Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Fluorogenic Bifunctional trans-Cyclooctenes as Efficient Tools for Investigating Click-to-Release Kinetics.

ABSTRACT: The inverse electron demand Diels-Alder pyridazine elimination reaction between tetrazines and allylic substituted trans-cyclooctenes (TCOs) is a key player in bioorthogonal bond cleavage reactions. Determining the rate of elimination of alkylamine substrates has so far proven difficult. Here, we report a fluorogenic tool consisting of a TCO-linked EDANS fluorophore and a DABCYL quencher for accurate determination of both the click and release rate constants for any tetrazine at physiologically relevant concentrations.

SUBMITTER: de Geus MAR

PROVIDER: S-EPMC7496853 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Fluorogenic Bifunctional trans-Cyclooctenes as Efficient Tools for Investigating Click-to-Release Kinetics.

de Geus Mark A R MAR Maurits Elmer E Sarris Alexi J C AJC Hansen Thomas T Kloet Max S MS Kamphorst Kiki K Ten Hoeve Wolter W Robillard Marc S MS Pannwitz Andrea A Bonnet Sylvestre A SA Codée Jeroen D C JDC Filippov Dmitri V DV Overkleeft Herman S HS van Kasteren Sander I SI

Chemistry (Weinheim an der Bergstrasse, Germany) 20200603 44

The inverse electron demand Diels-Alder pyridazine elimination reaction between tetrazines and allylic substituted trans-cyclooctenes (TCOs) is a key player in bioorthogonal bond cleavage reactions. Determining the rate of elimination of alkylamine substrates has so far proven difficult. Here, we report a fluorogenic tool consisting of a TCO-linked EDANS fluorophore and a DABCYL quencher for accurate determination of both the click and release rate constants for any tetrazine at physiologically ...[more]

PMID: 32154603

Similar Datasets

Efficient low-cost preparation of <i>trans</i>-cyclooctenes using a simplified flow setup for photoisomerization.

Project description:AbstractBioorthogonal ligations have emerged as highly versatile chemical tools for biomedical research. The exceptionally fast reaction between 1,2,4,5-tetrazines and trans-cyclooctenes (TCOs), also known as tetrazine ligation, is frequently used in this regard. Growing numbers of applications for the tetrazine ligation led to an increased demand for TCO compounds, whose commercial availability is still very limited. Reported photochemical procedures for the preparation of TCOs using flow chemistry are straightforward and high yielding but require expensive equipment. Within this contribution, we present the construction and characterization of a low-cost flow photoreactor assembled from readily accessible components. Syntheses of all commonly used trans-cyclooctene derivatives were successfully carried out using the described system. We are convinced that the presented system for photoisomerization will promote access to bioorthogonally reactive TCO derivatives.Graphical abstract

| S-EPMC4785212 | biostudies-literature

Genetic Encoding of bicyclononynes and trans-cyclooctenes for site-specific protein labeling in vitro and in live mammalian cells via rapid fluorogenic Diels-Alder reactions.

Project description:Rapid, site-specific labeling of proteins with diverse probes remains an outstanding challenge for chemical biologists. Enzyme-mediated labeling approaches may be rapid but use protein or peptide fusions that introduce perturbations into the protein under study and may limit the sites that can be labeled, while many "bioorthogonal" reactions for which a component can be genetically encoded are too slow to effect quantitative site-specific labeling of proteins on a time scale that is useful for studying many biological processes. We report a fluorogenic reaction between bicyclo[6.1.0]non-4-yn-9-ylmethanol (BCN) and tetrazines that is 3-7 orders of magnitude faster than many bioorthogonal reactions. Unlike the reactions of strained alkenes, including trans-cyclooctenes and norbornenes, with tetrazines, the BCN-tetrazine reaction gives a single product of defined stereochemistry. We have discovered aminoacyl-tRNA synthetase/tRNA pairs for the efficient site-specific incorporation of a BCN-containing amino acid, 1, and a trans-cyclooctene-containing amino acid 2 (which also reacts extremely rapidly with tetrazines) into proteins expressed in Escherichia coli and mammalian cells. We demonstrate the rapid fluorogenic labeling of proteins containing 1 and 2 in vitro, in E. coli , and in live mammalian cells. These approaches may be extended to site-specific protein labeling in animals, and we anticipate that they will have a broad impact on labeling and imaging studies.

| S-EPMC3687367 | biostudies-literature

Enhancing reactivity for bioorthogonal pretargeting by unmasking antibody-conjugated trans-cyclooctenes.

Project description:The bioorthogonal cycloaddition reaction between tetrazine and trans-cyclooctene (TCO) is rapidly growing in use for molecular imaging and cell-based diagnostics. We have surprisingly uncovered that the majority of TCOs conjugated to monoclonal antibodies using standard amine-coupling procedures are nonreactive. We show that antibody-bound TCOs are not inactivated by trans-cis isomerization and that the bulky cycloaddition reaction is not sterically hindered. Instead, TCOs are likely masked by hydrophobic interactions with the antibody. We show that introducing TCO via hydrophilic poly(ethylene glycol) (PEG) linkers can fully preserve reactivity, resulting in >5-fold enhancement in functional density without affecting antibody binding. This is accomplished using a novel dual bioorthogonal approach in which heterobifunctional dibenzylcyclooctyne (DBCO)-PEG-TCO molecules are reacted with azido-antibodies. Improved imaging capabilities are demonstrated for different cancer biomarkers using tetrazine-modified fluorophore and quantum dot probes. We believe that the PEG linkers prevent TCOs from burying within the antibody during conjugation, which could be relevant to other bioorthogonal tags and biomolecules. We expect the improved TCO reactivity obtained using the reported methods will significantly advance bioorthogonal pretargeting applications.

| S-EPMC5022565 | biostudies-literature

Development of benzothiazole 'click-on' fluorogenic dyes.

Project description:'Click-on' fluorogenic reaction: a non-fluorescent benzothiazole with an electron-deficient alkyne group at 2-position reacts with azide containing molecules could form fluorescent adducts.

| S-EPMC3010281 | biostudies-literature

Large-Scale Flow Photochemical Synthesis of Functionalized trans-Cyclooctenes Using Sulfonated Silica Gel.

Project description:Functionalized trans-cyclooctenes are useful bioorthogonal reagents that are typically prepared using a flow photoisomerization method where the product is captured by AgNO3 on silica gel. While this method is effective, the leaching of silver can be problematic when scaling up syntheses. It is shown here that Ag(I) immobilized on tosic silica gel can be used to capture trans-cyclooctene products at higher loadings without leaching. It is demonstrated that the sulfonated silica gel can be regenerated and reused with similar yields over multiple runs. Nine different trans-cyclooctenes were synthesized, including those commonly utilized in bioorthogonal chemistry as well as new amine and carboxylic acid derivatives.

| S-EPMC8224989 | biostudies-literature

Developing visible fluorogenic 'click-on' dyes for cellular imaging.

Project description:New fluorogenic dyes were designed and synthesized based on Cu(I)-catalyzed 'click' reaction. Conjugating weakly fluorescent benzothiazole derivatives with an electron-deficient alkyne group at the 2-position with azide-containing molecules in aqueous solution form 'click-on' fluorescent adducts. Model reactions and cell culture experiment indicated that the developed 'click-on' dye could be applied to labeling various biomolecules, such as nucleic acids, proteins, and other molecules, in cells.

| S-EPMC3178751 | biostudies-literature

Mechanism-Based Fluorogenic trans-Cyclooctene-Tetrazine Cycloaddition.

Project description:The development of fluorogenic reactions which lead to the formation of fluorescent products from two nonfluorescent starting materials is highly desirable, but challenging. Reported herein is a new concept of fluorescent product formation upon the inverse electron-demand Diels-Alder reaction of 1,2,4,5-tetrazines with particular trans-cyclooctene (TCO) isomers. In sharp contrast to known fluorogenic reagents the presented chemistry leads to the rapid formation of unprecedented fluorescent 1,4-dihydropyridazines so that the fluorophore is built directly upon the chemical reaction. Attachment of an extra fluorophore moiety is therefore not needed. The photochemical properties of the resulting dyes can be easily tuned by changing the substitution pattern of the starting 1,2,4,5-tetrazine. We support the claim with NMR measurements and rationalize the data by computational study. Cell-labeling experiments were performed to demonstrate the potential of the fluorogenic reaction for bioimaging.

| S-EPMC5299526 | biostudies-literature

Synthesis of a fluorogenic cyclooctyne activated by Cu-free click chemistry.

Project description:Cyclooctyne-based probes that become fluorescent upon reaction with azides are important targets for real-time imaging of azide-labeled biomolecules. The concise synthesis of a coumarin-conjugated cyclooctyne, coumBARAC, that undergoes a 10-fold enhancement in fluorescence quantum yield upon triazole formation with organic azides is reported. The design principles embodied in coumBARAC establish a platform for generating fluorogenic cyclooctynes suited for biological imaging.

| S-EPMC3219546 | biostudies-literature

Oligonucleotide labelling using a fluorogenic "click" reaction with a hemicarboxonium salt.

Project description:Two fluorescent streptocyanine labelled oligonucleotides have been synthesized by a simple "click" reaction between a non-fluorescent hemicarboxonium salt and aminoalkyl functionalized thymidines within the oligonucleotide and their spectrophotometric properties have been studied.

| S-EPMC6270631 | biostudies-literature

Genetically encoded tetrazine amino acid directs rapid site-specific in vivo bioorthogonal ligation with trans-cyclooctenes.

Project description:Bioorthogonal ligation methods with improved reaction rates and less obtrusive components are needed for site-specifically labeling proteins without catalysts. Currently no general method exists for in vivo site-specific labeling of proteins that combines fast reaction rate with stable, nontoxic, and chemoselective reagents. To overcome these limitations, we have developed a tetrazine-containing amino acid, 1, that is stable inside living cells. We have site-specifically genetically encoded this unique amino acid in response to an amber codon allowing a single 1 to be placed at any location in a protein. We have demonstrated that protein containing 1 can be ligated to a conformationally strained trans-cyclooctene in vitro and in vivo with reaction rates significantly faster than most commonly used labeling methods.

| S-EPMC3369569 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data