Oligomerization of the EK18 mutant of the trp repressor of Escherichia coli as observed by NMR spectroscopy

The regulation of the trp repressor system of Escherichia coli is frequentl y modeled by a single equilibrium, that between the aporepressor (TR) and t he corepressor, L-tryptophan (Trp), at their intracellular concentrations. The actual mechanism, which is much more complex and more finely tuned, inv olves multiple equilibria: TR and Trp association, TR oligomerization, spec ific and nonspecific binding of various states of TR to DNA, and interactio ns between these various species and ions. TR in isolation exists primarily as a homodimer, but the state of oligomerization increases as the TR conce ntration goes up and/or the salt concentration goes down, leading to specie s with lower affinity for DNA. We have used multinuclear, multidimensional NMR spectroscopy to investigate structural changes that accompany the oligo merization of TR, For these investigations, the superrepressor mutant EK18 (TR with Glu 18 replaced by Lys) was chosen because it exhibits less severe oligomerization at higher protein concentration than other known variants; this made it possible to study the dimer to tetramer oligomerization step by NMR. The NMR results suggest that the interaction between TR dimers is s tructurally linked to folding of the DNA binding domain and that it likely involves direct contacts between the C-terminal residues of the C-helix of one dimer with the next dimer, This implies that oligomerization can compet e with DNA binding and thus serves as a factor in the fine-tuning of gene e xpression. (C) 1999 Academic Press.