Evidence for tertiary structure in natural single stranded RNAs in solution.

Binding isotherms (20 degrees C) of ethidium bromide to a number of tRNA species at various ionic strengths indicate that i) the number ni of intercalation sites is high 7 to 11 per molecule, in the low salt form III, but small, 2 to 1, at high Mg2+ or Na+ when form I predominates. ii) modification of tRNA at strategic positions for 3D folding prevents full expression of intercalation restriction iii) maximal restriction is obtained at salt concentrations higher than needed for full conversion to form I. It is inferred that restriction, which is not observed with bihelical RNA (or DNA), requires the native tRNA 3D structure but also some physical coupling between the region of 3D folding and bihelical arms. Ribosomal RNAs, some viral RNAs, mRNA from sheep mammary gland as well as the random copolymers Poly UG, Poly AUG, Poly AUCG all exhibit intercalation restriction. Hence 3D folding of the polyribonucleotide chains appears to be a feature common to single-stranded RNAs when free in solution under physiological conditions.