Genetically encoded chemical crosslinking of carbohydrate

Li, Shanshan; Wang, Nanxi; Yu, Bingchen; Sun, Wei; Wang, Lei

doi:10.1038/s41557-022-01059-z

Cited by 42 publications

(30 citation statements)

References 52 publications

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“…Due to their enhanced protein solubility and proteolysis resistance, “active site transplant” variants of Methanomethylophilus alvus pyrrolysyl-tRNA synthetase ( Ma -PylRS) are reported to display higher UAA incorporation efficiency than their Methanosarcina mazei ( Mm ) and Methanosarcina barkeri ( Mb ) counterparts. ,, We wondered whether the transplantation of Ma -VtKRS with active site mutations directly from established Mb -VtKRS could further improve VtK incorporation (Figure b). We quantitatively compared the incorporation efficiency and fidelity of Mb -VtKRS and Ma -VtKRS by measuring the relative fluorescence units (RFUs) of cells coexpressing them with the sfGFP-2TAG and sfGFP-149TAG reporter genes in Escherichia coli DH10B, respectively, in the presence or absence of 1 mM VtK.…”

Section: Resultsmentioning

confidence: 99%

Oxidation-Induced Protein Cross-Linking in Mammalian Cells

Tang

et al. 2023

ACS Synth. Biol.

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A proximity-enabled protein cross-linking strategy with additional spatiotemporal control is highly desirable. Here, we report an oxidation-induced protein cross-linking strategy involving the incorporation of a vinyl thioether group into proteins in both Escherichia coli and mammalian cells via genetic code expansion. We demonstrated that vinyl thioether can be selectively induced by exogenously added oxidant or by reactive oxygen species from the cellular environment, as well as by photocatalysts, and converted into a Michael acceptor, enabling fluorescence labeling and protein cross-linking.

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Section: Resultsmentioning

confidence: 99%

Oxidation-Induced Protein Cross-Linking in Mammalian Cells

Tang

et al. 2023

ACS Synth. Biol.

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“…Three ncAAs for SuFEx have been genetically encoded in E . coli and mammalian cells: fluorosulfate‐tyrosine (FSY, 20 ; Wang et al, 2018 ), fluorosulfonyloxybenzoyl‐lysine (FSK, 21 ; Liu, Cao, et al, 2021 ), and o ‐sulfonyl fluoride‐O‐methyltyrosine (SFY, 22 ; Li et al, 2022 ). All three ncAAs have been shown to crosslink His, Tyr, and Lys proteins.…”

Section: Genetically Encoded Chemical Crosslinkersmentioning

confidence: 99%

“…Aryl fluorosulfate and sulphonyl fluoride moieties are weak electrophiles that are stable in the physiological milieu but react efficiently with proximal amines and alcohols in biomolecules to give stable covalent bonds, which makes them ideal probes for biocompatible proximity-induced chemical crosslinking. Three ncAAs for SuFEx have been genetically encoded in E. coli and mammalian cells: fluorosulfate-tyrosine (FSY, 20;Wang et al, 2018), fluorosulfonyloxybenzoyl-lysine (FSK, 21;, and o-sulfonyl fluoride-O-methyltyrosine (SFY, 22;Li et al, 2022). All three ncAAs have been shown to crosslink His, Tyr, and Lys proteins.…”

Section: Genetically Encoded Chemical Crosslinkersmentioning

confidence: 99%

Genetically encoded crosslinkers to address protein–protein interactions

Aydin

Coin

2023

Protein Science

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Noncanonical amino acids (ncAAs) for photo‐ and chemical crosslinking are powerful biochemical tools for studying and manipulating interactions between proteins both in vitro and in intact cells. Since the first crosslinking ncAAs were genetically encoded about 20 years ago, the technology has now ripened beyond the proof‐of‐principle demonstrations and is contributing to the study of relevant biological questions in the frame of modern integrative approaches. Here, we provide an overview of available photo‐activatable ncAAs for photo‐crosslinking and electrophilic ncAAs for genetically encoded chemical crosslinking (GECX), with a major focus on the most recent entries such as ncAAs for SuFEx click chemistry and photo‐activatable ncAAs for chemical crosslinking. We present recent examples of the application of genetically encoded crosslinkers to capture protein–protein interactions and identify interaction partners in live cells, to investigate molecular mechanisms of protein function, to stabilize protein complexes for structural studies, to derive structural information about protein complexes from the physiological cell environment, up to perspective applications of GECX‐ncAAs for the development of covalent drugs.

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“…Inspired by the on-the-shelf covalent small-molecule drugs and hottest covalent proteins, − recently several electrophilic groups have been considered as warheads to modify aptamers to develop covalent aptamers. For example, Tivon et al have developed tosyl or sulfonamide-modified covalent aptamers to specifically cross-link and label proteins of interest (POI) .…”

Section: Introductionmentioning

confidence: 99%

Robust Covalent Aptamer Strategy Enables Sensitive Detection and Enhanced Inhibition of SARS-CoV-2 Proteins

et al. 2023

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Aptamer-based detection and therapy have made substantial progress with cost control and easy modification. However, the conformation lability of an aptamer typically causes the dissociation of aptamer−target complexes during harsh washes and other environmental stresses, resulting in only moderate detection sensitivity and a decreasing therapeutic effect. Herein, we report a robust covalent aptamer strategy to sensitively detect nucleocapsid protein and potently neutralize spike protein receptor binding domain (RBD), two of the most important proteins of SARS-CoV-2, after testing different crosslink electrophilic groups via integrating the specificity and efficiency. Covalent aptamers can specifically convert aptamer−protein complexes from the dynamic equilibrium state to stable and irreversible covalent complexes even in harsh environments. Covalent aptamer-based ELISA detection of nucleocapsid protein can surpass the gold standard, antibody-based sandwich ELISA. Further, covalent aptamer performs enhanced functional inhibition to RBD protein even in a blood vessel-mimicking flowing circulation system. The robust covalent aptamer-based strategy is expected to inspire more applications in accurate molecular modification, disease biomarker discovery, and other theranostic fields.

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Genetically encoded chemical crosslinking of carbohydrate

Cited by 42 publications

References 52 publications

Oxidation-Induced Protein Cross-Linking in Mammalian Cells

Oxidation-Induced Protein Cross-Linking in Mammalian Cells

Genetically encoded crosslinkers to address protein–protein interactions

Robust Covalent Aptamer Strategy Enables Sensitive Detection and Enhanced Inhibition of SARS-CoV-2 Proteins

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