Click Chemistry: Refocusing our strategy

Part of the series: Biochemical Tools & Techniques

Imagine you are a researcher and you have a problem. You need to attach a fluorescent probe to a specific protein inside a living cell. You have the tag and the protein, but how do you connect the two? Where do you start? The complexity of the cell is remarkable, and the task appears daunting.

Additionally, how do you ensure that you are bonding the appropriate two constituents together with sufficient efficiency and specificity? For decades, the complex chemistry required to solve this problem was a maddening reality for researchers. Expensive, multistep organic syntheses that yielded hazardous side products were the primary method for producing and modifying complex biomolecules. This is the story of how click chemistry became an elegant and precise solution to this problem.

LEGO brick illustration for the concept of click chemistry

Let us start this story with the historical background. In 2001, the term "click chemistry" was coined by American stereochemist K. Barry Sharpless in his work to simplify the synthesis of complex organic molecules. In his work, he proposed an alternative approach for chemists: refocusing from mimicking nature's complex molecules through multistep, expensive syntheses to using reliable, reproducible reactions. Sharpless aimed to develop a toolkit of energetically favorable reactions that could function as molecular Lego bricks for connecting components.

Click chemistry is not a single reaction but a design principle aimed at simplicity and reliability. It's about choosing the most efficient, foolproof way to connect molecular building blocks, like snapping LEGO bricks together. This contrasts with many traditional strategies, which were often expensive, hazardous, and time-consuming.

How do we determine if a reaction is a click reaction?

A click reaction has the following characteristics:

• Modular and wide in scope

• High-yielding

• Generates few, harmless byproducts

• Stereospecific

• Simple to perform (in water or benign solvents, tolerant of atmospheric conditions)

• Easy to purify

Sharpless and Morten Meldal, another chemist working independently on the concept, identified the copper-catalyzed azide-alkyne cycloaddition reaction (CuAAC) as a prime click reaction candidate. The reaction is reliable, regioselective (forming a single isomer), and compatible with water and atmospheric conditions.

Copper-catalyzed azide-alkyne cycloaddition reaction (CuAAC)

The CuAAC reaction grew rapidly in popularity, and biochemists took notice. However, there was a major issue with applying the CuAAC reaction in biology. Copper is toxic to cells. It can bind to enzymes in metabolic pathways, such as the TCA cycle, thereby disrupting cellular metabolism. The challenge, then, was to retain the desirable characteristics of CuAAC without a copper catalyst.

This was the challenge addressed by biochemist Carolyn Bertozzi. She found that by incorporating the alkyne into a ring, the azide-alkyne cycloaddition reaction could be carried out at high efficiency without copper. The ring strain activates the alkyne, enabling the strain-promoted azide-alkyne cycloaddition reaction (SPAAC). This brilliant solution gave rise to bioorthogonal chemistry, the use of such selective reactions within living systems.

Strain-promoted azide-alkyne cycloaddition reaction (SPAAC)

The combined work of Sharpless, Meldal, and Bertozzi did more than earn them a shared Nobel Prize in Chemistry; it sparked a revolution in biochemical thinking. Their work built on one another's, transforming a powerful concept into a valuable toolkit for addressing the living cell.

The future of click chemistry and bioorthogonal reactions is exciting. They enable a new strategy for developing therapeutic drugs with superior efficiency and reduced off-target effects.

While the iconic azide-alkyne "click" is the family's flagship, the philosophy extends far beyond. Reactions like the incredibly fast inverse electron-demand Diels-Alder (IEDDA), thiol-ene coupling, and others continue to expand the bioorthogonal toolkit. This is why the future of biochemistry is so exciting. Click chemistry is paving the way for smarter therapeutics, dramatically increasing efficiency and reducing off-target effects.

The story of click chemistry is ultimately a story of biochemists taking a step back and choosing simplicity and precision to solve biochemistry's most complex puzzles. Our imagination is our only limitation.