The modification of nucleosides and nucleotides: IV. The reaction of semicarbazide with nucleic acids

Abstract

1. Cytidine residues of yeast RNA have been specifically converted into semicarbazide derivatives without altering any other bases. In this reaction the C-4 amino groups of cytidine residues were converted into a semicarbazido group. 2. tRNA modified with semicarbazide was completely hydrolysed to nucleoside 5′-phosphate with snake venom phosphodiesterase (orthophosphoric diester phosphohydrolase, EC 3.1.4.1). The base composition of the tRNA was estimated after separation of the nucleotides by ion-exchange column chromatography. A quantitative estimation of the modified cytidine residues in the tRNA was achieved by this method. 3. The results of sedimentation analysis and gel filtration of the tRNA modified with semicarbazide indicated that no splitting of the internucleotide bonds of the RNA occurred during the reaction. 4. The temperature-absorbance relationship plotted for the modified tRNA indicated that the secondary structure in the tRNA decreased with increasing extent of modification. 5. The rate of this modification reaction is low when cytosine groups are highly hydrogen bonded (double-stranded DNA). 6. A simple method for estimating the extent and rate of the modification reaction of cytidine residues in nucleic acids is proposed. In this procedure the specific light absorption of the 4-deamino-4-semicarbazidocytidine residue in the range 300–340 mμ at pH 13 is followed.