Modular Enzyme‐ and Light‐Based Activation of Cyclopropene–Tetrazine Ligation

Ji, Ting; Kumar, Pratik; Huang, Wei; Kao, Wei‐Siang; Thompson, Adrian; Camarda, Frank M.; Laughlin, Scott T.

doi:10.1002/cbic.201900137

Cited by 31 publications

(29 citation statements)

References 43 publications

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“…Taking these factors into account in the kinetic analysis, we determined the second-order rate constants with tetrazine S17 and 3,6-di-2-pyridyl-1,2,4,5-tetrazine to be 0.11 and 0.05 M –1 s –1 , respectively (Figure F and Figures S6 and S7). The more sluggish kinetics with 3,6-di-2-pyridyl-1,2,4,5-tetrazine than with S17 was expected on the basis of literature reports of cyclopropene kinetics with this tetrazine. ,, This rate of uncaged ketone cyclopropene (0.05–0.11 M –1 s –1 ) is substantially higher than that of the caged version and 125–270-fold higher than that of the first-generation cyclopropene, thereby improving its potential for applications that require faster ligation kinetics, spatial and/or temporal control, and small bioorthogonal tags like cyclopropene. ,,, …”

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confidence: 85%

“…23 Conversely, installing the C3-difluoro inhibits such ring-opening isomerization. 20 These first-generation caged cyclopropenes have excellent modularity of activation 25 but exhibit relatively slow ligation kinetics with s-tetrazines (k 2 = 0.0004 M −1 s −1 ) in a buffered solution at neutral pH. A modular activatable cyclopropene system with an improved tetrazine ligation rate can greatly expand the possible applications of cyclopropenes in which spatial and/or temporal control is a high priority.…”

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confidence: 99%

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Caged Cyclopropenes with Improved Tetrazine Ligation Kinetics

et al. 2019

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Activatable cyclopropenes are unreactive toward their inverse electron demand Diels–Alder reaction partner (e.g., s-tetrazines) until they are activated. The activation strategy is highly modular due to the cyclopropene’s ability to be caged by various light- and enzyme-activatable groups. This work describes the next generation of activatable cyclopropenes with a new core scaffold that maintains the activation modularity of the first generation but improves upon the ligation kinetics with s-tetrazines by ≤270-fold.

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confidence: 85%

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Caged Cyclopropenes with Improved Tetrazine Ligation Kinetics

et al. 2019

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“…[10] Very few reactions reported to date can be activated by an enzyme.F or example,F ox and co-workers reported horseradish-peroxidase-activated oxidation of dihydrotetrazine to tetrazine [11] and Laughlin and co-workers reported the enzymatic uncaging of as teric blocking group on cyclopropene. [12] However,n oe nzyme-activated bioorthogonal reactions have been used in vivo.…”

Section: Introductionmentioning

confidence: 99%

Exploring the Condensation Reaction between Aromatic Nitriles and Amino Thiols To Optimize In Situ Nanoparticle Formation for the Imaging of Proteases and Glycosidases in Cells

Chen

Cheng

et al. 2020

Angew Chem Int Ed

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The condensation reaction between 6‐hydroxy‐2‐cyanobenzothiazole (CBT) and cysteine has been shown for various applications such as site‐specific protein labelling and in vivo cancer imaging. This report further expands the substrate scope of this reaction by varying the substituents on aromatic nitriles and amino thiols and testing their reactivity and ability to form nanoparticles for cell imaging. The structure–activity relationship study leads to the identification of the minimum structural requirement for the macrocyclization and assembly process in forming nanoparticles. One of the scaffolds made of 2‐pyrimidinecarbonitrile and cysteine joined by a benzyl linker was applied to design fluorescent probes for imaging caspase‐3/7 and β‐galactosidase activity in live cells. These results demonstrate the generality of this system for imaging hydrolytic enzymes.

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“…47 The bioorthogonal Diels-Alder reactions of tetrazines with strained alkene and alkyne dienophiles has become increasingly important to the chemical biology community due to their exceptional kinetics with rates that can exceed 10 6 M -1 s -1 with conformationally strained trans-cyclooctenes. [48][49][50][51] Recent interest in the development of photochemically inducible variants of tetrazine ligation have prompted the discovery of new methods for uncaging cyclopropene 52,53 and bicyclononyne 28 dienophiles. Tetrazine (Tz) synthesis is commonly achieved through the oxidation of dihydrotetrazine (DHTz) precursors, 54 and the DHTz/Tz redox couple has been used in electrochemically controlled bioconjugation at electrode surfaces, 55 in batteries, 56 and for colorimetric nitrous gas detection.…”

Section: Introductionmentioning

confidence: 99%

Enabling in vivo Photocatalytic Activation of Rapid Bioorthogonal Chemistry by Repurposing Si-Rhodamine Fluorophores as Cytocompatible Far-Red Photocatalysts

Wang¹,

Zhang²,

Zhang³

et al. 2021

Preprint

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<p><a></a>Chromophores that absorb in the tissue-penetrant far-red/near-infrared window have long served as photocatalysts for the generation of singlet oxygen for photodynamic therapy. However, the cytotoxicity and side-reactions associated with singlet oxygen sensitization have posed a problem for using long wavelength photocatalysis to initiate other types of chemical reactions in biological environments. Described here is the use of Si-Rhodamine (SiR) dyes as photocatalysts for inducing rapid bioorthogonal chemistry using 660 nm light through the oxidation of a dihydrotetrazine to a tetrazine in the presence of trans-cyclooctene dienophiles. SiRs have been commonly used as fluorophores for applications in biology, but have not previously been applied to catalyze chemical reactions. A dihydrotetrazine/tetrazine pair is described that displays high stability in both oxidation states. A series of SiR derivatives were evaluated, and the Janelia-SiR dyes were found to be especially effective in catalyzing rapid photooxidation at low catalyst loadings (typically 1 µM). A protein that was site-selectively modified by trans-cyclooctene was quantitively conjugated upon exposure to 660 nm light and a dihydrotetrazine. By contrast, a previously described methylene blue catalyst was found to rapidly degrade the protein. SiR-red light photocatalysis was used to crosslink hyaluronic acid derivatives that were functionalized by dihydrotetrazine and trans-cyclooctenes, enabling 3D culture of human prostate cancer cells. This photoinducible hydrogel formation could also be carried out in vivo in live mice through subcutaneous injection of a solution containing SiR photocatalyst and a Cy7-labeled hydrogel precursor, followed by brief in vivo irradiation with 660 nm light to produce a stable hydrogel material. This cytocompatible method for using red light photocatalysis to activate bioorthogonal chemistry is anticipated to find broad applications where spatiotemporal control is needed in the in vivo environment. <br></p>

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Modular Enzyme‐ and Light‐Based Activation of Cyclopropene–Tetrazine Ligation

Cited by 31 publications

References 43 publications

Caged Cyclopropenes with Improved Tetrazine Ligation Kinetics

Caged Cyclopropenes with Improved Tetrazine Ligation Kinetics

Exploring the Condensation Reaction between Aromatic Nitriles and Amino Thiols To Optimize In Situ Nanoparticle Formation for the Imaging of Proteases and Glycosidases in Cells

Enabling in vivo Photocatalytic Activation of Rapid Bioorthogonal Chemistry by Repurposing Si-Rhodamine Fluorophores as Cytocompatible Far-Red Photocatalysts

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