Open AccessImproved procedure for the isolation of a double‐strand‐specific ribonuclease and its application to structural analysis of various 5S rRNAs and tRNAs
Author(s) -
DIGWEED Martin,
PIELER Tomas,
KLUWE Dagmar,
SCHUSTER Lennart,
WALKER Richard,
ERDMANN Volker A.
Publication year - 1986
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1986.tb09355.x
Subject(s) - rnase p , biology , nuclease , ribonuclease , ribosomal rna , transfer rna , biochemistry , rna , rnase h , escherichia coli , protein secondary structure , dna , microbiology and biotechnology , gene
An improved method for the isolation of a double‐strand‐specific RNase from snake venom is presented. This RNase, called CSV, was used to cleave yeast tRNA Phe and tRNA Glu 2 and tRNA Met f from Escherichia coli. In addition these RNAs and E. coli tRNA Phe were examined with the single‐strand‐specific nuclease S1. The results are discussed in terms of the specificity of CSV RNase and the structure of tRNAs. S1 nuclease digestions at increasing temperatures allowed the melting of tertiary and secondary structure to be monitored. 5S rRNA from E. coli, Thermoplasma acidophilum and the chloroplasts of Spinacia oleracea were digested with CSV and S1. The information these results give on the secondary‐structural differences between different classes of 5S rRNA are discussed. Supporting evidence is found for tertiary interactions between hairpin loop c and internal loop d of eubacterial 5S rRNA.