Revisiting Allostery in the Lac Repressor Protein

Abstract

The lac repressor protein (lac) is an allosterically regulated transcription factor which controls expression of the lac operon in bacteria. Binding of a small molecule inducer to a site 40A away from the DNA-binding domain relieves repression through what is thought to be local unfolding of the hinge helix. Despite decades of characterization, our understanding of this allosteric transition remains incomplete_mostly inferred from partial crystal structures. In principle, high-resolution solution NMR could provide detailed structural and dynamical information unobtainable by crystallography. However, due to lac's high molecular weight (70 kDa free, 85 kDa operator-bound), low solubility, and transient stability, such studies have been limited to the non-allosteric, isolated DNA-binding domain. We present, for the first time, a high-resolution solution NMR study of intact wild-type lac in all of its relevant functional states, including the poorly characterized lac-inducer-operator ternary complex. Non-uniform sampling was employed extensively in order to obtain backbone resonance assignments. Our initial results suggest that induction may not proceed simply through unfolding of the hinge helix but rather involves a more complex interplay between changes in structure and dynamics. Research supported by NIH GM102447.