Optical Control of Antibody Activity by Using Photocleavable Bivalent Peptide-DNA Locks.

Antibody-based molecular recognition plays a central role in today's life sciences, ranging from immunoassays to molecular imaging and antibody-based therapeutics. Control over antibody activity by using external triggers such as light could further increase the specificity of antibody-based targeting. Here we present bivalent peptide-DNA ligands containing photocleavable linkers as a noncovalent approach by which to allow photoactivation of antibody activity. Light-triggered cleavage of the 3-amino-3-(2-nitrophenyl)propionic acid peptide linker converted the high-affinity bivalent peptide-DNA lock into weakly binding monovalent ligands, effectively restoring antibody targeting of cell-surface receptors. In this work, a proof of principle was provided with an anti-hemagglutinin antibody, but the molecular design of the lock is generic and applicable to any monoclonal antibody for which an epitope or mimotope of sufficient affinity is available.

Generalizable Protein Biosensors Based on Synthetic Switch Modules.

Allosteric protein switches are key controllers of information and energy processing in living organisms and are desirable engineered control tools in synthetic systems. Here we present a generally applicable strategy for construction of allosteric signaling systems with inputs and outputs of choice. We demonstrate conversion of constitutively active enzymes into peptide-operated synthetic allosteric ON switches by insertion of a calmodulin domain into rationally selected sites. Switches based on EGFP, glucose dehydrogenase, NanoLuciferase, and dehydrofolate reductase required minimal optimization and demonstrated a dynamic response ranging from 1.8-fold in the former case to over 200-fold in the latter case. The peptidic nature of the calmodulin ligand enables incorporation of such synthetic switch modules into higher order sensory architectures. Here, a ligand-mediated increase in proximity of the allosteric switch and the engineered activator peptide modulates biosensor's activity. Created biosensors were used to measure concentrations of clinically relevant drugs and biomarkers in plasma, saliva, and urine with accuracy comparable to that of the currently used clinical diagnostic assays. The approach presented is generalizable as it allows rapid construction of efficient protein switches that convert binding of a broad range of analytes into a biochemical activity of choice enabling construction of artificial signaling and metabolic circuits of potentially unlimited complexity.