NMR Studies of lac Operator and lac Repressor
Résumé
For many years, and even now, the lac operon of Escherichia coli has been a model for gene regulation. The lac repressor is a tetrameric protein that binds strongly the lac operator and the affinity of which is modulated by an inducer molecule. The lac repressor–lac operator complex was the first nucleic acid–protein complex studied with success by NMR. The NMR studies began in 1973 and gave publications until now. 1H, 13C, 15N, 19F, 31P NMR and photo-CIDNP studies increased progressively the level of understanding of the system during these decades. At first, it appeared to be possible to duplicate the basic lac repressor–lac operator interaction by using a smaller protein: the lac repressor N-terminal fragment. In 1987, the Kaptein's group reported the first model of interaction where the motif helix-turn-helix plays a central role in the recognition process of the large groove of the operator by the protein. Later, heteronuclear NMR studies refined the NMR solution structure of both free protein and complexed protein. 13C and 15N relaxation rate measurements showed dynamic processes in both nucleic acid and protein moieties during the complexation. By studying a larger headpiece (1-62) Kaptein's group showed the fundamental role played by the hinge helix, which interacts, in the minor groove of the DNA. The protection factor of a dimeric lac repressor headpiece mutant, which has a high affinity for DNA, was measured and used to qualify the folding of this protein during the complexation. The role played by the asymmetry of the lac operator was studied and explained. NMR studies allowed proposing a scheme to explain the loss in affinity of the lac repressor for its operator when binding to an inducer molecule. Comparison of these data with crystal structures showed that, until now, the more detailed view of the lac repressor–lac operator interaction comes from NMR studies. The NMR solution structures and the dynamic processes revealed by relaxation are in agreement with all the genetic and biochemical data.