Laboratory for Dynamic Structure of Biomolecules

Membrane proteins and RNAs are fundamental to biological processes and serve as primary targets for drug development. While X-ray crystallography and cryo-EM provide high-resolution atomic structures, they often capture only static snapshots. To understand these molecules in their functional, in-situ environments, our team utilizes NMR spectroscopy, which reveals the critical relationships between dynamical structures and biological functions in solution.

     In 2025, we advanced the understanding of DDX3X, an ATP-dependent helicase that unwinds higher-order mRNA structures to regulate gene expression (Toyama, Takeuchi, Shimada. Nat Commun (2025)). Dysfunction in DDX3X is linked to medulloblastoma and neurological disorders. Using our 1 GHz NMR spectrometer, we discovered that the DDX3X core domain recognizes specific RNA structures only in its ATP-bound state. It exhibits a higher affinity for single-stranded RNA (ssRNA) than double-stranded RNA (dsRNA), adopting a "closed" conformation upon binding. To study these high-molecular-weight complexes, we developed a high-sensitivity system using ribose (2'-19F) probes. This allowed us to observe the dynamic equilibrium between RNA states, confirming that ATP-bound DDX3X facilitates unwinding by shifting the equilibrium toward the single-stranded form. Analysis of medulloblastoma-associated mutations (G302V and G325E) revealed a loss of this ssRNA-binding capacity, providing a direct biochemical link to disease progression. Our research further highlighted the role of the N-terminal intrinsically disordered region (N-IDR). We found that the N-IDR specifically recognizes G-quadruplex (GQ) structures with high affinity, while the core domain provides the enzymatic power to unwind them. Bioinformatic analysis corroborated this, showing that mRNAs regulated by DDX3X are enriched with GQ motifs in their untranslated regions (UTRs). Finally, we enhanced NMR methodology by establishing a theoretical framework for 1H-19F magnetic interactions, improving the detection of RNA-protein interactions (Toyama and Shimada, J Magn Reson (2025)). Collectively, these findings provide essential insights for drug discovery targeting DDX3X and mRNA structures.