic Acid Conformation; riboswitch; Riboswitch - physiology; RNA; RNA Stability; RNA, Transfer - chemistry; RNA, Transfer - metabolism; spermidine; Spermidine - chemistry; Spermidine - metabolism; T-box; Thermodynamics; riboswitch; spermidine; binding; RNA; antiterminator; T-box
• ISSN: 1747-0277; 1747-0285
• DOI: 10.1111/cbdd.12660

The T‐box transcription antitermination riboswitch controls bacterial gene expression by structurally responding to uncharged, cognate tRNA. Previous studies indicated that cofactors, such as the polyamine spermidine, might serve a specific functional role in enhancing riboswitch efficacy. As riboswitch function depends on key RNA structural changes involving the antiterminator element, the interaction of spermidine with the T‐box riboswitch antiterminator element was investigated. Spermidine binds antiterminator model RNA with high affinity (micromolar Kd) based on isothermal titration calorimetry and fluorescence‐monitored binding assays. NMR titration studies, molecular modeling, and inline and enzymatic probing studies indicate that spermidine binds at the 3′ portion of the highly conserved seven‐nucleotide bulge in the antiterminator. Together, these results support the conclusion that spermidine binds the T‐box antiterminator RNA preferentially in a location important for antiterminator function. The implications of these findings are significant both for better understanding of the T‐box riboswitch mechanism and for antiterminator‐targeted drug discovery efforts.
The T‐box transcription antitermination riboswitch controls bacterial gene expression by structurally responding to uncharged, cognate tRNA, in part by base pairing with an antiterminator element within the riboswitch. Spermidine binds the T‐box antiterminator RNA in a location important for antiterminator function. The implications of these findings are significant both for better understanding the T‐box riboswitch mechanism and for antiterminator‐targeted drug discovery efforts.