title: Importance of minor groove binding zinc fingers within the transcription factor IIIA-DNA complex.
abstract: The gene-specific transcription factor IIIA (TFIIIA) binds to the internal promoter element of the 5 S rRNA gene through nine zinc fingers which make specific DNA contacts. Seven of the nine TFIIIA zinc fingers participate in major groove DNA contacts while two fingers, 4 and 6, have been proposed to bind in or across the minor groove. Pyrrole-imidazole polyamides are minor groove binding ligands that recognize predetermined DNA sequences with affinity and specificity comparable to natural DNA-binding proteins. We have examined the DNA binding activity of nine finger TFIIIA and shorter recombinant analogs in the presence of polyamides that bind six base-pair sequences (Kd = 0.03 to 1.7 nM) in the minor groove of the binding site for zinc finger 4. DNase I footprint titrations demonstrate that the polyamides and a recombinant protein containing the three amino-terminal zinc fingers of TFIIIA (zf1-3) co-occupy the TFIIIA binding site, in agreement with the known location of zf1-3 in the major groove. In contrast, the polyamides block the specific interaction of TFIIIA or zf1-4 with the 5 S RNA gene, supporting a model for minor groove occupancy by zinc finger 4. Minor groove binding polyamides targeted to specific DNA sequences may provide a novel chemical approach to probing multidomain protein-DNA interactions.
authors.affiliation: Department of Molecular Biology, Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: L
authors.initials: L
authors.lastname: Neely
authors.forename: J W
authors.initials: JW
authors.lastname: Trauger
authors.forename: E E
authors.initials: EE
authors.lastname: Baird
authors.forename: P B
authors.initials: PB
authors.lastname: Dervan
authors.forename: J M
authors.initials: JM
authors.lastname: Gottesfeld
mesh_terms: D001665:Binding Sites; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D006147:Guanine / Q000737:chemistry / Q000378:metabolism; D009757:Nylons / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism*; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; D009757:Nylons; D011994:Recombinant Proteins; D035582:Transcription Factor TFIIIA; D014157:Transcription Factors; D006147:Guanine; D004247:DNA; D003850:Deoxyribonuclease I