Only two of the five zinc fingers of the eukaryotic transcriptional repressor PRDI-BF1 are required for sequence-specific DNA binding.
The eukaryotic transcriptional repressor PRDI-BF1 contains five zinc fingers of the C2H2 type, and the protein binds specifically to PRDI, a 14-bp regulatory element of the beta interferon gene promoter. We have investigated the amino acid sequence requirements for specific binding to PRDI and found that the five zinc fingers and a short stretch of amino acids N terminal to the first finger are necessary and sufficient for PRDI-specific binding. The contribution of individual zinc fingers to DNA binding was investigated by inserting them in various combinations into another zinc finger-containing DNA-binding protein whose own fingers had been removed. We found that insertion of PRDI-BF1 zinc fingers 1 and 2 confer PRDI-binding activity on the recipient protein. In contrast, the insertion of PRDI-BF1 zinc fingers 2 through 5, the insertion of zinc finger 1 or 2 alone, and the insertion of zinc fingers 1 and 2 in reverse order did not confer PRDI-binding activity. We conclude that the first two PRDI-BF1 zinc fingers together are sufficient for the sequence-specific recognition of PRDI.
Keller
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Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, Massachusetts 02138.
Maniatis
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002872:Chromosome Deletion; D003850:Deoxyribonuclease I; D005822:Genetic Vectors; D016899:Interferon-beta / Q000235:genetics; D007438:Introns; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes; D011401:Promoter Regions, Genetic; D012326:RNA Splicing; D012045:Regulatory Sequences, Nucleic Acid; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism*; D015183:Restriction Mapping; D014157:Transcription Factors; D014158:Transcription, Genetic; D016335:Zinc Fingers / Q000235:genetics*
D015345:Oligonucleotide Probes; D012097:Repressor Proteins; D014157:Transcription Factors; D016899:Interferon-beta; D003850:Deoxyribonuclease I