A new study in Nature Communications presents a detailed look at how a self-splicing ribozyme builds its functional structure. The work, led by Marco Marcia at Uppsala University, follows the folding of a Group II intron with a level of temporal and structural resolution not previously achieved for an RNA of this size.
The team combined cryo-electron microscopy, small-angle X-ray scattering, RNA biochemistry, enzymology, and molecular simulations. This integrative strategy produced a near-continuous series of conformations that document how the ribozyme avoids kinetic traps on its way to an active state.
“Determining RNA structures is a challenging task—the inherent flexibility and negative charge make RNA a notoriously difficult target for structural studies,” said Shekhar Jadhav, postdoc at Uppsala University. He added that extensive screening on electron microscopes finally made it possible to observe these RNA dynamics.
The results show that Domain 1 acts as a scaffold that cues the arrival of additional domains, guiding the ribozyme through a defined sequence of folding events that culminate in its catalytic form. Molecular simulations performed at the Istituto Italiano di Tecnologia helped clarify these transitions.
Image-processing methods developed at the Centre for Structural Systems Biology were essential for identifying intermediate conformations among hundreds of thousands of particles. “To capture these fleeting frames, we had to develop novel cryo-EM image-processing strategies,” said Maya Topf, collaborator and Group Leader at CSSB. SAXS and molecular dynamics simulations supported these reconstructions by estimating the small energetic shifts required for each structural transition.
Group II introns are considered ancestors of the spliceosome, which adds evolutionary context to their cooperative folding pathways. The detailed structural intermediates presented in this study have already been incorporated into CASP assessments and form a reference set for testing AI-based RNA structure prediction.
Publication Details
Jadhav, S., Maiorca, M., Manigrasso, J. et al. Dynamic assembly of a large multidomain ribozyme visualized by cryo-electron microscopy. Nat Commun 16, 10195 (2025). https://doi.org/10.1038/s41467-025-65502-8
The team combined cryo-electron microscopy, small-angle X-ray scattering, RNA biochemistry, enzymology, and molecular simulations. This integrative strategy produced a near-continuous series of conformations that document how the ribozyme avoids kinetic traps on its way to an active state.
“Determining RNA structures is a challenging task—the inherent flexibility and negative charge make RNA a notoriously difficult target for structural studies,” said Shekhar Jadhav, postdoc at Uppsala University. He added that extensive screening on electron microscopes finally made it possible to observe these RNA dynamics.
The results show that Domain 1 acts as a scaffold that cues the arrival of additional domains, guiding the ribozyme through a defined sequence of folding events that culminate in its catalytic form. Molecular simulations performed at the Istituto Italiano di Tecnologia helped clarify these transitions.
Image-processing methods developed at the Centre for Structural Systems Biology were essential for identifying intermediate conformations among hundreds of thousands of particles. “To capture these fleeting frames, we had to develop novel cryo-EM image-processing strategies,” said Maya Topf, collaborator and Group Leader at CSSB. SAXS and molecular dynamics simulations supported these reconstructions by estimating the small energetic shifts required for each structural transition.
Group II introns are considered ancestors of the spliceosome, which adds evolutionary context to their cooperative folding pathways. The detailed structural intermediates presented in this study have already been incorporated into CASP assessments and form a reference set for testing AI-based RNA structure prediction.
Publication Details
Jadhav, S., Maiorca, M., Manigrasso, J. et al. Dynamic assembly of a large multidomain ribozyme visualized by cryo-electron microscopy. Nat Commun 16, 10195 (2025). https://doi.org/10.1038/s41467-025-65502-8