The physical structure of an activated gene complexes is as yet unknown, although the formation of DNA loops or bends stabilized by protein binding or protein-protein interactions is considered likely. Previously the binding of divalent Zn to the 5s RNA gene from Xenopus laevis was shown to induce a strong bend in the DNA. The magnitude and center position of this bend from a series of DNA fragments with circularly permuted sequences and rotational relaxation times measured by transient electric birefringence have now been determined. The bend is some 50-60 degrees and is centered at about base pair +60 within the gene. This places the bend within the binding domain of the transcription regulatory factor TFIIIA. The rotational relaxation time of the TFIIIA-5s RNA gene complex is only slightly faster than the decay time found for the ZN-DNA complex. The binding of the regulatory factor is most probably stabilizing the same bent DNA conformation as Zn binding. Solution conditions that inhibit the formation of the Zn induced bend in DNA also inhibit the binding of TFIIIA to the gene. This is the first example of a bent DNA - protein complex that is stabilized by the formation of an alternate DNA conformation. The DNA sequence that appears responsible for the bent conformation is GGG. This triplet is prominent in several regulatory DNA binding sequences, most notably the sp1 binding domain of SV40. Further work is underway both to characterize the sequence requirements for the bent conformation, using a series of single base mutations in the TFIIIA binding domain of the 5s RNA gene, and the generality of inducing a bent conformation at this sequence by the binding of transcription regulatory proteins, using the 21 bp sequence from SV40.
