Binding by TRBP-dsRBD2 Does Not Induce Bending of Double-Stranded RNA

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Protein-nucleic acid interactions are crucial for biological processes, often involving nucleic acid helix bending.
• The protein TRBP (trans-acting regulatory protein) interacts with double-stranded RNA (dsRNA) via its double-stranded RNA-binding domains (dsRBDs).
• Previous models suggested dsRNA bending is necessary for TRBP binding to its A-form geometry.

Purpose of the Study:

• To investigate the conformational changes of dsRNA upon binding to TRBP's dsRBDs.
• To determine the binding affinity and thermodynamic properties of the dsRBD2-dsRNA interaction.
• To elucidate the mechanism by which TRBP interacts with dsRNA structure.

Main Methods:

• Isothermal titration calorimetry (ITC) to measure binding affinity and thermodynamics.
• Molecular-dynamics (MD) simulations to assess RNA bending during interaction.
• Analysis of existing crystal structures of dsRBD-dsRNA complexes.

Main Results:

• TRBP dsRBD2 binds dsRNA with a temperature-independent affinity of approximately 500 nM.
• A near-zero heat capacity change (ΔCp) indicates minimal conformational changes upon binding.
• MD simulations showed only modest dsRNA bending, contradicting previous hypotheses.
• TRBP preferentially binds to ideal A-form dsRNA structures, avoiding deformed regions.

Conclusions:

• The TRBP dsRBD-dsRNA interaction involves little to no alteration of dsRNA's A-form geometry.
• TRBP likely selects binding sites with ideal A-form structure, excluding deformed regions.
• This binding mechanism has significant implications for understanding micro-RNA processing efficiency.