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Purification of bacteriophage λO protein that specifically binds to the origin of replication

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Summary

By means of a nitrocellulose filter binding assay, DNA binding activities among proteins fractionated from extracts of Escherichia coli carrying λdv have been surveyed. An activity was found that binds specifically to a fragment of 164 base pairs that specifies the λ replication origin (λ ori). This activity was not detected in an extract of cells not carrying the λdv plasmid. The activity was detected in extracts of cells carrying a hybrid plasmid in which the entire λ O gene had been cloned and placed under the control of the lac promoter. Deletion of a 60 base pair segment in the ‘amino-terminal region’ of the O gene abolished this activity, indicating that the λ ori binding protein is coded for by the λ O gene.

The ori-specific binding protein was purified by five fractionation steps. The most purified preparation consists of a major polypeptide that migrates with a molecular weight of 32,000 in SDS-polyacrylamide gel electrophoresis. Binding of O protein to ori occurs in the absence of other protein aceous components.

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Authors and Affiliations

  1. Laboratory of Molecular Genetics, Osaka University Medical School, Kita-ku, 530, Osaka, Japan

    Toshiki Tsurimoto & Kenichi Matsubara

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  1. Toshiki Tsurimoto
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  2. Kenichi Matsubara
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Communicated by T. Yura

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Tsurimoto, T., Matsubara, K. Purification of bacteriophage λO protein that specifically binds to the origin of replication. Molec. Gen. Genet. 181, 325–331 (1981). https://doi.org/10.1007/BF00425606

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  • DOI: https://doi.org/10.1007/BF00425606

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