Inhibition of eukaryotic translation by analogs of messenger RNA 5'-cap: chemical and biological consequences of 5'-phosphate modifications of 7-methylguanosine 5'-monophosphate (original) (raw)

New analogues of 7-methylguanosine 5'-monophosphate (m7GMP) were synthesized with modified 5'-phosphate moieties by replacement of-0 with-H,-CH3, or-NH2. Additional analogues were synthesized with &methyl-or 8-aminoguanine base substitutions or ring-opened ribose (2',3'-diol). These compounds were analyzed by 'H and 31P N M R for solution conformation. In addition, they were also analyzed for biological activity as analogues of mRNA 5'-caps by competition as inhibitors of translation in reticulocyte lysate. Substitution of oxygen on the 5'-monophosphate moiety by-H and-CH3 diminished the activity of the cap analogue as a competitive inhibitor; however, replacement by-NH2 did not diminish the activity of the analogue as an inhibitor. It was inferred from this result that cap binding proteins require a hydrogen bond acceptor as opposed to having an exclusive requirement for a second anionic group on the a-phosphate moiety. Inhibition results obtained with C8-substituted m7GMP analogues indicated that the 8-amino derivative was a better inhibitor than the 8-methyl derivative of m7GMP. The former is primarily anti whereas the latter is primarily syn with respect to glycosidic bond conformation. This result further supports the model that the anti conformation is the preferred form of the cap structure for interaction with cap binding proteins. The 2',3'-diol derivative of m7GMP was inactive as an inhibitor of translation. T a n s l a t i o n of eukaryotic mRNA is dependent on several aspects of mRNA structure. These macromolecules are for the most part monocistronic, possess 5'-termini of the form m7G(5')ppp(5')N, and are polyadenylylated on the 3'-end. The 5' modification is known as a cap and is necessary for optimum translation. It is recognized by at least three translation initiation factors, namely, eIF-4A,' eIF-4B, and eIF-4F (Grifo et al., 1983; Edery et al., 1983, 1985). These are also collectively termed cap binding proteins (CBPs; Shatkin, 1985). These factors bind at or near the 5'-cap of mRNA early in the initiation phase of protein biosynthesis and facilitate mRNA attachment to the 40s ribosomal subunit. Cap analogues inhibit this step of initiation both in complete translation assay (