Spatiotemporally resolved mapping of extracellular proteomes via in vivo-compatible TyroID

Nature Communications, Journal Year: 2025, Volume and Issue: 16(1)

Published: March 15, 2025

Extracellular proteins play pivotal roles in both intracellular signaling and intercellular communications health disease. While recent advancements proximity labeling (PL) methods, such as peroxidase- photocatalyst-based approaches, have facilitated the resolution of extracellular proteomes, their vivo compatibility remains limited. Here, we report TyroID, an vivo-compatible PL method for unbiased mapping with high spatiotemporal resolution. TyroID employs plant- bacteria-derived tyrosinases to produce reactive o-quinone intermediates, enabling multiple residues on endogenous bioorthogonal handles, thereby allowing identification via chemical proteomics. We validate TyroID's specificity by proteomes HER2-neighboring using affibody-directed recombinant tyrosinases. Demonstrating its superiority over other enables including HER2-proximal tumor xenografts, quantifying turnover plasma hippocampal-specific live mouse brains. emerges a potent tool investigating protein localization molecular interactions within living organisms. is that maps through efficiently labels study interactions.

Language: Английский

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Advances in spatial proteomics: Mapping proteome architecture from protein complexes to subcellular localizations

Cell chemical biology, Journal Year: 2024, Volume and Issue: 31(9), P. 1665 - 1687

Published: Sept. 1, 2024

Language: Английский

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Enzymatic control of intermolecular interactions for generating synthetic nanoarchitectures in cellular environment

Science and Technology of Advanced Materials, Journal Year: 2024, Volume and Issue: 25(1)

Published: June 28, 2024

Nanoarchitectonics, as a technology to arrange nano-sized structural units such molecules in desired configuration, requires nano-organization, which usually relies on intermolecular interactions. This review briefly introduces the development of using enzymatic reactions control interactions for generating artificial nanoarchitectures cellular environment. We begin discussion with early examples and uniqueness enzymatically controlled self-assembly. Then, we describe intracellular nanostructures their relevant applications. Subsequently, discuss cases forming cell surface via reactions. Following that, highlight use creating intercellular nanostructures. Finally, provide summary outlook promises future direction this strategy. Our aim is give an updated introduction reaction regulating interactions, phenomenon ubiquitous biology but relatively less explored by chemists materials scientists. goal stimulate new developments simple versatile approach addressing societal needs.

Language: Английский

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Progress toward a comprehensive brain protein interactome

Biochemical Society Transactions, Journal Year: 2025, Volume and Issue: 53(01)

Published: Feb. 12, 2025

Protein–protein interactions (PPIs) in the brain play critical roles across all aspects of central nervous system, from synaptic transmission, glial development, myelination, to cell-to-cell communication, and more. Understanding these is crucial for deciphering neurological mechanisms underlying biochemical machinery affected disorders. Recently, advances proteomics techniques have significantly enhanced our ability study among proteins expressed brain. Here, we review some high-throughput studies characterizing PPIs, using affinity purification, proximity labeling, co-fractionation, chemical cross-linking mass spectrometry methods, as well yeast two-hybrid assays. We present current state field, discuss challenges, highlight promising future directions.

Language: Английский

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Profiling Proteins Involved in Peroxynitrite Homeostasis Using ROS/RNS Conditional Proteomics

Journal of the American Chemical Society, Journal Year: 2025, Volume and Issue: 147(9), P. 7305 - 7316

Published: Feb. 24, 2025

Peroxynitrite (ONOO–), the product of diffusion-controlled reaction superoxide (O2•–) with nitric oxide (NO•), plays a crucial role in oxidative and nitrative stress modulates key physiological processes such as redox signaling. While biological ONOO– is conventionally analyzed using 3-nitrotyrosine antibodies fluorescent sensors, probes lack specificity sensitivity, making high-throughput comprehensive profiling ONOO–-associated proteins challenging. In this study, we used conditional proteomics approach to investigate homeostasis by identifying its protein neighbors cells. We developed Peroxynitrite-responsive Labeling reagents (Porp-L) and, for first time, discovered 2,6-dichlorophenol an ideal moiety that can be selectively rapidly activated labeling proximal proteins. The Porp-L generated several short-lived reactive intermediates modify Tyr, His, Lys residues on surface. have demonstrated Porp-L-based immune-stimulated macrophages, which indeed identified known involved generation modification revealed endoplasmic reticulum (ER) hot spot. Moreover, previously unknown Ero1a, ER-resident protein, formation ONOO–. Overall, represent promising research tool advancing our understanding roles

Language: Английский

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Tyrosinase-Activated MRI and PET Probes for Selective Melanoma Imaging

ACS Sensors, Journal Year: 2025, Volume and Issue: unknown

Published: March 25, 2025

Melanoma is one of the most aggressive forms skin cancer. Accurate and early diagnosis melanoma crucial for improving patient outcomes. This study develops two TYR-activatable molecular probes, Mn-TyrEDTA Al-18F-TyrEDTA, selective detection in vivo. In vitro studies reveal that exhibits TYR activity-dependent relaxivity enhancement, undergoing TYR-mediated oxidative polymerization, resulting formation paramagnetic oligomers. UV-vis analysis supports this mechanism, showing time- concentration-dependent increases broad band absorbance region, specifically around 475 nm, due to o-quinone intermediates melanin-like HPLC further confirmed presence polar oligomeric products solutions incubated with TYR/O2. MRI demonstrate Mn-TyrEDTA's retention signal enhancement TYR-expressing tissues. Furthermore, PET imaging Al-18F-TyrEDTA conducted using a dual-xenograft mouse model reveals significantly higher uptake compared TYR-negative tumors. could be attributed proximity labeling where highly reactive quinones form covalent bonds nearby tumor proteins. summary, our findings establish as promising offering novel strategy prognosis melanoma.

Language: Английский

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Engineered Proteins and Chemical Tools to Probe the Cell Surface Proteome

Chemical Reviews, Journal Year: 2025, Volume and Issue: unknown

Published: April 3, 2025

The cell surface proteome, or surfaceome, is the hub for cells to interact and communicate with outside world. Many disease-associated changes are hard-wired within yet approved drugs target less than 50 proteins. In past decade, proteomics community has made significant strides in developing new technologies tailored studying surfaceome all its complexity. this review, we first dive into unique characteristics functions of emphasizing necessity specialized labeling, enrichment, proteomic approaches. An overview surfaceomics methods provided, detailing techniques measure protein expression how leads novel discovery. Next, highlight advances proximity labeling (PLP), showcasing various enzymatic photoaffinity can map protein-protein interactions membrane complexes on surface. We then review role extracellular post-translational modifications, focusing glycosylation, proteolytic remodeling, secretome. Finally, discuss identifying tumor-specific peptide MHC they have shaped therapeutic development. This emerging field neo-protein epitopes constantly evolving, where targets identified at proteome level encompass defined PTMs, complexes, dysregulated cellular tissue locations. Given functional importance biology therapy, view as a critical piece quest neo-epitope

Language: Английский

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Proximitomics by Reactive Species

ACS Central Science, Journal Year: 2024, Volume and Issue: 10(6), P. 1135 - 1147

Published: June 12, 2024

The proximitome is defined as the entire collection of biomolecules spatially in proximity a biomolecule interest. More broadly, concept can be extended to totality cells proximal specific cell type. Since spatial organization and essential for almost all biological processes, proximitomics has recently emerged an active area scientific research. One growing strategies leverages reactive species─which are generated situ confined, chemically tag capture systematic analysis. In this Outlook, we summarize different types species that have been exploited discuss their pros cons applications. addition, current challenges future directions exciting field.

Language: Английский

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Target protein identification in live cells and organisms with a non-diffusive proximity tagging system

Published: Oct. 29, 2024

Identifying target proteins for bioactive molecules is essential understanding their mechanisms, developing improved derivatives, and minimizing off-target effects. Despite advances in identification (target-ID) technologies, significant challenges remain, impeding drug development. Most target-ID methods use cell lysates, but maintaining an intact cellular context vital capturing specific drug-protein interactions, such as those with transient protein complexes membrane-associated proteins. To address these limitations, we developed POST-IT (Pup-On-target Small molecule Target Identification Technology), a non-diffusive proximity tagging system live cells, orthogonal to the eukaryotic system. utilizes engineered fusion of proteasomal accessory factor A (PafA) HaloTag transfer Pup proximal upon directly binding small molecule. After optimization eliminate self-pupylation polypupylation, minimize depupylation, optimize chemical linkers, successfully identified known targets discovered new binder, SEPHS2, dasatinib, VPS37C hydroxychloroquine, enhancing our drugs’ mechanisms action. Furthermore, demonstrated application zebrafish embryos, highlighting its potential broad biological research

Language: Английский

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Target protein identification in live cells and organisms with a non-diffusive proximity tagging system

eLife, Journal Year: 2024, Volume and Issue: 13

Published: Oct. 29, 2024

Identifying target proteins for bioactive molecules is essential understanding their mechanisms, developing improved derivatives, and minimizing off-target effects. Despite advances in identification (target-ID) technologies, significant challenges remain, impeding drug development. Most target-ID methods use cell lysates, but maintaining an intact cellular context vital capturing specific drug–protein interactions, such as those with transient protein complexes membrane-associated proteins. To address these limitations, we developed POST-IT (Pup-On-target Small molecule Target Identification Technology), a non-diffusive proximity tagging system live cells, orthogonal to the eukaryotic system. utilizes engineered fusion of proteasomal accessory factor A HaloTag transfer Pup proximal upon directly binding small molecule. After optimization eliminate self-pupylation polypupylation, minimize depupylation, optimize chemical linkers, successfully identified known targets discovered new binder, SEPHS2, dasatinib, VPS37C hydroxychloroquine, enhancing our drugs’ mechanisms action. Furthermore, demonstrated application zebrafish embryos, highlighting its potential broad biological research

Language: Английский

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