IMPORTANCE OF MINOR-GROOVE BINDING ZINC FINGERS WITHIN THE TRANSCRIPTION FACTOR IIIA-DNA COMPLEX

The gene-specific transcription factor IIIA (TFIIIA) binds to the inte rnal promoter element of the 5 S rRNA gene through nine zinc fingers w hich 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-imid azole polyamides are minor groove binding Ligands that recognize prede termined DNA sequences with affinity and specificity comparable to nat ural DNA-binding proteins. We have examined the DNA binding activity o f nine finger TFIIIA and shorter recombinant analogs in the presence o f polyamides that bind six base-pair sequences (K-d = 0.03 to 1.7 nM) in the minor groove of the binding site for zinc finger 4. DNase I foo tprint titrations demonstrate that the polyamides and a recombinant pr otein containing the three amino-terminal zinc fingers of TFIIIA (zf1- 3) co-occupy the TFIIIA binding site, in agreement with the known loca tion of zf1-3 in the major groove. In contrast, the polyamides block t he specific interaction of TFIIIA or zf1-4 with the 5S RNA gene, suppo rting a model for minor groove occupancy by zinc finger 4. Minor groov e binding polyamides targeted to specific DNA sequences may provide a novel chemical approach to probing multidomain protein-DNA interaction s. (C) 1997 Academic Press Limited.