Translational T-box riboswitches bind tRNA by modulating conformational flexibility.

Area of Science:

• Molecular Biology
• RNA Biology
• Bacterial Genetics

Background:

• T-box riboswitches are crucial noncoding RNA regulators in Gram-positive bacteria.
• They control amino acid metabolism by sensing tRNA aminoacylation status.
• This regulation impacts transcription or translation of related genes.

Purpose of the Study:

• To investigate the binding mechanism of the Mycobacterium tuberculosis IleS T-box riboswitch using single-molecule Förster Resonance Energy Transfer (smFRET).
• To elucidate the kinetic and conformational dynamics of translational T-box riboswitches.

Main Methods:

• Single-molecule Förster Resonance Energy Transfer (smFRET) studies.
• Analysis of tRNA binding and conformational changes in the T-box riboswitch.

Main Results:

• A two-step binding model was supported, with initial tRNA anticodon recognition followed by NCCA sequence interaction.
• Transient docking of tRNA into the discriminator domain occurs post-anticodon recognition, even without NCCA interactions.
• NC_CA-discriminator interactions significantly stabilize the fully bound state, suggesting a conformational selection model for NCCA recognition.

Conclusions:

• Translational T-box riboswitches exhibit significant conformational flexibility.
• The findings provide a kinetic framework for understanding RNA-protein binding specificity and affinity in gene regulation.
• This work deepens the understanding of genetic regulatory mechanisms in bacteria.