| Autor: |
de Smit MH; Department of Biochemistry, Gorlaeus Laboratories, Leiden University, The Netherlands., van Duin J, van Knippenberg PH, van Eijk HG |
| Jazyk: |
angličtina |
| Zdroj: |
Gene [Gene] 1994 May 27; Vol. 143 (1), pp. 43-7. |
| DOI: |
10.1016/0378-1119(94)90602-5 |
| Abstrakt: |
To activate expression of a human transferrin (Tf)-encoding cDNA in Escherichia coli by translational coupling, it was placed in an expression plasmid downstream from a 5'-terminal fragment from the replicase (R)-encoding gene of bacteriophage MS2. The resulting construct was found to produce, besides the desired Tf, a protein with the mobility of a fusion product (RTf) of the N-terminal R fragment and Tf. Analysis of available mutants showed that this fusion results from +1 ribosomal frameshifting at the end of the R reading frame. This region contains the sequence, CCC.UGA, suggesting that before termination occurs, tRNA(Pro) may dislodge from the CCC codon and reassociate with the +1 triplet CCU. By further site-directed mutagenesis, we demonstrate that both the CCC codon and the termination codon are indeed required for the observed 2-4% frameshifting. When either triplet is changed, the frequency of frameshifting drops to 0.3% or less. These results classify CCC.UGA as a new '+1 shifty stop'. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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