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Nucleic Acids Research, 1974, Vol. 1, No. 10 1305-1316
© 1974

Articles

A simplified method for study of RNA conformation - reaction of formylmethionine transfer RNA with [14c]methylamine-bisulfite

LaDonne H. Schulman*, Robert Shapiro+, David C. F. Law+ and Judith B. Louis+

*Department of Developmental Biology and Cancer, Division of Biology, Albert Einstein College of Medicine Bronx, New York, NY 10461, USA +Department of Chemistry, New York University New York, NY 10003, USA

Received August 8, 1974. A new chemical method for radioactive labeling of single-stranded regions of RNA has been used to probe the three-dimensional structure of E. coli tRNAfMet in solution. The procedure involves conversion of cytosine residues to N4-[14C]methylcytosines by treatment with. 14CH3NH2 and sodium bisulfite at pH7. Ribonuclease digestion of the modified tRNA produces 14C-labeled oligo-nucleotides which comigrate with the corresponding unlabeled oligormcleotides, facilitating structural analysis. By this procedure, E. coli tRNAfmet has been found to contain only six reactive cytosines: C1 C16, C17, C35, C75 and C76. In addition, slow reaction at C33m, was observed. These results are in excellent agreement with previously reported data on the sites of exposed cytosine residues in tRNAfmet obtained by two other chemical methods. The methylamine-bisulfite procedure is recommended for studying the ordered structure of more complex polyribonucleotides such as viral and ribosomal RNAs.


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