Function-guided proximity mapping unveils electrophilic-metabolite sensing by proteins not present in their canonical locales.

Enzyme-assisted posttranslational modifications (PTMs) constitute a major means of signaling across different cellular compartments. However, how nonenzymatic PTMs-despite their direct relevance to covalent drug development-impinge on cross-compartment signaling remains inaccessible as current target-identification (target-ID) technologies offer limited spatiotemporal resolution, and proximity mapping tools are also not guided by specific, biologically-relevant, ligand chemotypes. Here we establish a quantitative and direct profiling platform (Localis-rex) that ranks responsivity of compartmentalized subproteomes to nonenzymatic PTMs.

Luciferase Controlled Protein Interactions.

Protein trafficking and protein-protein interactions (PPIs) are central to regulatory processes in cells. Induced dimerization systems have been developed to control PPIs and regulate protein trafficking (localization) or interactions. Chemically induced dimerization (CID) has proven to be a robust approach to control protein interactions and localization.

Smartphone DNA or RNA Sensing Using Semisynthetic Luciferase-Based Logic Device.

Detection of specific oligonucleotide sequences is central to numerous applications, and technologies amenable to point-of-care diagnostics or end users are needed. Here, we report a technology making use of a bioluminescent readout and smartphone quantification. The sensor is a semisynthetic luciferase (H-Luc-PNA conjugate) that is turned on by a strand-displacement reaction.

Coupling of DNA Circuit and Templated Reactions for Quadratic Amplification and Release of Functional Molecules.

DNA-based circuitry empowers logic gated operations and amplifications but is restricted to nucleic acid output. Templated reactions enable the translation of nucleic acid cues into diverse small-molecule outputs but are more limited in their amplification. Herein, we demonstrate the coupling of a DNA circuit to templated reactions in order to achieve high levels of amplification in the output of small molecules, in response to nucleic acid input.

Luciferase-Induced Photouncaging: Bioluminolysis.

Bioluminescence resonance energy transfer (BRET) has been widely used for studying dynamic processes in biological systems such as protein-protein interactions and other signaling events. Aside from acting as a reporter, BRET can also turn on functions in living systems. Herein, we report the application of BRET to performing a biorthogonal reaction in living cells; namely, releasing functional molecules through energy transfer to a coumarin molecule, a process termed bioluminolysis.

Lysosome-targeted photoactivation reveals local sphingosine metabolism signatures.

Lipids are essential components of eukaryotic cell membranes and play crucial roles in cellular signaling and metabolism. While increasing evidence shows that the activities of lipids are dependent upon subcellular localization, tools to study local lipid metabolism and signaling are limited. Herein, we report an approach that enabled us to selectively deliver photo-caged lipids into lysosomes and thereafter to quickly release the lipid molecules by illumination.

Accelerating Turnover Frequency in Nucleic Acid Templated Reactions.

Nucleic acid templated reactions have attracted attention as an important technology to sense oligonucleotides and to translate nucleic acid-based instructions into diverse outputs. Great progress has been made in accelerating the reaction in order to improve signal amplification, reaching rates where substrate turnover rather than chemical reaction is rate limiting. Herein we explore the utility of architectures inspired by three-way junction that yield a cleavage of a strand thus accelerating substrate turnover.

Critical Analysis of Rate Constants and Turnover Frequency in Nucleic Acid-Templated Reactions: Reaching Terminal Velocity.

Nucleic acid-templated reactions have attracted significant attention for nucleic acid sensing and imaging. The level of signal amplification obtained from templated reactions is a function of the template turnover, wherein the template acts as the catalyst. Herein, we report the application of a pyridinium linker that immolates upon photocatalytic reduction with a ruthenium complex to yield the fastest nucleic acid templated reaction reported to date.

Pd-catalyzed difluoromethylation of vinyl bromides, triflates, tosylates, and nonaflates.

Pd-catalyzed difluoromethylation of di-, tri- or tetra-substituted vinyl bromides, triflates, tosylates and nonaflates under mild conditions is described. The reaction tolerates a wide range of functional groups, such as bromide, chloride, fluoride, ester, amine, nitrile, and protected carbonyl, thus providing a general route for the preparation of difluoromethylated alkenes.