Discrimination between initiation and elongation of protein biosynthesis in yeast: identity assured by a nucleotide modification in the initiator tRNA.
Cytoplasmic initiator tRNAs from plants and fungi possess an unique 2'-phosphoribosyl residue at position 64 of their sequence. In yeast tRNA(iMet), this modified nucleotide located in the T-stem of the tRNA is a 2'-1''-(beta-O-ribofuranosyl-5''-phosphoryl)-adenosine. The phosphoribosyl residue of this modified nucleoside was removed chemically by treatment involving periodate oxidation of tRNA(iMet) and regeneration of the 3'-terminal adenosine with ATP (CTP):tRNA nucleotidyl transferase. The role of phosphoribosylation at position 64 for interaction with elongation factor eEF-1 alpha and initiation factor 2 (eIF-2) was investigated in the homologous yeast system. Whereas the 5'-phosphoribosyl residue prevents the binding of Met-tRNA(iMet) to eEF-1 alpha, it does not influence the interaction with eIF-2. After removal of the ribosyl group, the demodified initiator tRNA showed binding to eEF-1 alpha, but no change was detected with respect to the interaction with the initiation factor eIF-2. This observation is interpreted to mean that a single modification of an eucaryotic initiator tRNA in yeast serves as a negative discriminant for eEF-1 alpha, thus preventing the initiator tRNA(iMet) from entering the elongation cycle of protein biosynthesis.
Förster
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C
Laboratorium für Biochemie, Universität Bayreuth, Germany.
Chakraburtty
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K
Sprinzl
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M
D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000096:biosynthesis*; D010441:Peptide Chain Elongation, Translational; D014176:Protein Biosynthesis; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism
D005656:Fungal Proteins; D012331:RNA, Fungal; D012358:RNA, Transfer, Met