Analysis of the structure and folding of the 3' genomic RNA of flaviviruses.

The structure of the 3' genomic RNA of flaviviruses has been analyzed by thermal melting, and by chemical and enzymatic probing of model RNAs. The 3' RNA had more transitions than could be assigned to individual stem-loops and a transition in the 40-45 degrees C range was assigned to tertiary structure unfolding. In support of this assignment, mutants designed to destabilize the proposed pseudoknot tertiary structure lose the 40-45 degrees C transition. Enzyme cleavage of the RNA and chemical probing as a function of temperature also support a pseudoknot tertiary structure for the 3' flavivirus RNA.

Persistence length of RNA.

A set of 10 double-stranded RNAs ranging in length from 92 to 317 base pairs has been synthesized, and their rotational diffusion times were measured by transient electric birefringence. A hydrodynamic analysis of the birefringence decay data yields a helix persistence length of 720 +/- 70 A, based on a helix with an effective hydrodynamic diameter of 26 A and a rise per base pair of 2.7 A in the presence of Mg2+ ions. Thus, duplex RNA is somewhat stiffer than DNA, for which the persistence length is 450-500 A. The measurements also suggest that (i) current hydrodynamic theory for the rotational decay times is applicable to RNA molecules that are longer than 100 base pairs and (ii) weak Mg(2+)-RNA association decreases the rise per base pair slightly, by no more than 0.1 A.