Efficiency of the 5'-terminal sequence (omega) of tobacco mosaic virus RNA for the initiation of eukaryotic gene translation in Escherichia coli.
Recent studies have demonstrated that the 5' leader (omega sequence) of tobacco mosaic virus RNA has a certain enhancing capacity for translation of mRNA in both prokaryotes and eukaryotes. In order to estimate the efficiency of omega to initiate translation of mRNA in Escherichia coli, in comparison to the Shine-Dalgarno (S/D) sequence, we have inserted eight different eukaryotic genes into two types of E. coli expression vectors containing one constitutive promoter (P1) but different translation-initiation sites (S/D or omega delta 3 sequence, respectively). The efficiency of transcription and translation in vivo was evaluated for these vectors by measuring the yield of protein and both the level and stability of mRNA. We report that substitution of omega delta 3 for S/D decreases the yield of expressed protein 4-1900-fold and the content of gene-specific mRNA is decreased by about sevenfold. However, in comparison with the S/D sequence, the level of protein expressed under the translational control of omega delta 3 is less sensitive to changes in the 5' coding region. We also report that the omega sequence contains a region of 10-12 nucleotides complementary to the small ribosomal subunit RNA (rRNA) of E. coli, Eikenella corrodens and Xenopus laevis, and to the rRNA of the (small ribosomal) subunit of Oryza sativa.
Ivanov
I G
IG
Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia.
Alexandrova
R
R
Dragulev
B
B
Leclerc
D
D
Saraffova
A
A
Maximova
V
V
Abouhaidar
M G
MG
D000595:Amino Acid Sequence; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid; D014027:Tobacco Mosaic Virus / Q000235:genetics*; D014158:Transcription, Genetic; D014169:Transformation, Bacterial
D009841:Oligonucleotides; D012333:RNA, Messenger; D012367:RNA, Viral