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Initiation of DNA synthesis on single-stranded DNA templates in vitro promoted by the bacteriophage lambda O and P

J H LeBowitz, R McMacken

    Advances in Experimental Medicine and Biology
    |January 1, 1984
    PubMed

    Insights

    Bacteriophage lambda O and P proteins initiate DNA replication by interacting with E. coli proteins to form a prepriming intermediate. This intermediate enables efficient replication of single-stranded phage chromosomes, offering insights into lambda DNA replication.

    Area of Science:

    • Molecular Biology
    • Virology
    • Biochemistry

    Background:

    • Bacteriophage lambda utilizes specific initiator proteins for DNA replication.
    • Replication of single-stranded DNA phages like M13 and phi X174 requires multiple host factors.

    Purpose of the Study:

    • To elucidate the mechanism by which bacteriophage lambda O and P proteins initiate DNA replication.
    • To investigate the roles of Escherichia coli replication proteins in M13 and phi X174 DNA synthesis.

    Main Methods:

    • In vitro DNA synthesis assays using purified proteins.
    • Gel filtration chromatography to isolate reaction intermediates.
    • Characterization of protein-protein interactions.

    Main Results:

    • Lambda O and P proteins, with six E. coli proteins, replicate M13 and phi X174 single-stranded DNA.
    • An ATP-dependent step involves lambda O/P and E. coli dnaJ/dnaK proteins, transferring dnaB protein to ssDNA.
    • A stable prepriming intermediate is formed, which is efficiently primed and replicated by primase and DNA polymerase III holoenzyme.

    Conclusions:

    • The identified proteins and mechanism are crucial for M13 and phi X174 replication.
    • The findings provide a model for O and P protein-dependent initiation of DNA synthesis.
    • This mechanism may be relevant to in vivo initiation events during bacteriophage lambda DNA replication.

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