The regulator of nitrate assimilation in ascomycetes is a dimer which binds a nonrepeated, asymmetrical sequence.
The regulation of nitrate assimilation seems to follow the same pattern in all ascomycetes where this process has been studied. We show here by in vitro binding studies and a number of protection and interference techniques that the transcription factor mediating nitrate induction in Aspergillus nidulans, a protein containing a binuclear zinc cluster DNA binding domain, recognizes an asymmetrical sequence of the form CTCC GHGG. We further show that the protein binds to its consensus site as a dimer. We establish the role of the putative dimerization element by its ability to replace the analogous element of the cI protein of phage lambda. Mutagenesis of crucial leucines of the dimerization element affect both the binding ability of the dimer and the conformation of the resulting protein-DNA complex. This is the first case to be described where a dimer recognizes such an asymmetrical nonrepeated sequence, presumably by each monomeric subunit making different contacts with different DNA half-sites.
Strauss
J
J
Institut de Génétique et Microbiologie, Université Paris-Sud, URA D2225, Orsay, France.
Muro-Pastor
M I
MI
Scazzocchio
C
C
D000595:Amino Acid Sequence; D001233:Aspergillus nidulans / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D002874:Chromosome Mapping; D018983:DNA Footprinting; D015342:DNA Probes; D004271:DNA, Fungal / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D019281:Dimerization; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009566:Nitrates / Q000378:metabolism*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid
D015342:DNA Probes; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D009566:Nitrates; D011993:Recombinant Fusion Proteins; C071245:NirA protein, Aspergillus nidulans