Effect of polyamines on translation fidelity in vivo

Abstract

Polyamines, when added to cell-free protein-synthesizing systems, have been shown either to reduce mistranslation or to increase it depending upon the composition of the reaction mixture. To study this question under conditions as natural as possible we investigated the effects of polyamines on the fidelity of protein synthesis in intact Escherichiu coli bacterial cells, using strains which were auxotrophic for polyamine synthesis. Error was measured in two ways: (a) the incorporation of [3H]histidine into coat protein of bacteriophage MS2, the gene of which does not code for histidine, and (b) the synthesis of a basic variant of MS2 coat protein in which a lysine erroneously replaces an asparagine, causing a change in isoelectric point. We found that when cell cultures were supplemented with polyamines there was no effect on the first type of error and the second type decreased twofold. The measured errors occurred at the level of translation because their frequency increased in the presence of streptomycin and decreased in cells bearing a streptomycin-resistance mutation known to lower the occurrence of translational misreading. The average erroneous incorporation per mol coat protein in the presence of polyamines was 1.43 ? 0.59 mmol histidine and 25 -34 mmol lysine/asparagine substitution. The reason for the different effect of polyamines on the two types of error is not known but could be related to the difference between their corresponding frequencies or to codon-specific effects.Polyamines are widely distributed organic cations which appear to have many roles, especially in processes involving nucleic acids, such as DNA replication, transcription and translation [I -51. Studies carried out with cell-free systems have demonstrated that polyamines can replace part of the magnesium requirement of protein synthesis and at the same time cause overall stimulation [6 -81. Polyamines have also been shown to have a qualitative effect on the proteins synthesized. Adenovirus proteins made in vitro in the presence of polyamines resembled more closely those synthesized in vivo, both in size and quantity [9]. Polypeptides of higher molecular mass were found in smaller amounts in the absence of polyamines in vitro [lo, 111 and in vivo [12]. These observations had suggested that polyamines might prevent premature peptide termination or otherwise affect the fidelity of translation. It was subsequently reported that polyamines, particularly spermidine, decreased the misincorporation of leucine for phenylalanine severalfold in a polyuridylic-acid-dependent in vitro translation system [13-151. However, in these studies the error determinations in presence of polyamines were done at lower Mg concentrations than those in the absence of polyamines because the Mg optimum for polyphenylalanine synthesis was higher under the latter conditions than in the presence of polyamines. In one study, other changes besides polyamine addition were made [16]. At some concentrations of Mg, however, the error was no different or [18] that when...

Roles of polyamines in translation

“…In both bacterial and wheat germ extracts, increasing the concentration of spermidine from 1 to 3 mM decreased the rate of misincorporation of leucine in place of phenylalanine by ribosomes programmed with poly(U) mRNA-encoding polyphenylalanine (53,54). Consistent with these findings, the relatively high level of misreading observed in E. coli strains that were defective or auxotrophic for polyamine synthesis was partially suppressed by addition of putrescine to the media (55). In contrast to these studies linking increased polyamines to enhanced fidelity during translation elongation, high polyamines were linked to stop codon readthrough on the ␤-globin mRNA in rabbit reticulocyte lysate (56).…”

Programmed Translational Frameshifting

“…They regulate the fidelity of aminoacylation; increasing concentrations of polyamines reduce the frequency of misacylation, especially in the absence of Mg 2ϩ (116). This effect might explain why increasing the concentration of polyamines increases the fidelity of protein synthesis in vitro (88) and in vivo (132). However, the effect of polyamines on translation appears to be more complex, affecting the conformation of aminoacyl-tRNAs and thus the efficiency and accuracy of codon-anticodon recognition (reference 145 and references therein).…”

Regioselective binding of spermine, N¹,N12-bismethylspermine, and N¹,N12-bisethylspermine to tRNA Phe as revealed by 750 MHz ¹H-NMR and its possible correlation with cell cycling and cytotoxicity

“…The same was true for the N α ,N ω -bismethyl derivatives; the strongest hydrogen bonds between the polyamine and tRNA were those formed by the central nitrogens (N 4 and N 9 ), independent of the alkyl substituent (methyl or ethyl) present at the terminal nitrogen groups. Binding of polyamines to tRNA has been shown to be relevant to the fidelity of the translation process [16][17][18][19] , as well as to codon-anticodon recognition and ribosomal binding [20][21] . Polyamines also stimulate aminoacylation in general and their effects on the structure of tRNA compelled attention to the possible role of these compounds for protein synthesis 22 .…”