Mutational analysis of the PapB transcriptional regulator in Escherichia coli. Regions important for DNA binding and oligomerization.
PapB is a transcriptional regulator in the control of pap operon expression in Escherichia coli. There are PapB homologous proteins encoded by many fimbrial gene systems that are involved in the regulation of fimbriae-adhesin production, and previous studies suggested that PapB binds DNA through minor groove contact. Both deletion and alanine-scanning mutagenesis were used to identify functionally important regions of the PapB protein. Mutations altering Arg61 or Cys65 caused deficiency in DNA binding, indicating that these residues are critical for PapB binding to DNA. Alanine substitutions at positions 35-36, 53-56, and 74-76 resulted in mutants that were impaired in oligomerization. All these amino acid residues are conserved among the PapB homologous proteins, suggesting their importance in the whole family of regulatory proteins. The transcriptional efficiency of all the mutants was clearly reduced as compared with that of wild-type PapB. Taken together, we have localized regions in the PapB protein that are involved in DNA binding and oligomerization, and our results show that both functions are required for its activity as a transcriptional regulator.
Xia
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Y
Department of Microbiology, Umeå University, S-90187 Umeå, Sweden.
Uhlin
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BE
D000595:Amino Acid Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010957:Plasmids / Q000235:genetics; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics*; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; C116245:atpD protein, E coli