Reaction of Psoralen with RNA: Specificity and Use as a Probe for Secondary-Structure Analysis

摘要

A variety of techniques has been used to study how psoralen and its derivatives react with RNA. This information has then been used to analyze the secondary structure of different ribosomal RNAs. Paper electrophoresis at pH 3.5 and 8.8 and HPLC has been used to get high-resolution separation of RNA-psoralen adducts. The separated adducts have been analyzed and shown to be primarily uridine adducts with the psoralen reacted at the furan end. The stereochemistry of the major adducts was determined by NMR. The effect of structural transitions on the number and type of adducts was found for several polymers. The effect of psoralen structure on cross linking ability was analyzed. Charged derivatives formed monoadducts very efficiently but did not produce the level of crosslinking obtainable with lower levels of reaction with uncharged derivatives. Secondary structure analysis of D. melanogaster 5S RNA yielded two definite and two tentative crosslinks which support the generally accepted models for 5S structure. Analysis of E. coli 16S RNA by gel techniques yielded 13 cross-links. Evidence is also presented for an interaction between eukaryotic mRNA (5' cap structure) and 18S RNA (hypermodified base am psi) which serves a function analogous to the Shine-Dalgarno sequence in pro karyotes. (ERA citation 08:010734)