Mg2+-dependent compaction and folding of yeast tRNAPhe and the catalytic domain of the B. subtilis RNase P RNA determined by small-angle X-ray scattering.

We apply synchrotron-based small-angle X-ray scattering to investigate the relationship between compaction, metal binding, and structure formation of two RNAs at 37 degrees C: the 76 nucleotide yeast tRNA(Phe) and the 255 nucleotide catalytic domain of the Bacillus subtilis RNase P RNA. For both RNAs, this method provides direct evidence for the population of a distinct folding intermediate. The relative compaction between the intermediate and the native state does not correlate with the size of the RNA but does correlate well with the amount of surface burial as quantified previously by the urea-dependent m-value. The total compaction process can be described in two major stages. Starting from a completely unfolded state (4-8 M urea, no Mg(2+)), the major amount of compaction occurs upon the dilution of the denaturant and the addition of micromolar amounts of Mg(2+) to form the intermediate. The native state forms in a single transition from the intermediate state upon cooperative binding of three to four Mg(2+) ions. The characterization of this intermediate by small-angle X-ray scattering lends strong support for the cooperative Mg(2+)-binding model to describe the stability of a tertiary RNA.