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Effect of Rauscher leukemia virus-specific proteins on reverse transcriptase. Binding between reverse transcriptase

A K Bandyopadhyay

    The Journal of Biological Chemistry
    |August 25, 1977
    PubMed
    Summary
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    Researchers purified Rauscher leukemia virus reverse transcriptase and found it forms a complex with protein p30. This reverse transcriptase-p30 complex shows enhanced enzymatic activity and can be dissociated with salt.

    Area of Science:

    • Virology
    • Molecular Biology
    • Biochemistry

    Background:

    • Rauscher leukemia virus (RLV) is a retrovirus known to possess reverse transcriptase activity.
    • The interaction between viral proteins and enzymes can significantly influence viral replication and function.

    Purpose of the Study:

    • To purify the reverse transcriptase enzyme from Rauscher leukemia virus.
    • To investigate the interaction between purified reverse transcriptase and the major internal protein p30.
    • To characterize the properties and enzymatic activity of the reverse transcriptase-p30 complex.

    Main Methods:

    • Purification of reverse transcriptase via sequential phosphocellulose and DNA-cellulose column chromatography.
    • Characterization of the enzyme and complex using molecular weight determination (SDS-PAGE, Sepharose 6B chromatography), sedimentation analysis (glycerol gradient centrifugation), and enzymatic assays.

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  • Immunoprecipitation using monospecific antisera against reverse transcriptase and p30.

    • Main Results:

    • Homogeneous reverse transcriptase was obtained as a single polypeptide chain (approx. 70,000 MW).
    • A complex formed between reverse transcriptase and p30, exhibiting a 2- to 3-fold stimulation of [3H]TMP incorporation.
    • The complex had an estimated molecular weight of 400,000 and dissociated in 0.8 M KCl.

    Conclusions:

    • Reverse transcriptase from Rauscher leukemia virus forms a functional complex with the p30 protein.
    • The p30 protein enhances the enzymatic activity of reverse transcriptase.
    • The reverse transcriptase-p30 complex is a distinct molecular entity that can be dissociated under specific conditions.