Purification and characterization of the Ner repressor of bacteriophage Mu.
The Ner protein of bacteriophage Mu acts as a lambda cro-like negative regulator of the phage's early (transposase) operon. Using the band retardation assay to monitor ner-operator-specific DNA-binding activity, the 8 kDa Ner protein was purified to homogeneity. DNase I footprinting revealed that the purified protein bound and protected a specific DNA operator that contains two 12 bp sites with the consensus sequence 5'-ANPyTAPuCTAAGT-3', separated by a 6 bp spacer region. Moreover, regions corresponding to a turn of the DNA helix flanking these 12 bp repeats are also protected by Ner. Unlike the functionally similar lambda cro protein, gel filtration experiments show that the native molecular mass of Mu Ner to be approx. 8 kDa. These results, plus the pattern of DNase I protection, suggest that the protein may bind as a monomer to each of its specific DNA substrates.
Kukolj
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Department of Microbiology and Immunology, McGill University, Montreal, Canada.
Tolias
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D001483:Base Sequence; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003090:Coliphages / Q000235:genetics*; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000302:isolation & purification*; D003850:Deoxyribonuclease I; D017886:Durapatite; D004926:Escherichia coli / Q000235:genetics*; D006882:Hydroxyapatites; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009876:Operon; D012097:Repressor Proteins / Q000302:isolation & purification* / Q000378:metabolism; D014157:Transcription Factors / Q000302:isolation & purification*; D014764:Viral Proteins / Q000302:isolation & purification* / Q000378:metabolism; D054334:Viral Regulatory and Accessory Proteins
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D006882:Hydroxyapatites; D012097:Repressor Proteins; D014157:Transcription Factors; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D017886:Durapatite; D003850:Deoxyribonuclease I