Nucleic Acids Research, Vol 24, Issue 22 4487-4494, Copyright © 1996 by Oxford University Press
M John, R Leppik, SJ Busch, M Granger-Schnarr and M Schnarr
We constructed plasmids encoding the sequences for the bZip modules of
c-Jun and c-Fos which could then be expressed as soluble proteins in
Escherichia coli. The purified bZip modules were tested for their binding
capacities of synthetic oligonucleotides containing either TRE or CRE
recognition sites in electrophoretic mobility shift assays and circular
dichroism (CD). Electrophoretic mobility shift assays showed that bZip Jun
homodimers and bZip Jun/Fos heterodimers bind a collagenase-like TRE
(CTGACTCAT) with dissociation constants of respectively 1.4 x 10(-7) M and
5 x 10(-8) M. As reported earlier [Patel et al. (1990) Nature 347,
572-575], DNA binding induces a marked change of the protein structure.
However, we found that the DNA also undergoes a conformational change. This
is most clearly seen with small oligonucleotides of 13 or 14 bp harboring
respectively a TRE (TGACTCA) or a CRE (TGACGTCA) sequence. In this case,
the positive DNA CD signal at 280 nm increases almost two-fold with a
concomitant blue-shift of 3- 4 nm. Within experimental error the same
spectral changes are observed for TRE and CRE containing DNA fragments. The
spectral changes observed with a non-specific DNA fragment are weaker and
the signal of free DNA is recovered upon addition of much smaller salt
concentrations than required for a specific DNA fragment. Surprisingly the
spectral changes induced by Jun/Jun homodimers are not identical to those
induced by Jun/Fos heterodimers. However, in both cases the increase of the
positive CD band and the concomitant blue shift would be compatible with a
B to A-transition of part of the binding site or a DNA conformation
intermediate between the canonical A and B structures.
ARTICLES
DNA binding of Jun and Fos bZip domains: homodimers and heterodimers induce a DNA conformational change in solution
Institut de Biologie Moleculaire et Cellulaire du CNRS, Strasbourg, France.
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