Mutations in the zinc fingers of ADR1 that change the specificity of DNA binding and transactivation.
ADR1 is a yeast transcription factor that contains two zinc fingers of the Cys-2-His-2 (C2H2) class. Mutations that change the specificity of DNA binding of ADR1 to its target site, upstream activation sequence 1 (UAS1), have been identified at three positions in the first zinc finger. Mutations Arg-115 to Gln, His-118 to Thr, and Arg-121 to Asn led to new specificities of DNA binding at adjacent positions 10, 9, and 8 (3'-GAG-5') in UAS1. Arg-115 is at the finger tip, and His-118 and Arg-121 are at positions 3 and 6, respectively, in the alpha helix of finger 1. One double mutant displayed the binding specificity expected from the properties of its constituent new-specificity mutations. Mutations in the second finger that allowed its binding site to be identified through loss-of-contact phenotypes were made. These mutations imply a tail-to-tail orientation of the two ADR1 monomers on their adjacent binding sites. Finger 1 is aligned on UAS1 in an amino-to-carboxyl-terminal orientation along the guanine-rich strand in a 3'-to-5' direction. One of the ADR1 mutants was functional in vivo with both its cognate binding site and wild-type UAS1, but the other two mutants were defective in transactivation despite their ability to bind with high affinity to their cognate binding sites.
Thukral
S K
SK
Department of Biochemistry, University of Washington, Seattle 98195.
Morrison
M L
ML
Young
E T
ET
D000595:Amino Acid Sequence; D001483:Base Sequence; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D016335:Zinc Fingers
C074682:ADR1 protein, S cerevisiae; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D009838:Oligodeoxyribonucleotides; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors