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The control of lambda DNA terminase synthesis.

H Murialdo, A Davidson, S Chow

    Nucleic Acids Research
    |January 12, 1987
    PubMed
    Summary
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    Bacteriophage lambda DNA terminase genes (Nu1 and A) have low translation in E. coli. Upstream DNA sequences control this, and their modification boosts terminase production for DNA packaging applications.

    Area of Science:

    • Molecular Biology
    • Genetics
    • Microbiology

    Background:

    • The Nu1 and A genes encode bacteriophage lambda DNA terminase, crucial for viral DNA packaging.
    • These genes exhibit notably low translation efficiency when expressed in E. coli.

    Purpose of the Study:

    • To investigate the regulatory mechanisms underlying the poor translation of bacteriophage lambda DNA terminase genes (Nu1 and A) in E. coli.
    • To identify DNA elements responsible for controlling the translation of these genes.

    Main Methods:

    • Cloning of Nu1 and A genes into expression plasmids.
    • Modification of DNA sequences upstream of the initiation codons.
    • Measurement of gene expression and terminase subunit production.
    • Analysis of the role of the 'cos' site in gene expression.

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    Main Results:

    • Translation control elements are located within the 100 base pairs upstream of the initiation codon.
    • Swapping upstream sequences with those from highly translated genes significantly increased terminase subunit translation.
    • Rare codons and mRNA secondary structure appear to play a minor role in translation control.
    • Elimination of the 'cos' site enhanced terminase production, suggesting its involvement in late gene expression regulation.

    Conclusions:

    • Upstream regulatory sequences are the primary determinants of low translation efficiency for bacteriophage lambda DNA terminase genes.
    • The 'cos' site may negatively regulate late gene expression.
    • Overproduction of terminase subunits can be achieved, offering potential for improved DNA packaging and cosmid mapping techniques.