The DNA binding specificity of the basic region of the yeast transcriptional activator GCN4 can be changed by substitution of a single amino acid.
The X-ray structure of a GCN4 DNA complex (1) shows, that specific DNA binding of the GCN4 basic region is mediated by a complicated network of base pair and DNA backbone contacts. According to the X-ray structure, alanine -14 of the basic region of GCN4 (we define the first leucine of the leucine zipper as +1) makes a hydrophobic contact to the methyl group of the thymine next to the center of the GCN4 binding site 5' ATGACTCAT 3'. We tested the DNA binding properties of the nineteen derivatives of GCN4, which carry all possible amino acids in position -14 of the basic region. Substitution of alanine -14 of GCN4 by either asparagine or cysteine changes the DNA binding specificity. Serine in this position broadens the specificity for position 1 of the target, whereas other amino acids either retain or decrease GCN4 specificity.
Institut für Genetik, Universität zu Köln, Germany.
M
M
Suckow
B
B
von Wilcken-Bergmann
B
B
Müller-Hill
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D001483:Base Sequence; D004247:DNA / Q000138:chemical synthesis / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D014961:X-Ray Diffraction
D000596:Amino Acids; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D011494:Protein Kinases