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  • Review Article
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Developing bioorthogonal probes to span a spectrum of reactivities

Nature Reviews Chemistry volume 4, pages 476–489 (2020)Cite this article

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Abstract

Bioorthogonal chemistries enable researchers to interrogate biomolecules in living systems. These reactions are highly selective and biocompatible, and can be performed in many complex environments. However, like any organic transformation, there is no perfect bioorthogonal reaction. Choosing the ‘best fit’ for a desired application is crucial. Correspondingly, there must be a variety of chemistries — spanning a range of rates and other features — to choose from. Over the past few years, considerable strides have been made towards not only expanding the number of bioorthogonal chemistries but also fine-tuning existing reactions for particular applications. In this Review, we highlight recent advances in the development of bioorthogonal reactions, focusing on how principles of physical organic chemistry have guided probe design. The continued expansion of this toolset will provide precisely tuned reagents for manipulating bonds in distinct environments.

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Fig. 1: Translating reactions from round-bottom flasks to living systems.
Fig. 2: Tuning transformations for biological application.
Fig. 3: Reactivities of cyclopropenone derivatives.
Fig. 4: An expansive set of reagents for dipolar cycloaddition.
Fig. 5: A collection of strained alkenes for IEDDA reactions.
Fig. 6: Steric modifications tune cyclopropene reactivity.
Fig. 7: Combining bioorthogonal chemistries for multicomponent labelling.

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References

  1. Rodriguez, E. A. et al. The growing and glowing toolbox of fluorescent and photoactive proteins. Trends Biochem. Sci. 42, 111–129 (2017).

    CAS  PubMed  Google Scholar 

  2. Row, R. D. & Prescher, J. A. Constructing new bioorthogonal reagents and reactions. Acc. Chem. Res. 51, 1073–1081 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Tian, Y. & Lin, Q. Fitness factors for bioorthogonal chemical probes. ACS Chem. Biol. 14, 2489–2496 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Sletten, E. M. & Bertozzi, C. R. Bioorthogonal chemistry: fishing for selectivity in a sea of functionality. Angew. Chem. Int. Ed. 48, 6974–6998 (2009).

    CAS  Google Scholar 

  5. Lang, K. & Chin, J. W. Cellular incorporation of unnatural amino acids and bioorthogonal labeling of proteins. Chem. Rev. 114, 4764–4806 (2014).

    CAS  PubMed  Google Scholar 

  6. Carrell, T. et al. in Cycloadditions in Bioorthogonal Chemistry (eds Vrabel, M. & Carell, T.) 1–157 (Springer, 2016).

  7. Andresen, M., Schmitz-Salue, R. & Jakobs, S. Short tetracysteine tags to β-tubulin demonstrate the significance of small labels for live cell imaging. Mol. Biol. Cell 15, 5616–5622 (2004).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Grammel, M. & Hang, H. C. Chemical reporters for biological discovery. Nat. Chem. Biol. 9, 475–484 (2013).

    CAS  PubMed  Google Scholar 

  9. Chai, Q.-Y., Yang, Z., Lin, H.-W. & Han, B.-N. Alkynyl-containing peptides of marine origin: a review. Mar. Drugs 14, 216 (2016).

    PubMed Central  Google Scholar 

  10. Jewett, J. C. & Bertozzi, C. R. Cu-free click cycloaddition reactions in chemical biology. Chem. Soc. Rev. 39, 1272–1279 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Selvaraj, R. & Fox, J. M. trans-Cyclooctene — a stable, voracious dienophile for bioorthogonal labeling. Curr. Opin. Chem. Biol. 17, 753–760 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Dommerholt, J., Rutjes, F. P. J. T. & van Delft, F. L. in Cycloadditions in Bioorthogonal Chemistry (eds Vrabel, M. & Carell, T.) 57–76 (Springer, 2016).

  13. Kölmel, D. K. & Kool, E. T. Oximes and hydrazones in bioconjugation: mechanism and catalysis. Chem. Rev. 117, 10358–10376 (2017).

    PubMed  PubMed Central  Google Scholar 

  14. Mahal, L. K., Yarema, K. J. & Bertozzi, C. R. Engineering chemical reactivity on cell surfaces through oligosaccharide biosynthesis. Science 276, 1125–1128 (1997).

    CAS  PubMed  Google Scholar 

  15. Datta, D., Wang, P., Carrico, I. S., Mayo, S. L. & Tirrell, D. A. A designed phenylalanyl-tRNA synthetase variant allows efficient in vivo incorporation of aryl ketone functionality into proteins. J. Am. Chem. Soc. 124, 5652–5653 (2002).

    CAS  PubMed  Google Scholar 

  16. Rabuka, D., Rush, J. S., deHart, G. W., Wu, P. & Bertozzi, C. R. Site-specific chemical protein conjugation using genetically encoded aldehyde tags. Nat. Protoc. 7, 1052–1067 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Hardisty, R. E., Kawasaki, F., Sahakyan, A. B. & Balasubramanian, S. Selective chemical labeling of natural T modifications in DNA. J. Am. Chem. Soc. 137, 9270–9272 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Dirksen, A., Dirksen, S., Hackeng, T. M. & Dawson, P. E. Nucleophilic catalysis of hydrazone formation and transimination: implications for dynamic covalent chemistry. J. Am. Chem. Soc. 128, 15602–15603 (2006).

    CAS  PubMed  Google Scholar 

  19. Dirksen, A., Hackeng, T. M. & Dawson, P. E. Nucleophilic catalysis of oxime ligation. Angew. Chem. Int. Ed. 45, 7581–7584 (2006).

    CAS  Google Scholar 

  20. Dirksen, A. & Dawson, P. E. Rapid oxime and hydrazone ligations with aromatic aldehydes for biomolecular labeling. Bioconjug. Chem. 19, 2543–2548 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Larsen, D. et al. Exceptionally rapid oxime and hydrazone formation promoted by catalytic amine buffers with low toxicity. Chem. Sci. 9, 5252–5259 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Saxon, E. & Bertozzi, C. R. Cell surface engineering by a modified Staudinger reaction. Science 287, 2007–2010 (2000).

    CAS  PubMed  Google Scholar 

  23. Kiick, K. L., Saxon, E., Tirrell, D. A. & Bertozzi, C. R. Incorporation of azides into recombinant proteins for chemoselective modification by the Staudinger ligation. Proc. Natl Acad. Sci. USA 99, 19–24 (2002).

    CAS  PubMed  Google Scholar 

  24. Saxon, E. et al. Investigating cellular metabolism of synthetic azidosugars with the Staudinger ligation. J. Am. Chem. Soc. 124, 14893–14902 (2002).

    CAS  PubMed  Google Scholar 

  25. Cai, J., Li, X., Yue, X. & Taylor, J. S. Nucleic acid-triggered fluorescent probe activation by the Staudinger reaction. J. Am. Chem. Soc. 126, 16324–16325 (2004).

    CAS  PubMed  Google Scholar 

  26. Hannoush, R. N. & Sun, J. The chemical toolbox for monitoring protein fatty acylation and prenylation. Nat. Chem. Biol. 6, 498–506 (2010).

    CAS  PubMed  Google Scholar 

  27. Chuh, K. N. & Pratt, M. R. Chemical methods for the proteome-wide identification of posttranslationally modified proteins. Curr. Opin. Chem. Biol. 24, 27–37 (2015).

    CAS  PubMed  Google Scholar 

  28. Prescher, J. A., Dube, D. H. & Bertozzi, C. R. Chemical remodelling of cell surfaces in living animals. Nature 430, 873–877 (2004).

    CAS  PubMed  Google Scholar 

  29. Meldal, M. & Tornøe, C. W. Cu-catalyzed azide–alkyne cycloaddition. Chem. Rev. 108, 2952–3015 (2008).

    CAS  PubMed  Google Scholar 

  30. Thirumurugan, P., Matosiuk, D. & Jozwiak, K. Click chemistry for drug development and diverse chemical–biology applications. Chem. Rev. 113, 4905–4979 (2013).

    CAS  PubMed  Google Scholar 

  31. Lampkowski, J. S., Villa, J. K., Young, T. S. & Young, D. D. Development and optimization of Glaser–Hay bioconjugations. Angew. Chem. Int. Ed. 54, 9343–9346 (2015).

    CAS  Google Scholar 

  32. Gaetke, L. M. & Chow, C. K. Copper toxicity, oxidative stress, and antioxidant nutrients. Toxicology 189, 147–163 (2003).

    CAS  PubMed  Google Scholar 

  33. Soares, E. V., Hebbelinck, K. & Soares, H. M. V. M. Toxic effects caused by heavy metals in the yeast Saccharomyces cerevisiae: a comparative study. Can. J. Microbiol. 49, 336–343 (2003).

    CAS  PubMed  Google Scholar 

  34. Kennedy, D. C. et al. Cellular consequences of copper complexes used to catalyze bioorthogonal click reactions. J. Am. Chem. Soc. 133, 17993–18001 (2011).

    CAS  PubMed  Google Scholar 

  35. del Amo, D. S. et al. Biocompatible copper(I) catalysts for in vivo imaging of glycans. J. Am. Chem. Soc. 132, 16893–16899 (2010).

    PubMed Central  Google Scholar 

  36. Besanceney-Webler, C. et al. Increasing the efficacy of bioorthogonal click reactions for bioconjugation: a comparative study. Angew. Chem. Int. Ed. 50, 8051–8056 (2011).

    CAS  Google Scholar 

  37. Uttamapinant, C. et al. Fast, cell-compatible click chemistry with copper-chelating azides for biomolecular labeling. Angew. Chem. Int. Ed. 51, 5852–5856 (2012).

    CAS  Google Scholar 

  38. Yang, M. et al. Biocompatible click chemistry enabled compartment-specific pH measurement inside E. coli. Nat. Commun. 5, 4981 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Wittig, G. & Krebs, A. Zur Existenz niedergliedriger cycloalkine, I [German]. Chem. Ber. 94, 3260–3275 (1961).

    CAS  Google Scholar 

  40. Agard, N. J., Prescher, J. A. & Bertozzi, C. R. A strain-promoted [3+2] azide–alkyne cycloaddition for covalent modification of biomolecules in living systems. J. Am. Chem. Soc. 126, 15046–15047 (2004).

    CAS  PubMed  Google Scholar 

  41. Rydahl, M. G., Hansen, A. R., Kračun, S. K. & Mravec, J. Report on the current inventory of the toolbox for plant cell wall analysis: proteinaceous and small molecular probes. Front. Plant Sci. 9, 581 (2018).

    PubMed  PubMed Central  Google Scholar 

  42. Yuet, K. P. et al. Cell-specific proteomic analysis in Caenorhabditis elegans. Proc. Natl Acad. Sci. USA 112, 2705–2710 (2015).

    CAS  PubMed  Google Scholar 

  43. tom Dieck, S. et al. Direct visualization of newly synthesized target proteins in situ. Nat. Methods 12, 411–414 (2015).

    CAS  PubMed  Google Scholar 

  44. Ernst, R. J. et al. Genetic code expansion in the mouse brain. Nat. Chem. Biol. 12, 776–778 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Alvarez-Castelao, B. et al. Cell-type-specific metabolic labeling of nascent proteomes in vivo. Nat. Biotechnol. 35, 1196–1201 (2017).

    CAS  PubMed  Google Scholar 

  46. Krogager, T. P. et al. Labeling and identifying cell-specific proteomes in the mouse brain. Nat. Biotechnol. 36, 156–159 (2018).

    CAS  PubMed  Google Scholar 

  47. van Geel, R., Prujin, G. J. M., van Delft, F. L. & Boelens, W. C. Preventing thiol-yne addition improves the specificity of strain-promoted azide–alkyne cycloaddition. Bioconjug. Chem. 23, 392–398 (2012).

    PubMed  Google Scholar 

  48. Blackman, M. L., Royzen, M. & Fox, J. M. Tetrazine ligation: fast bioconjugation based on inverse-electron-demand Diels–Alder reactivity. J. Am. Chem. Soc. 130, 13518–13519 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Taylor, M. T., Blackman, M. L., Dmitrenko, O. & Fox, J. M. Design and synthesis of highly reactive dienophiles for the tetrazine–trans-cyclooctene ligation. J. Am. Chem. Soc. 133, 9646–9649 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  50. Devaraj, N. K., Upadhyay, R., Haun, J. B., Hilderbrand, S. A. & Weissleder, R. Fast and sensitive pretargeted labeling of cancer cells through a tetrazine/trans-cyclooctene cycloaddition. Angew. Chem. Int. Ed. 48, 7013–7016 (2009).

    CAS  Google Scholar 

  51. Wang, M. et al. Conformationally strained trans-cyclooctene (sTCO) enables the rapid construction of 18F-PET probes via tetrazine ligation. Theranostics 6, 887–895 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Billaud, E. M. F. et al. Pretargeted PET imaging using a bioorthogonal 18F-labeled trans-cyclooctene in an ovarian carcinoma model. Bioconjug. Chem. 28, 2915–2920 (2017).

    CAS  PubMed  Google Scholar 

  53. Rossin, R. et al. Highly reactive trans-cyclooctene tags with improved stability for Diels–Alder chemistry in living systems. Bioconjug. Chem. 24, 1210–1217 (2013).

    CAS  PubMed  Google Scholar 

  54. Läppchen, T. et al. DOTA-tetrazine probes with modified linkers for tumor pretargeting. Nucl. Med. Biol. 55, 19–26 (2017).

    PubMed  Google Scholar 

  55. Bilton, H. A., Ahmad, Z., Janzen, N., Czorny, S. & Valliant, J. F. Preparation and evaluation of 99mTc-labeled tridentate chelates for pre-targeting using bioorthogonal chemistry. J. Vis. Exp. 120, e55188 (2017).

    Google Scholar 

  56. Versteegen, R. M., Rossin, R., ten Hoeve, W., Janssen, H. M. & Robillard, M. S. Click to release: instantaneous doxorubicin elimination upon tetrazine ligation. Angew. Chem. Int. Ed. 52, 14112–14116 (2013).

    CAS  Google Scholar 

  57. Rossin, R. et al. Chemically triggered drug release from an antibody–drug conjugate leads to potent antitumour activity in mice. Nat. Commun. 9, 1484 (2018).

    PubMed  PubMed Central  Google Scholar 

  58. Versteegen, R. M. et al. Click-to-release from trans-cyclooctenes: mechanistic insights and expansion of scope from established carbamate to remarkable ether cleavage. Angew. Chem. Int. Ed. 57, 10494–10499 (2018).

    CAS  Google Scholar 

  59. Mejia Oneto, J. M., Khan, I., Seebald, L. & Royzen, M. In vivo bioorthogonal chemistry enables local hydrogel and systemic pro-drug to treat soft tissue sarcoma. ACS Cent. Sci. 2, 476–482 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  60. Zhang, G. et al. Bioorthogonal chemical activation of kinases in living systems. ACS Cent. Sci. 2, 325–331 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  61. van der Gracht, A. M. F. et al. Chemical control over T-cell activation in vivo using deprotection of trans-cyclooctene-modified epitopes. ACS Chem. Biol. 13, 1569–1576 (2018).

    PubMed  PubMed Central  Google Scholar 

  62. Fan, X. et al. Optimized tetrazine derivatives for rapid bioorthogonal decaging in living cells. Angew. Chem. Int. Ed. 55, 14046–14050 (2016).

    CAS  Google Scholar 

  63. Schmidt, P., Zhou, L., Tishinov, K., Zimmermann, K. & Gillingham, D. Dialdehydes lead to exceptionally fast bioconjugations at neutral pH by virtue of a cyclic intermediate. Angew. Chem. Int. Ed. 53, 10928–10931 (2014).

    CAS  Google Scholar 

  64. Dilek, O., Lei, Z., Mukherjee, K. & Bane, S. Rapid formation of a stable boron–nitrogen heterocycle in dilute, neutral aqueous solution for bioorthogonal coupling reactions. Chem. Commun. 51, 16992–16995 (2015).

    CAS  Google Scholar 

  65. Schmidt, P., Stress, C. & Gillingham, D. Boronic acids facilitate rapid oxime condensations at neutral pH. Chem. Sci. 6, 3329–3333 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  66. Meadows, M. K., Roesner, E. K., Lynch, V. M., James, T. D. & Anslyn, E. V. Boronic acid mediated coupling of catechols and N-hydroxylamines: a bioorthogonal reaction to label peptides. Org. Lett. 19, 3179–3182 (2017).

    CAS  PubMed  Google Scholar 

  67. Bandyopadhyay, A., Cambray, S. & Gao, J. Fast diazaborine formation of semicarbazide enables facile labeling of bacterial pathogens. J. Am. Chem. Soc. 139, 871–878 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  68. Akgun, B. et al. Synergic “click” boronate/thiosemicarbazone system for fast and irreversible bioorthogonal conjugation in live cells. J. Am. Chem. Soc. 139, 14285–14291 (2017).

    CAS  PubMed  Google Scholar 

  69. Gu, H. et al. Formation of hydrazones and stabilized boron–nitrogen heterocycles in aqueous solution from carbohydrazides and ortho-formylphenylboronic acids. Org. Biomol. Chem. 15, 7543–7548 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  70. Bandyopadhyay, A., Cambray, S. & Gao, J. Fast and selective labeling of N-terminal cysteines at neutral pH via thiazolidino boronate formation. Chem. Sci. 7, 4589–4593 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  71. Faustino, H., Silva, M. J. S. A., Veiros, L. F., Bernardes, G. J. L. & Gois, P. M. P. Iminoboronates are efficient intermediates for selective, rapid and reversible N-terminal cysteine functionalisation. Chem. Sci. 7, 5052–5058 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  72. Sundhoro, M., Jeon, S., Park, J., Ramström, O. & Yan, M. Perfluoroaryl azide Staudinger reaction: a fast and bioorthogonal reaction. Angew. Chem. Int. Ed. 56, 12117–12121 (2017).

    CAS  Google Scholar 

  73. Meguro, T. et al. Staudinger reaction using 2,6-dichlorophenyl azide derivatives for robust aza-ylide formation applicable to bioconjugation in living cells. Chem. Commun. 54, 7904–7907 (2018).

    CAS  Google Scholar 

  74. Saneyoshi, H. et al. Triphenylphosphinecarboxamide: an effective reagent for the reduction of azides and its application to nucleic acid detection. Org. Lett. 16, 30–33 (2014).

    CAS  PubMed  Google Scholar 

  75. Luo, J., Liu, Q., Morihiro, K. & Deiters, A. Small-molecule control of protein function through Staudinger reduction. Nat. Chem. 8, 1027–1034 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  76. Lukasak, B., Morihiro, K. & Deiters, A. Aryl azides as phosphine-activated switches for small molecule function. Sci. Rep. 9, 1470 (2019).

    PubMed  PubMed Central  Google Scholar 

  77. Ren, G., Zheng, Q. & Wang, H. Aryl fluorosulfate trapped Staudinger reduction. Org. Lett. 19, 1582–1585 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  78. Böhrsch, V., Serwa, R., Majkut, P., Krause, E. & Hackenberger, C. P. R. Site-specific functionalisation of proteins by a Staudinger-type reaction using unsymmetrical phosphites. Chem. Commun. 46, 3176–3178 (2010).

    Google Scholar 

  79. Serwa, R. et al. Site-specific PEGylation of proteins by a Staudinger-phosphite reaction. Chem. Sci. 1, 596–602 (2010).

    CAS  Google Scholar 

  80. Nischan, N., Kasper, M.-A., Mathew, T. & Hackenberger, C. P. R. Bis(arylmethyl)-substituted unsymmetrical phosphites for the synthesis of lipidated peptides via Staudinger-phosphite reactions. Org. Biomol. Chem. 14, 7500–7508 (2016).

    CAS  PubMed  Google Scholar 

  81. Vallée, M. R. J. et al. Staudinger-phosphonite reactions for the chemoselective transformation of azido-containing peptides and proteins. Org. Lett. 13, 5440–5443 (2011).

    PubMed  Google Scholar 

  82. Kasper, M.-A. et al. Vinylphosphonites for Staudinger-induced chemoselective peptide cyclization and functionalization. Chem. Sci. 10, 6322–6329 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  83. Kasper, M.-A. et al. Cysteine-selective phosphonamidate electrophiles for modular protein bioconjugations. Angew. Chem. Int. Ed. 58, 11625–11630 (2019).

    CAS  Google Scholar 

  84. Lee, Y.-J., Kurra, Y. & Liu, W. R. Phospha-Michael addition as a new click reaction for protein functionalization. ChemBioChem 17, 456–461 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  85. Bos, J. & Muir, T. W. A chemical probe for protein crotonylation. J. Am. Chem. Soc. 140, 4757–4760 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  86. Shih, H.-W. & Prescher, J. A. A bioorthogonal ligation of cyclopropenones mediated by triarylphosphines. J. Am. Chem. Soc. 137, 10036–10039 (2015).

    CAS  PubMed  Google Scholar 

  87. Row, R. D., Shih, H.-W., Alexander, A. T., Mehl, R. A. & Prescher, J. A. Cyclopropenones for metabolic targeting and sequential bioorthogonal labeling. J. Am. Chem. Soc. 139, 7370–7375 (2017).

    CAS  PubMed  Google Scholar 

  88. Row, R. D. & Prescher, J. A. A cyclopropenethione-phosphine ligation for rapid biomolecule labeling. Org. Lett. 20, 5614–5617 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  89. Heiss, T. K. & Prescher, J. A. Cyclopropeniminium ions exhibit unique reactivity profiles with bioorthogonal phosphines. J. Org. Chem. 84, 7443–7448 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  90. Newberry, R. W., VanVeller, B., Guzei, I. A. & Raines, R. T. n→π* interactions of amides and thioamides: implications for protein stability. J. Am. Chem. Soc. 135, 7843–7846 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  91. Walters, C. R. et al. The effects of thioamide backbone substitution on protein stability: a study in α-helical, β-sheet, and polyproline II helical contexts. Chem. Sci. 8, 2868–2877 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  92. McKay, C. S., Moran, J. & Pezacki, J. P. Nitrones as dipoles for rapid strain-promoted 1,3-dipolar cycloadditions with cyclooctynes. Chem. Commun. 46, 931–933 (2010).

    CAS  Google Scholar 

  93. McKay, C. S., Chigrinova, M., Blake, J. A. & Pezacki, J. P. Kinetics studies of rapid strain-promoted [3+2]-cycloadditions of nitrones with biaryl-aza-cyclooctynone. Org. Biomol. Chem. 10, 3066–3070 (2012).

    CAS  PubMed  Google Scholar 

  94. MacKenzie, D. A. & Pezacki, J. P. Kinetics studies of rapid strain-promoted [3+2] cycloadditions of nitrones with bicyclo[6.1.0]nonyne. Can. J. Chem. 92, 337–340 (2014).

    CAS  Google Scholar 

  95. Gunawardene, P. N., Luo, W., Polgar, A. M., Corrigan, J. F. & Workentin, M. S. Highly electron-deficient pyridinium-nitrones for rapid and tunable inverse-electron-demand strain-promoted alkyne-nitrone cycloaddition. Org. Lett. 21, 5547–5551 (2019).

    CAS  PubMed  Google Scholar 

  96. McKay, C. S., Blake, J. A., Cheng, J., Danielson, D. C. & Pezacki, J. P. Strain-promoted cycloadditions of cyclic nitrones with cyclooctynes for labeling human cancer cells. Chem. Commun. 47, 10040–10042 (2011).

    CAS  Google Scholar 

  97. Sherratt, A. R. et al. Copper-catalysed cycloaddition reactions of nitrones and alkynes for bioorthogonal labelling of living cells. RSC Adv. 4, 46966–46969 (2014).

    CAS  Google Scholar 

  98. Sherratt, A. R. et al. Dual strain-promoted alkyne–nitrone cycloadditions for simultaneous labeling of bacterial peptidoglycans. Bioconjug. Chem. 27, 1222–1226 (2016).

    CAS  PubMed  Google Scholar 

  99. Decuypère, E., Plougastel, L., Audisio, D. & Taran, F. Sydnone–alkyne cycloaddition: applications in synthesis and bioconjugation. Chem. Commun. 53, 11515–11527 (2017).

    Google Scholar 

  100. Kolodych, S. et al. Discovery of chemoselective and biocompatible reactions using a high-throughput immunoassay screening. Angew. Chem. Int. Ed. 52, 12056–12060 (2013).

    CAS  Google Scholar 

  101. Wallace, S. & Chin, J. W. Strain-promoted sydnone bicyclo-[6.1.0]-nonyne cycloaddition. Chem. Sci. 5, 1742–1744 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  102. Plougastel, L. et al. 4-Halogeno-sydnones for fast strain promoted cycloaddition with bicyclo-[6.1.0]-nonyne. Chem. Commun. 50, 9376–9378 (2014).

    CAS  Google Scholar 

  103. Liu, H. et al. Ultrafast click chemistry with fluorosydnones. Angew. Chem. Int. Ed. 55, 12073–12077 (2016).

    CAS  Google Scholar 

  104. Mix, K. A., Aronoff, M. R. & Raines, R. T. Diazo compounds: versatile tools for chemical biology. ACS Chem. Biol. 11, 3233–3244 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  105. Moran, J., McKay, C. S. & Pezacki, J. P. Strain-promoted 1,3-dipolar cycloadditions of diazo compounds with cyclooctynes. Can. J. Chem. 89, 148–151 (2011).

    CAS  Google Scholar 

  106. Sanders, B. C. et al. Metal-free sequential [3+ 2]-dipolar cycloadditions using cyclooctynes and 1,3-dipoles of different reactivity. J. Am. Chem. Soc. 133, 949–957 (2011).

    CAS  PubMed  Google Scholar 

  107. Josa-Culleré, L., Wainman, Y. A., Brindle, K. M. & Leeper, F. J. Diazo group as a new chemical reporter for bioorthogonal labelling of biomolecules. RSC Adv. 4, 52241–52244 (2014).

    Google Scholar 

  108. Anderson, K. A., Aronoff, M. R., McGrath, N. A. & Raines, R. T. Diazo groups endure metabolism and enable chemoselectivity in cellulo. J. Am. Chem. Soc. 137, 2412–2415 (2015).

    Google Scholar 

  109. McGrath, N. A. & Raines, R. T. Diazo compounds as highly tunable reactants in 1,3-dipolar cycloaddition reactions with cycloalkynes. Chem. Sci. 3, 3237–3240 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  110. Aronoff, M. R., Gold, B. & Raines, R. T. 1,3-Dipolar cycloadditions of diazo compounds in the presence of azides. Org. Lett. 18, 1538–1541 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  111. Lim, R. K. V. & Lin, Q. Azirine ligation: fast and selective protein conjugation via photoinduced azirine–alkene cycloaddition. Chem. Commun. 46, 7993–7995 (2010).

    CAS  Google Scholar 

  112. King, M. & Wagner, A. Developments in the field of bioorthogonal bond forming reactions — past and present trends. Bioconjug. Chem. 25, 825–839 (2014).

    CAS  PubMed  Google Scholar 

  113. Schäfer, R. J. B. et al. The bioorthogonal isonitrile–chlorooxime ligation. J. Am. Chem. Soc. 141, 18644–18648 (2019).

    PubMed  Google Scholar 

  114. Yu, Z., Lim, R. K. V. & Lin, Q. Synthesis of macrocyclic tetrazoles for rapid photoinduced bioorthogonal 1,3-dipolar cycloaddition reactions. Chem. Eur. J. 16, 13325–13329 (2010).

    CAS  PubMed  Google Scholar 

  115. Yu, Z., Pan, Y., Wang, Z., Wang, J. & Lin, Q. Genetically encoded cyclopropene directs rapid, photoclick-chemistry-mediated protein labeling in mammalian cells. Angew. Chem. Int. Ed. 51, 10600–10604 (2012).

    CAS  Google Scholar 

  116. Kamber, D. N. et al. Isomeric cyclopropenes exhibit unique bioorthogonal reactivities. J. Am. Chem. Soc. 135, 13680–13683 (2013).

    CAS  PubMed  Google Scholar 

  117. An, P., Yu, Z. & Lin, Q. Design of oligothiophene-based tetrazoles for laser-triggered photoclick chemistry in living cells. Chem. Commun. 49, 9920–9922 (2013).

    CAS  Google Scholar 

  118. Yu, Z., Ohulchanskyy, T. Y., An, P., Prasad, P. N. & Lin, Q. Fluorogenic, two-photon-triggered photoclick chemistry in live mammalian cells. J. Am. Chem. Soc. 135, 16766–16769 (2013).

    CAS  PubMed  Google Scholar 

  119. Tasdelen, M. A. & Yagci, Y. Light-induced click reactions. Angew. Chem. Int. Ed. 52, 5930–5938 (2013).

    CAS  Google Scholar 

  120. Li, J. et al. Visible light-initiated bioorthogonal photoclick cycloaddition. J. Am. Chem. Soc. 140, 14542–14546 (2018).

    CAS  PubMed  Google Scholar 

  121. Zhang, X. et al. Photo-accelerated “click” reaction between diarylsydnones and ring-strained alkynes for bioorthogonal ligation. Chem. Commun. 55, 7187–7190 (2019).

    CAS  Google Scholar 

  122. Lim, R. K. V. & Lin, Q. Photoinducible bioorthogonal chemistry: a spatiotemporally controllable tool to visualize and perturb proteins in live cells. Acc. Chem. Res. 44, 828–839 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  123. An, P., Lewandowski, T. M., Erbay, T. G., Liu, P. & Lin, Q. Sterically shielded, stabilized nitrile imine for rapid bioorthogonal protein labeling in live cells. J. Am. Chem. Soc. 140, 4860–4868 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  124. de Almeida, G., Sletten, E. M., Nakamura, H., Palaniappan, K. K. & Bertozzi, C. R. Thiacycloalkynes for copper-free click chemistry. Angew. Chem. Int. Ed. 51, 2443–2447 (2012).

    Google Scholar 

  125. King, M., Baati, R. & Wagner, A. New tetramethylthiepinium (TMTI) for copper-free click chemistry. Chem. Commun. 48, 9308–9309 (2012).

    CAS  Google Scholar 

  126. de Almeida, G., Townsend, L. C. & Bertozzi, C. R. Synthesis and reactivity of dibenzoselenacycloheptynes. Org. Lett. 15, 3038–3041 (2013).

    PubMed  PubMed Central  Google Scholar 

  127. Ning, X., Guo, J., Wolfert, M. A. & Boons, G.-J. Visualizing metabolically labeled glycoconjugates of living cells by copper-free and fast Huisgen cycloadditions. Angew. Chem. Int. Ed. 47, 2253–2255 (2008).

    CAS  Google Scholar 

  128. Stöckmann, H. et al. Development and evaluation of new cyclooctynes for cell surface glycan imaging in cancer cells. Chem. Sci. 2, 932–936 (2011).

    PubMed  PubMed Central  Google Scholar 

  129. Friscourt, F. et al. Polar dibenzocyclooctynes for selective labeling of extracellular glycoconjugates of living cells. J. Am. Chem. Soc. 134, 5381–5389 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  130. Varga, B. R., Kállay, M., Hegyi, K., Béni, S. & Kele, P. A non-fluorinated monobenzocyclooctyne for rapid copper-free click reactions. Chem. Eur. J. 18, 822–828 (2012).

    CAS  PubMed  Google Scholar 

  131. Dommerholt, J. et al. Readily accessible bicyclononynes for bioorthogonal labeling and three-dimensional imaging of living cells. Angew. Chem. Int. Ed. 49, 9422–9425 (2010).

    CAS  Google Scholar 

  132. Agard, N. J., Baskin, J. M., Prescher, J. A., Lo, A. & Bertozzi, C. R. A comparative study of bioorthogonal reactions with azides. ACS Chem. Biol. 1, 644–648 (2006).

    CAS  PubMed  Google Scholar 

  133. Baskin, J. M. et al. Copper-free click chemistry for dynamic in vivo imaging. Proc. Natl Acad. Sci. USA 104, 16793–16797 (2007).

    CAS  PubMed  Google Scholar 

  134. Sletten, E. M., Nakamura, H., Jewett, J. C. & Bertozzi, C. R. Difluorobenzocyclooctyne: synthesis, reactivity, and stabilization by β-cyclodextrin. J. Am. Chem. Soc. 132, 11799–11805 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  135. Li, W. et al. Fluorodibenzocyclooctynes: a trackable click reagent with enhanced reactivity. Chem. Eur. J. 25, 10328–10332 (2019).

    CAS  PubMed  Google Scholar 

  136. Sletten, E. M. & Bertozzi, C. R. A hydrophilic azacyclooctyne for Cu-free click chemistry. Org. Lett. 10, 3097–3099 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  137. Jewett, J. C., Sletten, E. M. & Bertozzi, C. R. Rapid Cu-free click chemistry with readily synthesized biarylazacyclooctynones. J. Am. Chem. Soc. 132, 3688–3690 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  138. Debets, M. F. et al. Aza-dibenzocyclooctynes for fast and efficient enzyme PEGylation via copper-free (3+2) cycloaddition. Chem. Commun. 46, 97–99 (2010).

    CAS  Google Scholar 

  139. Gold, B., Shevchenko, N. E., Bonus, N., Dudley, G. B. & Alabugin, I. V. Selective transition state stabilization via hyperconjugative and conjugative assistance: stereoelectronic concept for copper-free click chemistry. J. Org. Chem. 77, 75–89 (2012).

    CAS  PubMed  Google Scholar 

  140. Gold, B., Batsomboon, P., Dudley, G. B. & Alabugin, I. V. Alkynyl crown ethers as a scaffold for hyperconjugative assistance in noncatalyzed azide–alkyne click reactions: ion sensing through enhanced transition-state stabilization. J. Org. Chem. 79, 6221–6232 (2014).

    CAS  PubMed  Google Scholar 

  141. Kaneda, K., Naruse, R. & Yamamoto, S. 2-Aminobenzenesulfonamide-containing cyclononyne as adjustable click reagent for strain-promoted azide–alkyne cycloaddition. Org. Lett. 19, 1096–1099 (2017).

    CAS  PubMed  Google Scholar 

  142. Burke, E. G., Gold, B., Hoang, T. T., Raines, R. T. & Schomaker, J. M. Fine-tuning strain and electronic activation of strain-promoted 1,3-dipolar cycloadditions with endocyclic sulfamates in SNO-OCTs. J. Am. Chem. Soc. 139, 8029–8037 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  143. Ni, R., Mitsuda, N., Kashiwagi, T., Igawa, K. & Tomooka, K. Heteroatom-embedded medium-sized cycloalkynes: concise synthesis, structural analysis, and reactions. Angew. Chem. Int. Ed. 54, 1190–1194 (2015).

    CAS  Google Scholar 

  144. Del Grosso, A. et al. Strained alkynes derived from 2,2′-dihydroxy-1,1′-biaryls; synthesis and copper-free cycloaddition with azides. Org. Biomol. Chem. 15, 4517–4521 (2017).

    PubMed  Google Scholar 

  145. Mistry, A. et al. Synthesis and cycloaddition reactions of strained alkynes derived from 2,2′-dihydroxy-1,1′-biaryls. Org. Biomol. Chem. 16, 8965–8975 (2018).

    CAS  PubMed  Google Scholar 

  146. Harris, T. et al. Twisted cycloalkynes and remote activation of “click” reactivity. Chem 3, 629–640 (2017).

    CAS  Google Scholar 

  147. Meier, H., Schuh-Popitz, C. & Peiersen, H. Isolation of a highly strained bicyclic alkyne. Angew. Chem. Int. Ed. Engl. 20, 270–271 (1981).

    Google Scholar 

  148. Ho, S. H. & Tirrell, D. A. Enzymatic labeling of bacterial proteins for super-resolution imaging in live cells. ACS Cent. Sci. 5, 1911–1919 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  149. Liu, F., Liang, Y. & Houk, K. N. Bioorthogonal cycloadditions: computational analysis with the distortion/interaction model and predictions of reactivities. Acc. Chem. Res. 50, 2297–2308 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  150. Bickelhaupt, F. M. & Houk, K. N. Analyzing reaction rates with the distortion/interaction-activation strain model. Angew. Chem. Int. Ed. 56, 10070–10086 (2017).

    CAS  Google Scholar 

  151. Gordon, C. G. et al. Reactivity of biarylazacyclooctynones in copper-free click chemistry. J. Am. Chem. Soc. 134, 9199–9208 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  152. Tao, H. et al. Origins of halogen effects in bioorthogonal sydnone cycloadditions. Chem. Commun. 54, 5082–5085 (2018).

    CAS  Google Scholar 

  153. Liu, F., Paton, R. S., Kim, S., Liang, Y. & Houk, K. N. Diels–Alder reactivities of strained and unstrained cycloalkenes with normal and inverse-electron-demand dienes: activation barriers and distortion/interaction analysis. J. Am. Chem. Soc. 135, 15642–15649 (2013).

    CAS  PubMed  Google Scholar 

  154. Liu, F., Liang, Y. & Houk, K. N. Theoretical elucidation of the origins of substituent and strain effects on the rates of Diels–Alder reactions of 1,2,4,5-tetrazines. J. Am. Chem. Soc. 136, 11483–11493 (2014).

    CAS  PubMed  Google Scholar 

  155. Darko, A. et al. Conformationally strained trans-cyclooctene with improved stability and excellent reactivity in tetrazine ligation. Chem. Sci. 5, 3770–3776 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  156. Kozma, E. et al. Hydrophilic trans-cyclooctenylated noncanonical amino acids for fast intracellular protein labeling. ChemBioChem 17, 1518–1524 (2016).

    CAS  PubMed  Google Scholar 

  157. Knorr, G., Kozma, E., Herner, A., Lemke, E. A. & Kele, P. New red-emitting tetrazine-phenoxazine fluorogenic labels for live-cell intracellular bioorthogonal labeling schemes. Chem. Eur. J. 22, 8972–8979 (2016).

    CAS  PubMed  Google Scholar 

  158. Lambert, W. D. et al. Computationally guided discovery of a reactive, hydrophilic trans-5-oxocene dienophile for bioorthogonal labeling. Org. Biomol. Chem. 15, 6640–6644 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  159. Siegl, S. J. et al. Design and synthesis of aza-bicyclononene dienophiles for rapid fluorogenic ligations. Chem. Eur. J. 24, 2426–2432 (2018).

    CAS  PubMed  Google Scholar 

  160. Bach, R. D. Ring strain energy in the cyclooctyl system. The effect of strain energy on [3+2] cycloaddition reactions with azides. J. Am. Chem. Soc. 131, 5233–5243 (2009).

    CAS  PubMed  Google Scholar 

  161. Santucci, J. III, Sanzone, J. R. & Woerpel, K. A. [4+2] cycloadditions of seven- membered-ring trans-alkenes: decreasing reactivity with increasing substitution of the seven-membered ring. Eur. J. Org. Chem. 2016, 2933–2943 (2016).

    CAS  Google Scholar 

  162. Fang, Y. et al. Photochemical syntheses, transformations, and bioorthogonal chemistry of trans-cycloheptene and sila trans-cycloheptene Ag(I) complexes. Chem. Sci. 9, 1953–1963 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  163. Patterson, D. M., Nazarova, L. A., Xie, B., Kamber, D. N. & Prescher, J. A. Functionalized cyclopropenes as bioorthogonal chemical reporters. J. Am. Chem. Soc. 134, 18638–18643 (2012).

    CAS  PubMed  Google Scholar 

  164. Yang, J., Šečkutė, J., Cole, C. M. & Devaraj, N. K. Live-cell imaging of cyclopropene tags with fluorogenic tetrazine cycloadditions. Angew. Chem. Int. Ed. 51, 7476–7479 (2012).

    CAS  Google Scholar 

  165. Ravasco, J. M. J. M., Monteiro, C. M. & Trindade, A. F. Cyclopropenes: a new tool for the study of biological systems. Org. Chem. Front. 4, 1167–1198 (2017).

    CAS  Google Scholar 

  166. Elliot, T. S. et al. Proteome labeling and protein identification in specific tissues and at specific developmental stages in an animal. Nat. Biotechnol. 32, 465–472 (2014).

    Google Scholar 

  167. Yu, Z. & Lin, Q. Design of spiro[2.3]hex-1-ene, a genetically encodable double-strained alkene for superfast photoclick chemistry. J. Am. Chem. Soc. 136, 4153–4156 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  168. Ramil, C. P. et al. Spirohexene-tetrazine ligation enables bioorthogonal labeling of class B G protein-coupled receptors in live cells. J. Am. Chem. Soc. 139, 13376–13386 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  169. An, P., Wu, H.-Y., Lewandowski, T. M. & Lin, Q. Hydrophilic azaspiroalkenes as robust bioorthogonal reporters. Chem. Commun. 54, 14005–14008 (2018).

    CAS  Google Scholar 

  170. Kumar, P., Jiang, T., Li, S., Zainul, O. & Laughlin, S. T. Caged cyclopropenes for controlling bioorthogonal reactivity. Org. Biomol. Chem. 16, 4081–4085 (2018).

    CAS  PubMed  Google Scholar 

  171. Kumar, P. et al. Caged cyclopropenes with improved tetrazine ligation kinetics. Org. Lett. 21, 3721–3725 (2019).

    CAS  PubMed  Google Scholar 

  172. Tu, J., Xu, M., Parvez, S., Peterson, R. T. & Franzini, R. M. Bioorthogonal removal of 3-isocyanopropyl groups enables the controlled release of fluorophores and drugs in vivo. J. Am. Chem. Soc. 140, 8410–8414 (2018).

    CAS  PubMed  Google Scholar 

  173. Tu, J. et al. Isonitrile-responsive and bioorthogonally removable tetrazine protecting groups. Chem. Sci. 11, 169–179 (2020).

    CAS  PubMed  Google Scholar 

  174. Xu, M., Deb, T., Tu, J. & Franzini, R. M. Tuning isonitrile/tetrazine chemistry for accelerated deprotection and formation of stable conjugates. J. Org. Chem. 84, 15520–15529 (2019).

    CAS  PubMed  Google Scholar 

  175. Stöckmann, H., Neves, A. A., Stairs, S., Brindle, K. M. & Leeper, F. J. Exploring isonitrile-based click chemistry for ligation with biomolecules. Org. Biomol. Chem. 9, 7303–7305 (2011).

    PubMed  Google Scholar 

  176. Chen, Y. et al. Addition of Isocyanide-containing amino acids to the genetic code for protein labeling and activation. ACS Chem. Biol. 14, 2793–2799 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  177. Stairs, S. et al. Metabolic glycan imaging by isonitrile–tetrazine click chemistry. ChemBioChem 14, 1063–1067 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  178. Niederwieser, A. et al. Two-color glycan labeling of live cells by a combination of Diels–Alder and click chemistry. Angew. Chem. Int. Ed. 52, 4265–4268 (2013).

    CAS  Google Scholar 

  179. Lee, Y.-J. et al. Genetically encoded unstrained olefins for live cell labeling with tetrazine dyes. Chem. Commun. 50, 13085–13088 (2014).

    CAS  Google Scholar 

  180. Eising, S., Lelivelt, F. & Bonger, K. M. Vinylboronic acids as fast reacting, synthetically accessible, and stable bioorthogonal reactants in the Carboni–Lindsey reaction. Angew. Chem. Int. Ed. 55, 12243–12247 (2016).

    CAS  Google Scholar 

  181. Eising, S. et al. Coordination-assisted bioorthogonal chemistry: orthogonal tetrazine ligation with vinylboronic acid and a strained alkene. ChemBioChem 19, 1648–1652 (2018).

    CAS  PubMed  Google Scholar 

  182. Eising, S., Engwerda, A. H. J., Riedijk, X., Bickelhaupt, F. M. & Bonger, K. M. Highly stable and selective tetrazines for the coordination-assisted bioorthogonal ligation with vinylboronic acids. Bioconjug. Chem. 29, 3054–3059 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  183. Eising, S., van der Linden, N. G. A., Kleinpenning, F. & Bonger, K. M. Vinylboronic acids as efficient bioorthogonal reactants for tetrazine labeling in living cells. Bioconjug. Chem. 29, 982–986 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  184. Engelsma, S. B. et al. Acylazetine as a dienophile in bioorthogonal inverse electron-demand Diels–Alder ligation. Org. Lett. 16, 2744–2747 (2014).

    CAS  PubMed  Google Scholar 

  185. de Montes, E. G. et al. Azabicyclic vinyl sulfones for residue-specific dual protein labelling. Chem. Sci. 10, 4515–4522 (2019).

    Google Scholar 

  186. Oliveira, B. L. et al. A minimal, unstrained S-allyl handle for pre-targeting Diels–Alder bioorthogonal labeling in live cells. Angew. Chem. Int. Ed. 55, 14683–14687 (2016).

    CAS  Google Scholar 

  187. Oliveira, B. L., Guo, Z. & Bernardes, G. J. L. Inverse electron demand Diels–Alder reactions in chemical biology. Chem. Soc. Rev. 46, 4895–4950 (2017).

    CAS  PubMed  Google Scholar 

  188. Chen, W., Wang, D., Dai, C., Hamelberg, D. & Wang, B. Clicking 1,2,4,5-tetrazine and cyclooctynes with tunable reaction rates. Chem. Commun. 48, 1736–1738 (2012).

    CAS  Google Scholar 

  189. Wang, D. et al. 3,6-Substituted-1,2,4,5-tetrazines: tuning reaction rates for staged labeling applications. Org. Biomol. Chem. 12, 3950–3955 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  190. Blizzard, R. J. et al. Ideal bioorthogonal reactions using a site-specifically encoded tetrazine amino acid. J. Am. Chem. Soc. 137, 10044–10047 (2015).

    CAS  PubMed  Google Scholar 

  191. Lambert, W. D. et al. Installation of minimal tetrazines through silver-mediated Liebeskind–Srogl coupling with arylboronic acids. J. Am. Chem. Soc. 141, 17068–17074 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  192. Šečkutė, J. & Devaraj, N. K. Expanding room for tetrazine ligations in the in vivo chemistry toolbox. Curr. Opin. Chem. Biol. 17, 761–767 (2013).

    PubMed  Google Scholar 

  193. Zhang, H. et al. Rapid bioorthogonal chemistry turn-on through enzymatic or long wavelength photocatalytic activation of tetrazine ligation. J. Am. Chem. Soc. 138, 5978–5983 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  194. Ehret, F., Wu, H., Alexander, S. C. & Devaraj, N. K. Electrochemical control of rapid bioorthogonal tetrazine ligations for selective functionalization of microelectrodes. J. Am. Chem. Soc. 137, 8876–8879 (2015).

    CAS  PubMed  Google Scholar 

  195. Kamber, D. N. et al. 1,2,4-Triazines are versatile bioorthogonal reagents. J. Am. Chem. Soc. 137, 8388–8391 (2015).

    CAS  PubMed  Google Scholar 

  196. Horner, K. A., Valette, N. M. & Webb, M. E. Strain-promoted reaction of 1,2,4-triazines with bicyclononynes. Chem. Eur. J. 21, 14376–14381 (2015).

    CAS  PubMed  Google Scholar 

  197. Baalmann, M., Ziegler, M. J., Werther, P., Wilhelm, J. & Wombacher, R. Enzymatic and site-specific ligation of minimal-size tetrazines and triazines to proteins for bioconjugation and live-cell imaging. Bioconjug. Chem. 30, 1405–1414 (2019).

    CAS  PubMed  Google Scholar 

  198. Peewasan, K. & Wagenknecht, H.-A. 1,2,4-Triazine-modified 2′-deoxyuridine triphosphate for efficient bioorthogonal fluorescent labeling of DNA. ChemBioChem 18, 1473–1476 (2017).

    CAS  PubMed  Google Scholar 

  199. Reisacher, U. et al. Triazine-modified 7-deaza-2′-deoxyadenosines: better suited for bioorthogonal labeling of DNA by PCR than 2′-deoxyuridines. Bioconjug. Chem. 30, 1773–1780 (2019).

    CAS  PubMed  Google Scholar 

  200. Siegl, S. J., Dzijak, R., Vázquez, A., Pohl, R. & Vrabel, M. The discovery of pyridinium 1,2,4-triazines with enhanced performance in bioconjugation reactions. Chem. Sci. 8, 3593–3598 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  201. Kozhevnikov, V. N., Deary, M. E., Mantso, T., Panayiotidis, M. I. & Sims, M. T. Iridium(III) complexes of 1,2,4-triazines as potential bioorthogonal reagents: metal coordination facilitates luminogenic reaction with strained cyclooctynes. Chem. Commun. 55, 14283–14286 (2019).

    CAS  Google Scholar 

  202. Kamber, D. N. et al. Isomeric triazines exhibit unique profiles of bioorthogonal reactivity. Chem. Sci. 10, 9109–9114 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  203. Levandowski, B. J., Abularrage, N. S., Houk, K. N. & Raines, R. T. Hyperconjugative antiaromaticity activates 4H-pyrazoles as inverse-electron-demand Diels–Alder dienes. Org. Lett. 21, 8492–8495 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  204. Bruins, J. J., Albada, B. & van Delft, F. ortho-Quinones and analogues thereof: highly reactive intermediates for fast and selective biofunctionalization. Chem. Eur. J. 24, 4749–4756 (2018).

    CAS  PubMed  Google Scholar 

  205. Borrmann, A. et al. Strain-promoted oxidation-controlled cyclooctyne–1,2-quinone cycloaddition (SPOCQ) for fast and activatable protein conjugation. Bioconjug. Chem. 26, 257–261 (2015).

    CAS  PubMed  Google Scholar 

  206. Bruins, J. J. et al. Inducible, site-specific protein labeling by tyrosine oxidation–strain-promoted (4+2) cycloaddition. Bioconjug. Chem. 28, 1189–1193 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  207. Gahtory, D., Sen, R., Kuzmyn, A. R., Escorihuela, J. & Zuilhof, H. Strain-promoted cycloaddition of cyclopropenes with o-quinones: a rapid click reaction. Angew. Chem. Int. Ed. 57, 10118–10122 (2018).

    CAS  Google Scholar 

  208. Li, Q., Dong, T., Liu, X. & Lei, X. A bioorthogonal ligation enabled by click cycloaddition of o-quinolinone quinone methide and vinyl thioether. J. Am. Chem. Soc. 135, 4996–4999 (2013).

    CAS  PubMed  Google Scholar 

  209. Minard, A., Liano, D., Wang, X. & Di Antonio, M. The unexplored potential of quinone methides in chemical biology. Bioorg. Med. Chem. 27, 2298–2305 (2019).

    CAS  PubMed  Google Scholar 

  210. Shih, H.-W., Kamber, D. N. & Prescher, J. A. Building better bioorthogonal reactions. Curr. Opin. Chem. Biol. 21, 103–111 (2014).

    CAS  PubMed  Google Scholar 

  211. Chang, P. V., Prescher, J. A., Hangauer, M. J. & Bertozzi, C. R. Imaging cell surface glycans with bioorthogonal chemical reporters. J. Am. Chem. Soc. 129, 8400–8401 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  212. Karver, M. R., Weissleder, R. & Hilderbrand, S. A. Bioorthogonal reaction pairs enable simultaneous, selective, multi-target imaging. Angew. Chem. Int. Ed. 51, 920–922 (2012).

    CAS  Google Scholar 

  213. Schoch, J., Staudt, M., Samanta, A., Wiessler, M. & Jäschke, A. Site-specific one-pot dual labeling of DNA by orthogonal cycloaddition chemistry. Bioconjug. Chem. 23, 1382–1386 (2012).

    CAS  PubMed  Google Scholar 

  214. Hudak, J. E., Alvarez, D., Skelly, A., von Adrian, U. H. & Kasper, D. L. Illuminating vital surface molecules of symbionts in health and disease. Nat. Microbiol. 2, 17099 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  215. Zhang, X. et al. Second generation TQ-ligation for cell organelle imaging. ACS Chem. Biol. 10, 1676–1683 (2015).

    CAS  PubMed  Google Scholar 

  216. Narayanam, M. K., Liang, Y., Houk, K. N. & Murphy, J. M. Discovery of new mutually orthogonal bioorthogonal cycloaddition pairs through computational screening. Chem. Sci. 7, 1257–1261 (2016).

    CAS  PubMed  Google Scholar 

  217. Shao, Z. et al. Bioorthogonal release of sulfonamides and mutually orthogonal liberation of two drugs. Chem. Commun. 54, 14089–14092 (2018).

    CAS  Google Scholar 

  218. Winz, M.-L., Linder, E. C., Becker, J. & Jäschke, A. Site-specific one-pot triple click labeling for DNA and RNA. Chem. Commun. 54, 11781–11784 (2018).

    CAS  Google Scholar 

  219. Simon, C. et al. One, two, three: a bioorthogonal triple labelling strategy for studying the dynamics of plant cell wall formation in vivo. Angew. Chem. Int. Ed. 57, 16665–16671 (2018).

    CAS  Google Scholar 

  220. Schart, V. F. et al. Triple orthogonal labeling of glycans by applying photoclick chemistry. ChemBioChem 20, 166–171 (2019).

    CAS  PubMed  Google Scholar 

  221. Sletten, E. M. & Bertozzi, C. R. A bioorthogonal quadricyclane ligation. J. Am. Chem. Soc. 133, 17570–17573 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  222. Italia, J. S. et al. Mutually orthogonal nonsense-suppression systems and conjugation chemistries for precise protein labeling at up to three distinct sites. J. Am. Chem. Soc. 141, 6204–6212 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  223. Tu, J. et al. Stable, reactive, and orthogonal tetrazines: dispersion forces promote the cycloaddition with isonitriles. Angew. Chem. Int. Ed. 58, 9043–9048 (2019).

    CAS  Google Scholar 

  224. Kim, J. & Bertozzi, C. R. A bioorthogonal reaction of N-oxide and boron reagents. Angew. Chem. Int. Ed. 54, 15777–15781 (2015).

    CAS  Google Scholar 

  225. Dong, J., Krasnova, L., Finn, M. G. & Sharpless, K. B. Sulfur(VI) fluoride exchange (SuFEx): another good reaction for click chemistry. Angew. Chem. Int. Ed. 53, 9430–9448 (2014).

    CAS  Google Scholar 

  226. Barrow, A. S. et al. The growing applications of SuFEx click chemistry. Chem. Soc. Rev. 48, 4731–4758 (2019).

    CAS  PubMed  Google Scholar 

  227. Zheng, Q. et al. SuFEx-enabled, agnostic discovery of covalent inhibitors of human neutrophil elastase. Proc. Natl Acad. Sci. USA 116, 18808–18814 (2019).

    CAS  PubMed  Google Scholar 

  228. Mortenson, D. E. et al. “Inverse drug discovery” strategy to identify proteins that are targeted by latent electrophiles as exemplified by aryl fluorosulfates. J. Am. Chem. Soc. 140, 200–210 (2018).

    CAS  PubMed  Google Scholar 

  229. Shannon, A. et al. Sulfonyl fluoride analogues as activity-based probes for serine proteases. ChemBioChem 13, 2327–2330 (2012).

    CAS  PubMed  Google Scholar 

  230. Hett, E. C. et al. Rational targeting of active-site tyrosine residues using sulfonyl fluoride probes. ACS Chem. Biol. 10, 1094–1098 (2015).

    CAS  PubMed  Google Scholar 

  231. Wang, N. et al. Genetically encoding fluorosulfate-l-tyrosine to react with lysine, histidine, and tyrosine via SuFEx in proteins in vivo. J. Am. Chem. Soc. 140, 4995–4999 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  232. Yang, B. et al. Proximity-enhanced SuFEx chemical cross-linker for specific and multitargeting cross-linking mass spectrometry. Proc. Natl Acad. Sci. USA 115, 11162–11167 (2018).

    CAS  PubMed  Google Scholar 

  233. Meng, G. et al. Modular click chemistry libraries for functional screens using a diazotizing reagent. Nature 574, 86–89 (2019).

    CAS  PubMed  Google Scholar 

  234. Kabakoff, D. S., Bünzil, J. C. G., Oth, J. F. M., Hammond, W. B. & Berson, J. A. Enthalpy and kinetics of isomerization of quadricyclane to norbornadiene. Strain energy of quadricyclane. J. Am. Chem. Soc. 97, 1510–1512 (1975).

    CAS  Google Scholar 

  235. Tomlin, F. M. et al. Site-specific incorporation of quadricyclane into a protein and photocleavage of the quadricyclane ligation adduct. Bioorg. Med. Chem. 26, 5280–5290 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  236. Martínez-Calvo, M. & Mascareñas, J. L. Organometallic catalysis in biological media and living settings. Coord. Chem. Rev. 359, 57–79 (2018).

    Google Scholar 

  237. Jang, S.-Y., Murale, D. P., Kim, A. D. & Lee, J.-S. Recent developments in metal- catalyzed bio-orthogonal reactions for biomolecule tagging. ChemBioChem 20, 1498–1507 (2019).

    CAS  PubMed  Google Scholar 

  238. Vidal, C., Tomás-Gamasa, M., Destito, P., López, F. & Mascareñas, J. L. Concurrent and orthogonal gold(I) and ruthenium(II) catalysis inside living cells. Nat. Commun. 9, 1913 (2018).

    PubMed  PubMed Central  Google Scholar 

  239. Pérez-López, A. M. et al. Gold-triggered uncaging chemistry in living systems. Angew. Chem. Int. Ed. 56, 12548–12552 (2017).

    Google Scholar 

  240. Tsubokura, K. et al. In vivo gold complex catalysis within live mice. Angew. Chem. Int. Ed. 56, 3579–3584 (2017).

    CAS  Google Scholar 

  241. Chalker, J. M., Wood, C. S. C. & Davis, B. G. A convenient catalyst for aqueous and protein Suzuki–Miyaura cross-coupling. J. Am. Chem. Soc. 131, 16346–16347 (2009).

    CAS  PubMed  Google Scholar 

  242. Clavadetscher, J., Indrigo, E., Chankeshwara, S. V., Lilienkampf, A. & Bradley, M. In-cell dual drug synthesis by cancer-targeting palladium catalysts. Angew. Chem. Int. Ed. 56, 6864–6868 (2017).

    CAS  Google Scholar 

  243. Li, N., Lim, R. K. V., Edwardraja, S. & Lin, Q. Copper-free Sonogashira cross-coupling for functionalization of alkyne-encoded proteins in aqueous medium and in bacterial cells. J. Am. Chem. Soc. 133, 15316–15319 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  244. Li, N., Ramil, C. P., Lim, R. K. V. & Lin, Q. A genetically encoded alkyne directs palladium-mediated protein labeling on live mammalian cell surface. ACS Chem. Biol. 10, 379–384 (2015).

    CAS  PubMed  Google Scholar 

  245. Li, J. et al. Palladium-triggered deprotection chemistry for protein activation in living cells. Nat. Chem. 6, 352–361 (2014).

    CAS  PubMed  Google Scholar 

  246. Wang, J. et al. Chemical remodeling of cell-surface sialic acids through a palladium-triggered bioorthogonal elimination reaction. Angew. Chem. Int. Ed. 54, 5364–5368 (2015).

    CAS  Google Scholar 

  247. Yusop, R. M., Unciti-Broceta, A., Johansson, E. M. V., Sánchez-Martin, R. M. & Bradley, M. Palladium-mediated intracellular chemistry. Nat. Chem. 3, 239–243 (2011).

    CAS  PubMed  Google Scholar 

  248. Wang, F., Zhang, Y., Du, Z., Ren, J. & Qu, X. Designed heterogeneous palladium catalysts for reversible light-controlled bioorthogonal catalysis in living cells. Nat. Commun. 9, 1209 (2018).

    PubMed  PubMed Central  Google Scholar 

  249. Tomás-Gamasa, M., Martínez-Calvo, M., Couceiro, J. R. & Mascareñas, J. L. Transition metal catalysis in the mitochondria of living cells. Nat. Commun. 7, 12538 (2016).

    PubMed  PubMed Central  Google Scholar 

  250. Lin, Y. A., Chalker, J. M., Floyd, N., Bernardes, G. J. L. & Davis, B. G. Allyl sulfides are privileged substrates in aqueous cross-metathesis: application to site-selective protein modification. J. Am. Chem. Soc. 130, 9642–9643 (2008).

    CAS  PubMed  Google Scholar 

  251. Lin, Y. A. et al. Rapid cross-metathesis for reversible protein modifications via chemical access to Se-allyl-selenocysteine in proteins. J. Am. Chem. Soc. 135, 12156–12159 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  252. Bloom, S. et al. Decarboxylative alkylation for site-selective bioconjugation of native proteins via oxidation potentials. Nat. Chem. 10, 205–211 (2018).

    CAS  PubMed  Google Scholar 

  253. Hooker, J. M., Kovacs, E. W. & Francis, M. B. Interior surface modification of bacteriophage MS2. J. Am. Chem. Soc. 126, 3718–3719 (2004).

    CAS  PubMed  Google Scholar 

  254. Sengupta, S. & Chandrasekaran, S. Modifications of amino acids using arenediazonium salts. Org. Biomol. Chem. 17, 8308–8329 (2019).

    CAS  PubMed  Google Scholar 

  255. Addy, P. S., Erickson, S. B., Italia, J. S. & Chatterjee, A. A chemoselective rapid azo-coupling reaction (CRACR) for unclickable bioconjugation. J. Am. Chem. Soc. 139, 11670–11673 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  256. Lin, S. et al. Redox-based reagents for chemoselective methionine bioconjugation. Science 355, 597–602 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  257. Taylor, M. T., Nelson, J. E., Suero, M. & Gaunt, M. J. A protein functionalization platform based on selective reactions at methionine residues. Nature 562, 563–568 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  258. Alvarez-Dorta, D. et al. Electrochemically promoted tyrosine-click-chemistry for protein labeling. J. Am. Chem. Soc. 140, 17120–17126 (2018).

    CAS  PubMed  Google Scholar 

  259. Geng, J. et al. Radical polymerization inside living cells. Nat. Chem. 11, 578–586 (2019).

    CAS  PubMed  Google Scholar 

  260. Vidal, C., Tomás-Gamasa, M., Gutiérrez-González, A. & Mascareñas, J. L. Ruthenium-catalyzed redox isomerizations inside living cells. J. Am. Chem. Soc. 141, 5125–5129 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

S.S.N. is an Allergan Graduate Research Fellow. J.A.P. is a Cottrell Scholar, Alfred P. Sloan Fellow and Dreyfus Scholar. This work was funded by the US National Institutes of Health (award no. R01 GM126226). The authors thank members of the Prescher lab for helpful discussions during manuscript preparation.

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  1. Department of Chemistry, University of California, Irvine, Irvine, CA, USA

    Sean S. Nguyen & Jennifer A. Prescher

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Nguyen, S.S., Prescher, J.A. Developing bioorthogonal probes to span a spectrum of reactivities. Nat Rev Chem 4, 476–489 (2020). https://doi.org/10.1038/s41570-020-0205-0

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