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Mutant EF-Tu increases missense error in vitro.

S Tapio, C G Kurland

    Molecular & General Genetics : MGG
    |October 1, 1986
    PubMed
    Summary
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    Mutations in elongation factor EF-Tu affect bacterial ribosome proofreading and missense errors. The EF-Tu Ar mutant increases errors during polypeptide synthesis, similar to kanamycin action.

    Area of Science:

    • Molecular Biology
    • Bacterial Protein Synthesis

    Background:

    • Elongation Factor EF-Tu (EF-Tu) is crucial for bacterial protein synthesis, mediating aminoacyl-tRNA binding to the ribosome.
    • Ribosomal fidelity is maintained through initial selection and proofreading steps, minimizing errors during translation.

    Purpose of the Study:

    • To investigate the impact of EF-Tu structural mutations on missense errors and ribosomal proofreading activity in vitro.
    • To elucidate the kinetic basis for altered fidelity in mutant EF-Tu systems.

    Main Methods:

    • In vitro studies using bacterial ribosomes.
    • Analysis of polypeptide synthesis with wild-type and mutant EF-Tu forms (EF-Tu Bo, EF-Tu Ar).
    • Kinetic analysis of missense error incorporation using specific leucine isoacceptors.

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    Main Results:

    • The EF-Tu Bo mutant is inactive in polypeptide synthesis despite binding aminoacyl-tRNA.
    • The EF-Tu Ar mutant exhibits significantly increased missense incorporation for leucine isoacceptors 2 and 4.
    • This enhanced missense frequency is attributed to increased errors in both proofreading and initial selection steps.

    Conclusions:

    • Mutational alterations in EF-Tu can directly impact ribosomal fidelity.
    • The EF-Tu Ar mutation enhances missense errors by compromising both proofreading and initial selection mechanisms.
    • The observed effect of EF-Tu Ar is analogous to the mechanism of action of the antibiotic kanamycin.