NMR solution structure of Max in absence of DNA: Insight into the mechanism of
DNA recognition by B-HLH-LZ transcription factors

The NMR solution structure of a mutant of the Max b/HLH/LZ free of DNA: insights into the specific and reversible DNA binding mechanism of dimeric transcription factors.

Max is a member of the b-HLH-LZ (basic region-helix1-loop-helix2-leucine zipper)
family of eukaryotic transcription factors. It is the obligate partner of the
related b-HLH-LZ proteins, c-Myc and Mad1, with which it forms heterodimers on
target DNA. While c-Myc and Mad1 require Max for DNA-binding, Max itself can
form a homodimer that recognizes E-box DNA sequences (CACGTG) in gene promoters
that are targeted by c-Myc. Evidence suggests that this mode of binding by Max
may repress c-Myc transcriptional activity, and this may have applications in
the control of the aberrant activity of c-Myc during certain oncogenic
transformations. To enhance this repressive potential of Max, we sought to
stabilize Max homodimers. We have designed a double mutant (N78V/H81L) located
in the coiled-coil interface of the leucine zipper domain and we demonstrate
that these mutations do indeed increase the stability of the protein. The
mutations also improve the stability of the complex with cognate DNA. Thermal
denaturations monitored by circular dichroism reveal two transitions that are
due to intermediate folding states for both the wild-type and mutant proteins;
this is supported by detailed thermodynamic analyses. A formalism to
characterize the temperature-dependence of the unfolding, including the effect
of intermediates, is presented.

Jean-Francois N, Frederic G, Raymund W, Benoit C, Lavigne P.
Improving the thermodynamic stability of the leucine zipper of max increases
the stability of its b-HLH-LZ:E-box complex.
J Mol Biol. 2003 Mar 7;326(5):1577-95.

Improving the thermodynamic stability of the leucine zipper of max increases the stability of its b-HLH-LZ:E-box complex.

For more information on the construction, please refer to
Jean-Francois, N. et al. J. Mol. Biol. (2003) 326, 1577-1595.