Conformational dynamics at microsecond timescale in the RNA-binding regions of dsRNA-binding domains

Abstract

Double-stranded RNA-binding domains (dsRBDs) are involved in a variety of biological functions via recognition and processing of dsRNAs. Though the primary substrate of the dsRBDs are dsRNAs with A-form helical geometry; they are known to interact with structurally diverse dsRNAs. Here, we have employed two model dsRBDs -TAR-RNA binding protein and Adenosine deaminase that acts on RNA -to understand the role of intrinsic protein dynamics in RNA binding. We have performed a detailed characterization of the residue-level dynamics by NMR spectroscopy for the two dsRBDs.While the dynamics profiles at the ps-ns timescale of the two dsRBDs were found to be different, a striking similarity was observed in the μ s-ms timescale dynamics for both the dsRBDs. Motions at fast μ s timescale (k ex > 50000 s -1 ) were found to be present not only in the RNA-binding residues but also in some allosteric residues of the dsRBDs. We propose that this intrinsic μ s timescale dynamics in two distinct dsRBDs allows them to undergo conformational rearrangement that may aid dsRBDs to target substrate dsRNA from the pool of structurally different RNAs in cellular environment.

Role of protein dynamics in enthalpy-driven recognition of topologically distinct dsRNAs by dsRBDs

“…A recent study from our lab has reported the presence of microsecond timescale dynamics in the two dsRBDs. Based on these results, we have proposed that the conformational ensemble of the dsRBDs might be responsible for the recognition of conformationally distinct target dsRNAs (38).…”

“…NMR relaxation data analysis for the D10RNA-bound TRBP2-dsRBD1 was carried out in a similar way as was done for apo-TRBP2-dsRBD1 described elsewhere (38).…”

“…The presence of all the peaks in the D10RNA-bound dsRBD 1 H-15 N HSQC spectrum allowed us to probe the dynamics of TRBP2-dsRBD1 in the presence of D10RNA, which could not be done in the presence of larger dsRNAs due to extensive line-broadening. Fast dynamics data (R 1 , R 2 , [ 1 H]-15 N nOe) could be measured for 58 residues of TRBP2-dsRBD1 bound to D10RNA as against for 69 residues for apo-dsRBD (38). Overall profile for the relaxation parameters (R 1 , R 2 , [ 1 H]-15 N nOe) was found to be strikingly similar between apo-and dsRNA-bound state ( Figure S8) except for a marginal dip in R 1 and R 2 values for the core residues of the protein (highlighted by a horizontal red line in Figure S8) (38).…”

“…Fast dynamics data (R 1 , R 2 , [ 1 H]-15 N nOe) could be measured for 58 residues of TRBP2-dsRBD1 bound to D10RNA as against for 69 residues for apo-dsRBD (38). Overall profile for the relaxation parameters (R 1 , R 2 , [ 1 H]-15 N nOe) was found to be strikingly similar between apo-and dsRNA-bound state ( Figure S8) except for a marginal dip in R 1 and R 2 values for the core residues of the protein (highlighted by a horizontal red line in Figure S8) (38). It was concluded from the data that the D10RNA binding did not perturb the overall fast dynamics of the protein.…”

“…In HARD experiments, the relaxation rates (R 1 , R 1ρ , and R 2ρ ) along with rotational correlation time calculated from quadric_diffusion program (54) was used to fit the data to the time-dependent Bloch-McConnell equation assuming a two-site exchange model as was done for the apo-state of the protein (38,51). The solution to the equation by using geometric approximation method allowed to extract the dynamic parameters (the conformational exchange rate, k ex , the equilibrium populations of the two states, p A or p B , and the chemical shift difference, Δ ω…”