Design of stimulus-responsive two-state hinge proteins

In nature, proteins that switch between two conformations in response to environmental stimuli structurally transduce biochemical information in a manner analogous to how transistors control information flow in computing devices. Designing proteins with two distinct but fully structured conformations is a challenge for protein design as it requires sculpting an energy landscape with two distinct minima. Here we describe the design of “hinge” proteins that populate one designed state in the absence of ligand and a second designed state in the presence of ligand. X-ray crystallography, electron microscopy, double electron-electron resonance spectroscopy, and binding measurements demonstrate that despite the significant structural differences the two states are designed with atomic level accuracy and that the conformational and binding equilibria are closely coupled. Natural proteins often adopt multiple conformational states, thereby changing their activity or binding partners in response to another protein, small molecule, or other stimulus. It has been difficult to engineer such conformational switching between two folded states in human-designed proteins. Praetorius et al . developed a hinge-like protein by simultaneously considering both desired states in the design process. The successful designs exhibited a large shift in conformation upon binding to a target peptide helix, which could be tailored for specificity. The authors characterized the protein structures, binding kinetics, and conformational equilibrium of the designs. This work provides the groundwork for generating protein switches that respond to biological triggers and can produce conformational changes that modulate protein assemblies. —Michael A. Funk A two-state design of protein switches that couple effector binding to a conformational change is discussed.