Identification of three residues in the basic regions of the bZIP proteins GCN4, C/EBP and TAF-1 that are involved in specific DNA binding.
The bZIP regions of the eukaryotic transcription factors GCN4 and C/EBP have similar protein sequences but they recognize different DNA sequences. In order to understand their specificity, a vector was constructed which permits overexpression in Escherichia coli of those domains of GCN4 that are necessary and sufficient for specific DNA binding i.e. the basic region and the leucine zipper. Specific DNA binding was monitored with gel shift experiments. The residues of the basic region of GCN4 were systematically replaced by those of C/EBP to transform GCN4 into C/EBP with respect to DNA binding. Residues -17, -16 and -14 were found to be responsible for switching GCN4 to C/EBP binding specificity (we define as residue +1 the first leucine of the first leucine heptad repeat of GCN4). We broadened the specificity of GCN4 to TAF-1 by replacing residues -15 and -17 and we changed the specificity of C/EBP to TAF-1 by swapping residue -17 of a particular hybrid. Thus residues positioned from -14 to -17 of the basic region play a key role in recognizing specific DNA sequences.
Suckow
M
M
Institut für Genetik, Universität zu Köln, Germany.
von Wilcken-Bergmann
B
B
Müller-Hill
B
B
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins; D002868:Chromosomal Proteins, Non-Histone; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli; D005656:Fungal Proteins / Q000378:metabolism*; D051790:G-Box Binding Factors; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D014157:Transcription Factors / Q000378:metabolism
D000596:Amino Acids; D050976:Basic-Leucine Zipper Transcription Factors; D022762:CCAAT-Enhancer-Binding Proteins; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D051790:G-Box Binding Factors; D009687:Nuclear Proteins; D010940:Plant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D004247:DNA; D011494:Protein Kinases