A relation between inhibition of protein synthesis and conformation of 5′-phosphorylated 7-methylguanosine derivatives

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The inhibition of protein synthesis by 5′-phosphorylated derivatives of 7-methylguanosine and 7-methylinosine has been studied in a wheat germ cell-free system programmed by exogenous messenger RNA. The di- and triphosphate derivatives were found to inhibit protein synthesis at lower concentrations than 7-methylguanosine-5′-monophosphate. Inosine derivatives were consistently poor inhibitors when tested with different eucaryotic and viral RNAs. Dinucleotides derived from G5′ppp5′A by methylation of guanosine in the 7 position and/or of the adenosine in the 2′-O position were also tested as inhibitors of protein synthesis. A dinucleotide without the 7-methyl group is not inhibitory. Dinucleotides containing m7G‡ inhibit protein synthesis at about the same concentration as m7G5′ppp. The presence of the 2′-O methyl group has no effect on the inhibition of protein synthesis. A correlation between inhibition of protein synthesis and conformation in solution of the phosphorylated derivatives of m7G has been sought by nuclear magnetic resonance analysis. In aqueous solutions m7G exhibits considerable conformational freedom and various conformers are present. In the 5′-phosphorylated derivatives of m7G a very dramatic effect on the conformational freedom of the backbone is observed. This results in an increase in conformational purity and loss of the flexibility about the C-4′−C-5′ bond and a significant increase in the population of 3E conformers of the ribofuranose ring. Little conformational difference is observed between the di- and triphosphate derivatives of m7G. It seems likely that the electrostatic interaction between the positively charged N-7 and the negatively charged phosphate groups plays the dominant role in imparting conformational “rigidity” to the backbone of the 5′-phosphorylated derivatives of m7G.