Click chemistry mechanism, click chemistry applications

Click chemistry applications in drug discovery

Article on Click Chemistry


Click chemistry is a chemical philosophy introduced by K. Barry Sharpless of The Scripps Research Institute, in 2001 and describes chemistry tailored to generate substances quickly and reliably by joining small units together. This is inspired by the fact that nature also generates substances by joining small modular units. Click chemistry is not a specific reaction; it is a concept that mimics nature.

Click chemistry is a modular approach that uses only the most practical and reliable chemical transformations. Its applications are increasingly found in all aspects of drug discovery, ranging from lead finding through combinatorial chemistry and target-templated in situ chemistry, to proteomics and DNA research, using bioconjugation reactions.

Numerous examples of click reactions have been reported for preparation and functionalization of polymeric micelles and nanoparticles, liposomes and polymersomes, capsules, microspheres, metal and silica nanoparticles, carbon nanotubes and fullerenes, or bionanoparticles. Among these click processes, Cu (I)-catalyzed azide-alkyne cycloaddition (CuAAC) has attracted most attention based on its high orthogonality, reliability and experimental simplicity for non-specialists. The main aim of the write up is to focus on the importance of Click chemistry in drug discovery.

Taking cues from nature

Click chemistry is a chemical philosophy introduced by K Barry Sharpless of The Scripps Research Institute, in 2001 and describes chemistry tailored to generate substances quickly and reliably by joining small units together. This is inspired by the fact that nature also generates substances by joining small modular units.

“Generating substances by joining small units together with heteroatom links (C-X-C)“ is termed as Click Chemistry. The goal is to develop an expanding set of powerful, selective, and modular ‘blocks’ that work reliably in both small- and large-scale applications.

Click Chemistry reaction takes place only between Azide and Alkyne components. It is does not interfere with most any other organic groups present in DNA and proteins being labeled, such as amino and carboxy groups.

Click Chemistry is a reaction between azide and alkyne yielding covalent product triazole conjugate. This process is also known as CuAAC-Cu catalyzed alkyne azide cycloaddition.

Fig. 1) Azide-alkyne reaction

One of the most useful is the reaction between organic azides and alkynes, catalyzed by Cu (I) compounds. This reaction is a [3+2] cycloaddition which is highly specific, can be carried out in water and fully bioorthogonal. As a result, it has rapidly established a prominent role in materials science, medicinal, and bio conjugation chemistry.

Fig. 2) Click chemistry applications

It is important to recognize that click reactions achieve their required characteristics by having a high thermodynamic driving force, usually greater than 20 kcal/mol.

Carbon-heteroatom bond forming reactions comprise the most common examples, including the following classes of chemical transformations.

  • Cycloadditions of unsaturated species, especially 1,3-dipolar cycloaddition reactions, but also the Diels-Alder family of transformations
  • Nucleophilic substitution chemistry, particularly ring-opening reactions of strained heterocyclic electrophiles such as epoxides, aziridines, aziridinium ions, and episulfonium ions;
  • Carbonyl chemistry of the “non-aldol” type, such as formation of ureas, thioureas, aromatic heterocycles, oxime ethers, hydrazones, and amides; and
  • Additions to carbon-carbon multiple bonds, especially oxidative cases such as epoxidation, dihydroxylation, aziridination, and sulfenyl halide addition, but also Michael additions of Nu-H reactants

It describe reactions that are high yielding, wide in scope, create only byproducts that can be removed without chromatography, are stereo specific, simple to perform, and can be conducted in easily removable or benign solvents. It was developed in parallel with the interest within the pharmaceutical, materials, and other industries in capabilities for generating large libraries of compounds for screening in discovery research.

Click chemistry is a newer approach to the synthesis of drug-like molecules that can accelerate the drug discovery process by utilizing a few practical and reliable reactions.

In the search for new drugs, the Click Chemistry started to give very promising results.


The very potent inhibitors 1-3 of various enzymes were discovered by the fragment approach made possible by the use of the appropriate linkers (highlighted in red), constructed by [3+2] azide-alkyne cycloaddition.

Fig. 3) Enzyme inhibitors reaction


Click chemistry has widespread applications.

  • Preparative organic synthesis of 1, 4-substituted triazoles
  • Modification of peptide function with Triazoles
  • Modification of natural products and pharmaceuticals
  • Click chemistry is being used increasingly in biomedical research, ranging from lead discovery and optimization, to tagging of biological systems, such as proteins, nucleotides and whole organisms.
  • Macrocyclizations using Cu (I) catalyzed triazole couplings
  • Modification of DNA and nucleotides by triazole ligation
  • In Supramolecular chemistry
  • Dendrimer design
  • Carbohydrate clusters and carbohydrate conjugation by Cu(I) catalyzed triazole ligation reactions
  • Polymers
  • Material science
  • Nanotechnology and
  • Bioconjugation

Fig. 4) Click chemistry applications

Fig. 5) Click chemistry in Dendron’s design

Click chemistry in bioconjugation

Click chemistry has become a burgeoning strategy of bioconjugation in the development of bifunctional molecules. Bioconjugation involves the attachment of synthetic labels to biomolecular building blocks, such as fusing two or more proteins together or linking a carbohydrate with a peptide, and covers a wide range of science between molecular biology and chemistry. Although bioconjugation is applicable to the in vivo labeling of biomolecules, only a handful of reactions are actually useful.

The possibility of applying click chemistry in bioconjugation was first demonstrated by Tornoe et al. for the preparation of peptidotriazoles via solid-state synthesis. Their goal was to develop new, more efficient synthetic methods to prepare various [1,2,3]-triazole pharmacophores for potential biologic targets. This initial report makes possible the introduction of various novel functional and reporter groups into biomolecules, such as peptides and proteins, for DNA labeling and modification, and for cell-surface labeling.

Click chemistry continues to attract attention for the labeling of proteins and live organisms. Wang et al. successfully labeled Cowpea mosaic virus (CPMV) particles with fluorescein with >95% yield. The labeling was performed by modifying the surface of viral protein (either lysine or cysteine residues) with azides or alkynes, followed by reaction with fluorescein-bearing complementary groups. Similarly, Link and Tirrell were able to modify Eschericia coli with an azide-bearing outer membrane protein C (OmpC). The modified cell was then biotinylated by reacting with a biotinalkyne derivative under copper-catalyzed click chemistry conditions.

Deiters et al. developed a method to genetically encode proteins of Saccharomyces cerevisiae with azide- or acetylene-based synthetic amino acids. The genetic modification was done by reacting an alkyne- or an azide-bearing protein with the counterpart unnatural amino acid. In the same study, the possibility of inserting organic molecules to proteins by an azide-alkyne [3+2] cycloaddition reaction was demonstrated by reacting azide- or alkyne-bearing proteins with azide- or alkyne-bearing dyes.

Fig.6) Structures of synthetic amino acids (top) and dyes (bottom)


Click chemistry has proven to be a powerful tool in biomedical research, ranging from combinatorial chemistry and target-templated in situ chemistry for lead discovery, to bioconjugation strategies for proteomics and DNA research et al. It has become synonymous with the Huisgen 1, 3-dipolar cycloaddition reaction and has gained potential recognition in drug discovery. Moving forwards, the technology will play an influential role in the design of future drugs, owing to its simplicity. Looking at the current market trends, one can safely say that click chemistry can serve as a promising route in drug discovery.


[1] Shekh Yunus, Nirmal Das Adhikary and Partha Chattopadhyay, Click chemistry - A New Approach for Drug Discovery; Available from -  

[2] Enrique Lallana, Ana Sousa-Herves, Francisco Fernandez-Trillo, Ricardo Riguera, Eduardo Fernandez-Megia, Click Chemistry for Drug Delivery Nanosystems, 10.1007/s11095-011-0568-5,Sep 13, 2011, PAGES: 1-34-  

[3] Gregory C. Patton, DEVELOPMENT AND APPLICATIONS OF CLICK CHEMISTRY, November 8, 2004, Available from-  

[4] Dr. Joost A. Opsteen, Dr. Lee Ayres and Prof. Jan C.M. van Hest, "Click" Chemistry in (Bio) Materials Science, Material Matters Volume 3 Issue 3, Available from -  

[5] Piyush Chaturvedi,Nupur Chaturvedi,Setu Gupta, Amlan Mishra,Mithilesh Singh,Tarun Siddhartha, CLICK CHEMISTRY: A NEW APPROACH FOR DRUG DISCOVERY, Volume 10, Issue 2, September – October 2011; Article-022, Available from - 

Image Reference

Fig. 1):  Lumiprobe Corporation. Website,  

Fig. 2):  Hartmuth C. Kolb, M. G. Finn, and K. Barry Sharpless, Click Chemistry: Diverse Chemical Function from a Few Good Reactions; Available from- 

Fig. 3):  ENAMINE Ltd. Website, 

Fig. 4):  IRIS Biotech GmbH Website,

Fig. 5):  IRIS Biotech GmbH Website,

Fig. 6):  Kido Nwe and Martin W. Brechbiel, Growing Applications of “Click Chemistry” for Bioconjugation in Contemporary Biomedical Research, 2009 June; 24(3): 289–302-

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