Structure and dynamics of the tetrameric Mnt repressor and a model for its DNA complex

The tetrameric Mnt repressor of bacteriophage P22 consists of two dimeric DNA-binding domains and a tetramerization domain. The NOE and chemical shift data demonstrate that the structures of the domains in the wild-type repressor protein are similar to those of the separate domains, the three-dimensional structures of which have been determined previously. N-15 relaxation measurements show that the linker that connects the anti-parallel four-helix bundle with the two beta -sheet DNA-binding dimers is highly flexible. No evidence was found for interactions between the distinct modules. The N-15 relaxation properties of the two domains differ substantially, confirming their structural independence. A model in which one two-stranded coiled coil of the four-helix bundle is attached to one N-terminal dimer is most consistent with the biochemical data and N-15 relaxation data. For the Mnt-DNA complex this geometry fits with a model in which the two beta -sheet DNA-binding domains are bound at two successive major grooves of the Mnt operator and the tetramerization domain is packed between these two DNA-bound dimers. In such a model the two-fold symmetry axis of the four-helix bundle coincides with that of the operator sequence and the two bound dimers. Bending of the Mnt operator of approximately 30 degrees upon binding of the tetramer. as measured by gel-shift assays, is in agreement with this model of the Mnt-DNA complex.