The role of membrane potential in most intracellular organelles remains unexplored because of the lack of suitable tools. Here, we describe a fluorescent DNA-nanodevice that reports absolute membrane potential and can be targeted to organelles in live cells. This nanodevice, denoted Voltair , bears a voltage-sensitive fluorophore, a reference fluorophore for ratiometry, and acts as an endocytic tracer. Using Voltair we could measure the membrane potential of different organelles in situ in live cells. Voltair can potentially guide the rational design of biocompatible electronics as well as expand our understanding of how membrane potential regulates organelle biology.

The role of membrane potential in most intracellular organelles remains unexplored because of the lack of suitable probes. We describe a DNA-based fluorescent reporter that 20

Methylglyoxal (MGO), a reactive metabolite byproduct of glucose metabolism, is known to form a variety of posttranslational modifications (PTMs) on nucleophilic amino acids. For example, cysteine, the most nucleophilic proteinogenic amino acid, forms reversible hemithioacetal and stable mercaptomethylimidazole adducts with MGO. The high reactivity of cysteine toward MGO and the rate of formation of such modifications provide the opportunity for mechanisms by which proteins and pathways might rapidly sense and respond to alterations in levels of MGO. This indirect measure of alterations in glycolytic flux would thereby allow disparate cellular processes to dynamically respond to changes in nutrient availability and utilization. Here we report the use of quantitative LC−MS/MS-based chemoproteomic profiling approaches with a cysteine-reactive probe to map the proteome-wide landscape of MGO modification of cysteine residues. This approach led to the identification of many sites of potential functional regulation by MGO. We further characterized the role that such modifications have in a catalytic cysteine residue in a key metabolic enzyme and the resulting effects on cellular metabolism.

Functional regulation of cell signaling through dynamic changes in protein activity state as well as spatial organization represent two dynamic, complex, and conserved phenomena in biology. Seemingly separate areas of -omics method development have focused on building tools that can detect and quantify protein activity states, as well as map sub-cellular and intercellular protein organization. Integration of these efforts, through the development of chemical tools and platforms that enable detection and quantification of protein functional states with spatial resolution provide opportunities to better understand heterogeneity in the proteome within cell organelles, multi-cellular tissues, and whole organisms. This review provides an overview of and considerations for major classes of chemical proteomic probes and technologies that enable protein activity mapping from sub-cellular compartments to live animals.