title: Interaction of wild-type and truncated forms of transcription factor IIIA from Saccharomyces cerevisiae with the 5 S RNA gene.
abstract: Transcription factor (TF) IIIA, which contains nine zinc finger motifs, binds to the internal control region of the 5S RNA gene as the first step in the assembly of a multifactor complex that promotes accurate initiation of transcription by RNA polymerase III. We have monitored the interaction of wild-type and truncated forms of yeast TFIIIA with the 5 S RNA gene. The DNase I footprints obtained with full-length TFIIIA and a polypeptide containing the amino-terminal five zinc fingers (TF5) were indistinguishable, extending from nucleotides +64 to +99 of the 5 S RNA gene. This suggests that fingers 6 through 9 of yeast TFIIIA are not in tight association with DNA. The DNase I footprint obtained with a polypeptide containing the amino-terminal four zinc fingers (TF4) was 14 base pairs shorter than that of TF5, extending from nucleotides +78 to +99 on the nontranscribed strand and from nucleotides +79 to +98 on the transcribed strand of the 5 S RNA gene. Protection provided by a polypeptide containing the first three zinc fingers (TF3) was similar to that provided by TF4, with the exception that protection on the nontranscribed strand ended at nucleotide +80, rather than nucleotide +78. Methylation protection analysis indicated that finger 5 makes major groove contacts with guanines +73 and +74. The amino-terminal four zinc fingers make contacts that span the internal control region, which extends from nucleotides +81 to +94 of the 5 S RNA gene, with finger 4 appearing to contact guanine +82. Measurements of the apparent Kd values of the TFIIIA.DNA complexes indicated that the amino-terminal three zinc fingers of TFIIIA have a binding energy that is similar to that of the full-length protein.
authors.affiliation: Department of Biochemistry, University of Toronto, Ontario, Canada.
authors.forename: O
authors.initials: O
authors.lastname: Rowland
authors.forename: J
authors.initials: J
authors.lastname: Segall
mesh_terms: D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000494:pharmacology; D008745:Methylation; D012341:RNA, Ribosomal, 5S / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000378:metabolism*; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; D012341:RNA, Ribosomal, 5S; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors; D004247:DNA; D003850:Deoxyribonuclease I