Halobacterium halobium strains lysogenic for phage phi H contain a protein resembling coliphage repressors.
DNA-binding proteins such as bacteriophage repressors belong to the helix-turn-helix family. Ionic interactions drive DNA binding, which means that repressors bind DNA most tightly at low salt concentrations. This raises the question of who gene expression might be regulated in obligate halophiles, which maintain internal salt concentrations of about 5 M. As a model system we have investigated the phage phi H, which infects the archaebacterium Halobacterium halobium. Previous genetic data and transcriptional mapping had suggested a region of the phage genome where a repressor might bind. A modified electrophoretic mobility shift assay was used to identify an activity, present only in lysogens, that specifically binds this region. Methylation interference and DNA sequencing were used to identify four similar binding sites, which are arranged so that two copies of a dimer might bind on one face of the DNA helix. Binding of a protein at these sites could block RNA polymerase from initiating a transcript found only during lytic growth. A nearby divergent promoter produces a lysogen-specific transcript, T6, which encodes a member of the helix-turn-helix family of DNA-binding proteins. By expressing the gene in Escherichia coli, we confirmed that T6 specifies the DNA binding activity detected biochemically. The data show that the basic DNA-binding motif of repressors can be adapted even for the unfavorable conditions of high salt concentration.
Ken
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Department of Molecular Biology, Vanderbilt University, Nashville, Tennessee 37235.
Hackett
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D000595:Amino Acid Sequence; D001435:Bacteriophages / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D003090:Coliphages / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005814:Genes, Viral; D006217:Halobacterium / Q000235:genetics*; D008242:Lysogeny; D008745:Methylation; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D004268:DNA-Binding Proteins; D012097:Repressor Proteins