DNA-directed oligomerization of the monomeric Ner repressor from the Mu-like bacteriophage D108.
We have purified the 8.6 kd ner gene product (a lambda Cro-like protein which negatively regulates transcription from two divergent and overlapping promoters) from the Mu-like transposable bacteriophage D108. Chemical and enzymatic protection experiments show the D108 ner-operator to contain two perfect 11 bp (5'-CCG-TGAGCTAC-3') inverted repeats separated by an 8 bp AT-rich region. Ner makes base-specific contacts in the major groove spanning the 11 bp repeats and also interacts with regions flanking these sites such that its operator comprises five turns of the DNA helix. Furthermore, gel filtration chromatography and dimethyl suberimidate crosslinking experiments indicate that D108 Ner (at concentrations exceeding 5 microM) is a monomer in solution, yet crosslinks as a dimer when bound to its operator site. As a small (73 amino acids) monomeric protein, Ner does not display strong homology with any known DNA-binding proteins. By virtue of the interactions with its operator it appears to bind DNA in a markedly different manner from other known prokaryotic repressors thus adding to the growing catalog of protein motifs used for specific binding to DNA.
Kukolj
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Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada.
Tolias
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PP
Autexier
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DuBow
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D010583:Bacteriophage mu / Q000235:genetics*; D001483:Base Sequence; D002850:Chromatography, Gel; D004247:DNA / Q000378:metabolism*; D046911:Macromolecular Substances; D008745:Methylation; D012091:Repetitive Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000235:genetics* / Q000302:isolation & purification; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics* / Q000302:isolation & purification; D054334:Viral Regulatory and Accessory Proteins
D046911:Macromolecular Substances; D012097:Repressor Proteins; D014157:Transcription Factors; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D004247:DNA