Metal Messengers: Communication in the Bacterial World through Transition-Metal-Sensing Two-Component Systems.

Bacteria survive in highly dynamic and complex environments due, in part, to the presence of systems that allow the rapid control of gene expression in the presence of changing environmental stimuli. The crosstalk between intra- and extracellular bacterial environments is often facilitated by two-component signal transduction systems that are typically composed of a transmembrane histidine kinase and a cytosolic response regulator. Sensor histidine kinases and response regulators work in tandem with their modular domains containing highly conserved structural features to control a diverse array of genes that respond to changing environments. Bacterial two-component systems are widespread and play crucial roles in many important processes, such as motility, virulence, chemotaxis, and even transition metal homeostasis. Transition metals are essential for normal prokaryotic physiological processes, and the presence of these metal ions may also influence pathogenic virulence if their levels are appropriately controlled. To do so, bacteria use transition-metal-sensing two-component systems that bind and respond to rapid fluctuations in extracytosolic concentrations of transition metals. This perspective summarizes the structural and metal-binding features of bacterial transition-metal-sensing two-component systems and places a special emphasis on understanding how these systems are used by pathogens to establish infection in host cells and how these systems may be targeted for future therapeutic developments.

A de novo binuclear zinc enzyme with DNA cleavage activity.

Metallohydrolases are broadly used throughout biology, often to catalyze the degradation of macromolecules such as DNA and proteins. Many of these enzymes function with zinc in their active site, and an important subset of these enzymes utilize a binuclear zinc active site. Mimics of these enzymes have been developed, some of which catalyze the digestion of DNA. However, the majority of the mimics that utilize zinc are small molecules, and most are mononuclear. Herein, we report DNA cleavage activity by the de novo designed Due Ferri single-chain (DFsc) protein containing a binuclear zinc active site. This binuclear zinc-protein complex is able to digest plasmid DNA at rates up to 50 ng/h, and these cleavage rates are affected by changes to amino acid residues near the zinc-binding site. These results indicate that the DFsc scaffold is a good model system to carry out careful structure-function relationship studies to understand key structural features that influence reactivity in natural binuclear zinc hydrolases, as it is the first report of a binuclear model system in a protein scaffold.

DNA Cleavage by a De Novo Designed Protein-Titanium Complex.

Titanium is one of the most abundant elements on Earth but is commonly thought to have no role in biology because of its propensity to hydrolyze. Nature stabilizes hard Lewis acidic metals from hydrolysis using a variety of mechanisms, providing inspiration for how titanium can be stabilized using biological ligands. The well-characterized Due Ferri single-chain (DFsc) de novo designed protein was developed to bind and stabilize iron and provides a binding site with hard Lewis basic residues able to bind two metal ions. We demonstrate that the DFsc scaffold stably binds 2 equiv of titanium and protects them from unwanted hydrolysis. The Ti4+-DFsc protein complex was tested for its ability to hydrolytically cleave DNA, where it was seen to linearize plasmid DNA in an overnight reaction. Ti4+-DFsc is thus the first example of a functional, soluble titanium-protein complex.