Click chemistry copper-free

The azide-aUcyne cycloaddition is highly relevant for biological applications. However, in vivo applications are limited by the fact of the toxicity of copper ions for living organisms. 

Electron-withdrawing groups tend to lower the LUMO of the alkyne and thus increase the interaction with the HOMO of the azide. The kinetics are comparable to the Cu-catalyzed Click reaction however, cycloadditions with these compounds are not regiose-lective. Specific DFT-measurements of the transition state were taken and previously published by Houk and coworkers.  

Although cyclooctynes enable copper-free Click reactions, they are not compatible with bioconjugations in aqueous media, because of their hydrophobic character. Thus, Bertozzi and coworkers uncovered a new method for the synthesis of a biocompatible cyclooctyne (6,7-dimethoxyazacyclooct-4-yne = DIMAC) to detect azide-labeled biomolecules via copper-free Click chemistry. The first conjugations to biotin as well as ceU-surface glycans were arranged. 

In the last years, other substrates like oxanorbomadienes by Comelissen et al. as well as dibenzocyclooctynes by Boon et al. were tested as (high) potential compounds for copper-free azide-aUcyne bioconjugations. 

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Codelli JA, Baskin JM, Agard NJ et al (2008) Second-generation difluorinated cyclooctynes for copper-free click chemistry. J Am Chem Soc 130 11486—11493... 

Baskin JM, Prescher JA, Laughlin ST, Agard NJ, Chang PV, Miller lA, Lo A, Codelli JA, Bertozzi CR (2007) Copper-free click chemistry for dynamic in vivo imaging. Proc Natl Acad Sci USA 104 16793-16797... 

For an example of such interaction in the TS of non-catalyzed alkyne/azide cycloaddition (click reaction), see Gold, B., Shevchenko, N. E., Bonus, N., Dudley, G. B., Alabugin, 1. V. (2012). Selective Transition State Stabihzation via Hyperconjugative and Conjugative Assistance Stereoelectronic Concept for Copper-Free Click Chemistry. The Journal of Organic Chemistry, 77(1), 75-89. 

High Density Orthogonal Surface Immobilization via Photoactivated Copper-Free Click Chemistry."... 

De Almeida G, Sletten EM, Nakamura H, Palaniappan KK, Bertozzi CR (2012) TTiiacycloalkynes for Copper-Free Click Chemistry. Angew Chem Int Ed 51 2443-2447... 

Copper-Free Click Chemistry for Nucleic Acid Functionalization... 

Copper-catalyzed and copper-free click chemistries have been used for metabolic labeling and fluorescent detection of DNA and RNA synthesis in vivo. Alkyne or alkene modifications on natural nucleosides are sufficiently small to allow efficient incorporation into DNA/RNA by cellular polymerases and circumvent drawbacks of immunohistochemical staining methods using 5-bromo-2 -deoxyuridine (Br-dU), such as the poor permeability of antibodies into various tissues. 

In summary, click chemistry on nucleic acid has revolutionized the synthesis and assembly of chemically modified oligonucleotides for various biophysical and biochemical applications. In vitro preparation of labeled DNA and RNA via copper-catalyzed and copper-free click chemistry is well established to date. 

Jayaprakash KN, Peng CG, Butler D, Varghese JP, Maier MA, Rajeev KG, Manoharan M (2010) Non-nucleoside building blocks for copper-assisted and copper-free click chemistry for the efficient synthesis of RNA conjugates. Org Lett 12(23) 5410-5413. doi 10.1021/oll02205j... 

Shelbourne M, Brown T Jr, El-Sagheer AH, Brown T (2012) Fast and efficient DNA crosslinking and multiple orthogonal labelling by copper-free click chemistry. Chem Commun 48(91) 1184-11186. doi 10.1039/c2cc35084j... 

Campbell-Verduyn, L.S., Mirfeizi, L., Schoonen, A.K., Dierckx, R.A., Elsinga, P.H., Feringa, B.L., 2011. Strain-promoted copper-free click chemistry for 18F radiolabeling of bombesin. Angew. Chem. Int. Edit. 50, 11117—11120. 

Johnson JA, Baskin JM, Bertozzi CR, Koberstein JT, Turro NJ. Copper-free click chemistry for the in sim crosslinking of photodegradable star polymers. Chem Comm 2008 44 3064-3066. 

Scheme 9.4 Biolabeling via copper-free Click chemistry with 6,7-dimethoxyazacyclooct-4-yne (33) ...

Gouin, S.G. and Kovensky, J. (2009) A procedure for fast and regioselective copper-free click chemistry at room temperature with p-toluenesulfonyl alkyne. Synlett, 1409. 

Fig. 1 (a) Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC)- and (b) copper-free click chemistry... 

Today, CuAAC and copper-free click chemistry are serving as complementary approaches to modify biomolecules for both in vitro and in vivo applications. In this chapter, we will highlight the recent applications of these triazole-forming reactions to the imaging and profiling of biomolecules. Comprehensive reviews of click chemistry-based approaches for bioconjugation have been published recently [12, 29]. 

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