Click Chemistry has been used prominently in pharmaceutical and biotech R&D. It has become increasingly popular due its biocompatibility and its faster reaction speeds.1 Click Chemistry can be categorized as one of three generations.
The 1st generation of Click Chemistry uses a Cu(I) catalyst in the reaction between a terminal alkyne and an azide. Although the coupling reaction is successful, the addition of a Cu(I) catalyst can cause damage to the biomolecule.
The 2nd generation Click Chemistry utilizes the reaction between cyclooctyne and an azide via strain-promoted azide-alkyne Click Chemistry. This reaction is copper-free and proceeds due to the high-activation energy of the strained molecule. In this copper-free Click Chemistry, commonly used reagents include DBCO, OCT and BCN. The use of reagents such as BCN, prevents the production of regioselective mixtures.
The 3rd generation of Click Chemistry involves the ligation between a tetrazine and trans-Cyclooctene (TCO) without the use of a Cu(I) catalyst. The omission of the Cu(I) catalyst is ideal for in vivo cell labelling while still enabling a high-speed reaction.2 This has been found to allow protein modification at lower concentrations.3
The introduction of a PEG linker to Click Chemistry reagents enhances its water-solubility properties and makes it more convenient for biolabeling in aqueous buffers.
1 Biocojug Chem. 2008 December; 19(12):2297-2299. Doi: 10.1021/bc8004446.
2 Biocojug Chem. 2008 December; 19(12):2297-2299. Doi: 10.1021/bc8004446.
3 J. Am. Chem. Soc. 2008, 130, 13518-13519.