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Biochemistry. 2019 May 21;58(20):2463-2473. doi: 10.1021/acs.biochem.9b00111. Epub 2019 May 10.

Conformational Stability Adaptation of a Double-Stranded RNA-Binding Domain to Transfer RNA Ligand.

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Laboratoire de Chimie des Processus Biologiques, CNRS-UMR 8229, Collège De France , Université Pierre et Marie Curie , 11 place Marcelin Berthelot , 75231 Paris Cedex 05 , France.
Expression Génétique Microbienne , UMR 8261, CNRS, Université Paris, Institut de Biologie Physico-Chimique , 13 rue Pierre et Marie Curie , 75005 Paris , France.
Laboratoire de Biochimie Théorique, CNRS UPR9080 , Institut de Biologie Physico-Chimique , 13 rue Pierre et Marie Curie , 75005 Paris , France.


The double-stranded RNA-binding domain (dsRBD) is a broadly distributed domain among RNA-maturing enzymes. Although this domain recognizes dsRNA's structures via a conserved canonical structure adopting an α11β2β32 topology, several dsRBDs can accommodate discrete structural extensions expanding further their functional repertoire. How these structural elements engage cooperative communications with the canonical structure and how they contribute to the dsRBD's overall folding are poorly understood. Here, we addressed these issues using the dsRBD of human dihydrouridine synthase-2 (hDus2) (hDus2-dsRBD) as a model. This dsRBD harbors N- and C-terminal extensions, the former being directly involved in the recognition of tRNA substrate of hDus2. These extensions engage residues that form a long-range hydrophobic network (LHN) outside the RNA-binding interface. We show by coarse-grain Brownian dynamics that the Nt-extension and its residues F359 and Y364 rigidify the major folding nucleus of the canonical structure via an indirect effect. hDus2-dsRBD unfolds following a two-state cooperative model, whereas both F359A and Y364A mutants, designed to destabilize this LHN, unfold irreversibly. Structural and computational analyses show that these mutants are unstable due to an increase in the dynamics of the two extensions favoring solvent exposure of α2-helix and weakening the main folding nucleus rigidity. This LHN appears essential for maintaining a thermodynamic stability of the overall system and eventually a functional conformation for tRNA recognition. Altogether, our findings suggest that functional adaptability of extended dsRBDs is promoted by a cooperative hydrophobic coupling between the extensions acting as effectors and the folding nucleus of the canonical structure.

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