The Effect of Polynucleotides on Aminoacyl Transfer Ribonucleic Acid Synthetases

The inhibitory effects of polyuridylic or polyinosinic acid (poly U, poly I) on the formation of glutamyl-RNA by purified rat liver glutamyl transfer RNA (tRNA) synthetase and rabbit liver tRNA have been investigated. Although double reciprocal plots of the data indicated that the inhibition was competitive with respect to tRNA, high levels of poly U or poly I did not lead to 100% inhibition. Instead, a plateau of inhibition was reached, its value depending on the amount of tRNA present. The percentage inhibition observed did not change over a 10-min period. Preincubation of the synthetase with either tRNA or the polynucleotide inhibitors did not lead to any change in the level of inhibition, suggesting that both tRNA and inhibitors interacted with the enzyme reversibly. The results of experiments in which both poly U and poly I were added to the reaction mixture were consistent with both polymers acting at the same site. In addition to a requirement for specific bases, the polynucleotides also required the presence of single stranded regions in order to elicit inhibition. However, lowering the incubation temperature increased the inhibition by poly U but decreased the effect of poly I, suggesting that at different temperatures the affinity of the enzyme for the various polynucleotides may change. Rat liver glutamyl-tRNA synthetase catalyzes an anomalously low ATP-PPi exchange reaction which is stimulated by tRNA or very high concentrations of glutamic acid. Poly U and poly I had no effect on the exchange reaction at the high glutamate concentrations, but the polynucleotides did inhibit the stimulation of this reaction by tRNA. From these results and the competitive nature of the inhibition, it is concluded that the synthetic polynucleotides interfere with the interaction of tRNA and glutamyl-tRNA synthetase. The partial competitive character of the inhibition is discussed in relation to possible models for the interaction of aminoacyl-tRNA synthetases and nucleic acids.