Enzymatic acylation of histidine to tobacco mosaic virus RNA

R. Salomon*, I. Sela, H. Soreq, D. Giveon, U. Z. Littauer

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

A method for the purification of rat liver histidyl-tRNA synthetase was developed. The enzyme, freed of trace nuclease contamination by affinity chromatography on DNA-Sepharose, could catalyze the aminoacylation of tobacco mosaic virus (TMV) RNA with histidine. The aminoacylation reaction of TMV RNA was inhibited by KCl, while that of rat liver tRNA exhibited a maximum rate at 140 mM KCl. TMV N-acetyl [3H]histidyl-RNA was prepared in order to stabilize the TMV histidyl-RNA ester bond. The TMV N-acetyl [3H]histidyl-RNA showed a migration identical to that of unacylated TMV RNA upon electrophoresis in polyacrylamide agarose gels. Removal of 5 to 10 nucleoside residues from the 3′-terminus using Escherichia coli polynucleotide phosphorylase or oxidation of the 3′-terminal ribose with periodate (to produce TMV RNAox) eliminated the aminoacylation capacity of TMV RNA as well as its infectivity in tobacco leaves. It is concluded that the aminoacylation site is located at the 3′-terminal adenosine of TMV RNA. Fragmentation of TMV RNAox with a crude rat liver enzyme did not expose additional sites for acylation with histidine, nor did it produce acylation capacity for other amino acids. The reduction of TMV RNAox with sodium borohydride converted the terminal dialdehyde group to a dialcohol-lacking covalent bond between the C2′ and C3′ of the terminal adenosine, but did not restore its ability to accept histidine or its infectivity. Similar reduction of rat liver tRNAox to tRNAox-red, however, did restore its ability to accept phenylalanine while activity for histidine was not regained.

Original languageEnglish
Pages (from-to)74-84
Number of pages11
JournalVirology
Volume71
Issue number1
DOIs
StatePublished - May 1976
Externally publishedYes

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