Protein chain initiation by methionyl-tRNA in wheat embryo.
The mechanism of protein chain initiation has been investigated in a cell-free amino acid incorporation system from wheat embryos dependent on tobacco mosaic virus RNA. Analysis of the N-termini of the labeled peptide products of short-term incubations showed the presence of unblocked methionine. In addition, methionyl-tRNA (Met-tRNA) could be bound to ribosomes at 1.3 mM Mg(++) in a reaction requiring viral RNA, ATP, GTP, and soluble protein factors. Incorporation experiments with the two cytoplasmic Met-tRNAs of wheat germ, an initiating species designated Met-tRNA(i) and a Met-tRNA(m), showed that methionine transfer from Met-tRNA(i) was linear from zero time, while that from Met-tRNA(m) occurred only after an appreciable lag. Analysis of the peptide products showed that methionine transfer from Met-tRNA(i) was predominantly N-terminal. In contrast, methionine transfer from Met-tRNA(m) was exclusively into internal positions. Similar selectivity was observed in the ribosome binding assay; only Met-tRNA(i) showed a strong reaction. These experiments provide strong evidence that in the wheat embryo, cytoplasmic Met-tRNA(i) functions without formylation in the initiation of protein synthesis.
D000596:Amino Acids / Q000032:analysis; D002247:Carbon Isotopes; D002474:Cell-Free System; D066298:In Vitro Techniques; D007700:Kinetics; D008715:Methionine / Q000378:metabolism; D010455:Peptides / Q000032:analysis; D010940:Plant Proteins / Q000096:biosynthesis*; D012343:RNA, Transfer / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D012639:Seeds / Q000378:metabolism*; D014908:Triticum / Q000378:metabolism*
D000596:Amino Acids; D002247:Carbon Isotopes; D010455:Peptides; D010940:Plant Proteins; D012367:RNA, Viral; D012343:RNA, Transfer; D008715:Methionine