Evidence on the conformation of HeLa-cell 5.8S ribosomal ribonucleic acid from the reaction of specific cytidine residues with sodium bisulphite

Abstract

The reaction of HeLa-cell 5.8 S rRNA with NaHSO3 under conditions in which exposed cytidine residues are deaminated to uridine was studied. It was possible to estimate the reactivities of most of the 46 cytidine residues in the nucleotide sequence by comparing 'fingerprints' of the bisulphite-treated RNA with those of untreated RNA. The findings were consistent with the main features of the secondary-structure model for mammalian 5.8 S rRNA proposed by Nazar, Sitz & Busch [J. Biol. Chem. (1975) 250,[8591][8592][8593][8594][8595][8596][8597]. Five out of six regions that are depicted in the model as single-stranded loops contain cytidine residues that are reactive towards bisulphite at 25°C (the other loop contains no cytidine). The cytidine residue nearest to the 3'-terminus is also reactive. Several cytidine residues that are internally located within proposed double-helical regions show little or no reactivity towards bisulphite, but the cytidine residues of several C. G pairs at the ends of helical regions show some reactivity, and one of the proposed loops appears to contain six nucleotides, rather than the minimum of four suggested by the primary structure. Two cytidine residues that are thought to be 'looped out' by small helix imperfections also show some reactivity.

Chemical modification studies and the secondary structure of HeLa cell 5.8S rRNA

“…Several G residues were inferred to be unreactive where the previous bisulphite data (11) and the structural models strongly indicated base-pairing with C.…”

“…5.8 S rRNA was obtained as in ref. 3 and was purified by rerunning on a 10-25% sucrose gradient (11).…”

“…However, a number of these streaked or were recovered in low yields and a complete analysis was not undertaken. Tables 1 and 2 give the recoveries of (11,25).…”

“…Nucleic Acids Research Nucleic Acids Research have described the reaction of specific cytidine residues in HeLa cell 5.8 S rRNA with sodium bisulphite (11).…”

“…Removal of unreacted NaS03 and of bisulphite adducts, and further preparation of the treated 5.8 S rRNA for fingerprinting, were carried out as described (11).…”