PLoS Biol. doi:10.1371/journal.pbio.1001987

Credit: PLoS BIOLOGY

Bacteria have several families of metal-binding proteins that enable them to survive when the extracellular concentrations of metal ions are very low or extremely high. One such protein—the zinc uptake regulator (Zur) protein—is a transcription factor that maintains zinc homeostasis in Escherichia coli by regulating the expression of several proteins, including zinc transporter proteins, a ribosomal protein and a periplasmic zinc-trafficking protein. Gilston et al. have now solved the X-ray crystal structure of two Zur dimers bound to a DNA duplex derived from the znuABC operator ((Zur2)2–PznuABC). The authors identified two zinc-binding sites in each of the Zur monomers and determined that mutations that eliminated metal binding abolished Zur-mediated transcription in vivo. Native gel shift experiments were used to show that the binding of the two Zur dimers to the znuABC promoter was highly cooperative, and the authors determined that a pair of salt bridges that link the Zur dimers mediates the observed cooperativity. Examination of the protein–DNA interface in the (Zur2)2–PznuABC structure and the sequences of two other Zur-binding promoters yielded a putative Zur recognition sequence. This DNA sequence was used to identify a previously unknown Zur-binding promoter in E. coli, upstream of the periplasmic lysozyme inhibitor pliG gene. Comparison of the binding affinities for this transcription factor and the four known Zur-binding promoters indicated that there is a ∼20,000-fold difference in affinity between the strongest and weakest Zur–DNA interaction. These in vitro binding affinities nicely correlate with the levels of repression observed in vivo, as the promoters with tightest Zur binding interactions are the most strongly repressed by Zur in cells (and vice versa). Additional work is needed to determine whether the in vitro thermodynamic properties of other metal- and DNA-binding proteins correlate with their behavior in vivo.