A comparison of the solution structures and conformational properties of the somatic and oocyte 5S rRNAs of Xenopus laevis.
The secondary and tertiary structures of Xenopus oocyte and somatic 5S rRNAs were investigated using chemical and enzymatic probes. The accessibility of both RNAs towards single-strand specific nucleases (T1, T2, A and S1) and a helix-specific ribonuclease from cobra venom (RNase V1) was determined. The reactivity of nucleobase N7, N3 and N1 positions towards chemical probes was investigated under native (5 mM MgCl2, 100 mM KCl, 20 degrees C) and semi-denaturing (1 mM EDTA, 20 degrees C) conditions. Ethylnitrosourea was used to identify phosphates not reactive towards alkylation under native conditions. The results obtained confirm the presence of the five helical stems predicted by the consensus secondary structure model of 5S rRNA. The chemical reactivity data indicate that loops C and D are involved in a number of tertiary interactions, and loop E folds into an unusual secondary structure. A comparison of the data obtained for the two types of Xenopus 5S rRNA indicates that the conformations of the oocyte and somatic 5S rRNAs are very similar. However, the data obtained with nucleases under native conditions, and chemical probes under semi-denaturing conditions, reveal that helices III and IV in the somatic 5S rRNA are less stable than the same structures in oocyte 5S rRNA. Using chimeric 5S rRNAs, it was possible to demonstrate that the relative resistance of oocyte 5S rRNA to partial denaturation in 4 M urea is conferred by the five oocyte-specific nucleotide substitutions in loop B/helix III. In contrast, the superior stability of oocyte 5S rRNA in the presence of EDTA is related to a single C substitution at position 79.
Romaniuk
P J
PJ
Department of Biochemistry and Microbiology, University of Victoria, British Columbia, Canada.
de Stevenson
I L
IL
Ehresmann
C
C
Romby
P
P
Ehresmann
B
B
D000818:Animals; D001483:Base Sequence; D055598:Chemical Phenomena; D002621:Chemistry; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009865:Oocytes; D012335:RNA, Ribosomal; D012341:RNA, Ribosomal, 5S; D012260:Ribonucleases / Q000378:metabolism; D012996:Solutions; D014508:Urea; D014982:Xenopus laevis
D012335:RNA, Ribosomal; D012341:RNA, Ribosomal, 5S; D012996:Solutions; D014508:Urea; D012260:Ribonucleases