Measuring the 3D Size of Large RNA Molecules

Large single-stranded (ss) RNAs are ubiquitous in cells and constitute the genomic content of many viral species. Besides being the primary means of intra-cellular information transfer, some of their functions require them to form stable structural motifs. ssRNA molecules possess intrinsic self-complementarity leading to a partially double-stranded, branched, secondary structure. We measure, in solution, the physical dimensions of several sequences of ssRNA ranging from a few hundred to a few thousand nucleotides in length. Sizes are reported as radii of gyration ($R_g$) and hydrodynamic radii ($R_h$), respectively determined by small-angle x-ray scattering (SAXS) and fluorescence correlation spectroscopy (FCS). For RNAs of fixed nucleotide length ($\sim$2000) and composition, we find that $R_g$s and $R_h$s can vary by over 30$\%$. By changing solvent conditions, we demonstrate that these size discrepancies are a generic property of the secondary structure arising from sequence-dependent base-pairing. Some viral RNAs that self-assemble into spherical protein capsids have highly evolved sequences that code for unusually compact size and shape.