Small-Angle X-ray Scattering (SAXS) Combined with SAXS-Driven Molecular Dynamics for Structural Analysis of...

Nucleic acids (RNA and DNA) play crucial roles in all living organisms and find wide utility in clinical settings. The convergence of rationally designed nucleic acid multistranded assemblies with embedded therapeutic properties has led to the development of a platform based on nucleic acid nanoparticles (NANPs). NANPs incorporate various functional moieties to deliver their combinations to diseased cells in a highly controlled manner. Given that the structure and composition of NANPs can also influence their immunorecognition and biological activities, thorough verification of all designs is essential. We introduce an experimental pipeline for small-angle X-ray scattering (SAXS) to gather structural details about the solution-state NANPs assembled from up to 12 RNA strands. To the best of our knowledge, this study represents the largest multistranded RNA nanoassemblies characterized in this manner to date. We show that synchronized implementation of SAXS-driven molecular dynamics simulations reveals the diverse conformational landscape inhabited by these assemblies and provides insights into their immunorecognition. The developed strategy expands the capabilities of therapeutic nucleic acids and emerging nucleic acid nanotechnologies.