Berkeley Scientific Journal

Source: https://bsj.studentorg.berkeley.edu/bioorthogonal-chemistry-a-revolution-in-chemical-biology/ Parent: https://bsj.studentorg.berkeley.edu/features-department/

2025, BSJ Features, Spring 2025

Bioorthogonal Chemistry: A Revolution in Chemical Biology

Ruben Levya

December 6, 2025

INTRODUCTION

Life is built on chemistry, but until recently, chemistry had to be studied outside of living systems. The development of bioorthogonal chemistry has completely revolutionized this. Bioorthogonal chemistry refers to a class of specialized reactions that occur inside living organisms without interfering with natural biochemical processes.2 First introduced by Dr. Carolyn Bertozzi, Professor of Organic Chemistry at Stanford University, bioorthogonal chemistry has rapidly become a remarkable tool in research and medicine, from tracking molecules in cells to delivering drugs with incredible accuracy. But how did this field emerge, and where is it headed?\

HOW BIOORTHOGONAL CHEMISTRY WORKS

In a regular chemistry lab, reactions happen in carefully controlled flasks—with specific temperatures, concentrations, and solvents. However, inside the body, things are far more difficult to control. In the body, water is the main solvent, and the environment is surrounded by enzymes and other reactive biomolecules that are constantly reacting with one another.2, 11 Bioorthogonal chemistry changes these rules. It uses chemical groups that do not naturally exist in biology and will not react with anything except a similar group. When introduced into cells or animals, these groups ignore everything else and react only with each other2. This kind of precision allows\ scientists to label, manipulate, and track molecules in real time, making it incredibly useful for imaging and drug delivery.5,4, 11

\ HOW DID BIOORTHOGONAL CHEMISTRY EMERGE?

Bioorthogonal chemistry began with a simple but unresolved problem in biology: how can we study glycans? These complex sugars are found on the surface of cells an play key roles in communication, immune response, and disease. But unlike proteins or DNA, scientists did not have the tools to detect or visualize them.2,8 While working at UC Berkeley, Dr. Bertozzi imagined a two-step approach. First, feed cells a sugar with a unique chemical handle that could be added into the glycans. Second, use a chemical reaction to tag that handle with a probe, like a fluorescent dye. However, for this strategy to work, the reaction had to be safe and selective—meaning it had to happen inside cells without affecting anything else.6,7 That is where the term “bioorthogonal” came in: reactions that occur independently of other native biological processes. The first example was the Staudinger Ligation, originally developed by Hermann Staudinger in 1919. It involved a reaction between azides and phosphines. While it was highly selective, the reaction was slow and unstable in water, making it difficult to use in living organisms.1,3

Figure 1: Staudinger Ligation. In this reaction, the azide group (left red) reacts with the phosphine (left blue) to form an intermediate that eventually produces an amine (right red) and a phosphine oxide (right blue) in water. This reaction was one of the first example of bioorthogonal chemistry due to its hight selectivity.

Later, “click chemistry,” specifically the copper-catalyzed azide alkyne cycloaddition (CuAAC), developed by Dr. Barry Sharpless and Dr. Morten Medal, offered a much faster and stable reaction in water. However, the copper used in the reaction was toxic to cells, ruling out its use in biological systems.1,3 Dr. Bertozzi’s group solved this problem by developing copper-free click chemistry, which used strained ring molecules like cyclooctyne to enable the reaction without toxic metals. Further innovations, like the tetrazine ligation, made things even faster and more efficient, by using reactions between tetrazines and strained alkenes.1,3

Figure 2: Copper-Catalyzed Click Chemistry. This reaction shows an azide )red_and terminal alkyne (blue) react in the presence of copper to form a triazole ring. While highly efficient and works well with water, the copper is toxic to the cells.

Figure 3: Copper-Free Azide-Alkyne Cycloaddition. In this reaction the azide (red_reacts with a strained alkyne (blue) on a ring to form a stable triaxole (red and blue). Beacuse no copper was needed, this reaction is safe for use in living organisms.

Figure 4: Tetrazine Ligation. In this reaction, a strained alkene (red) reacts with a tetrazine (blue) to form a stable product after releasing nitrogen gas. The new ring (red and blue) is the result of a highly selective reaction that works well in living systems.

\ CURRENT APPLICATIONS OF BIOORTHOGONAL CHEMISTRY

Figure 5: BONCAT and FUNCAT. These are two bioorthogonal techniques used to study proteins in the cell. A special chemical handle (yellow, R1) is built into newly protein, which is then tagged with an affinity label (green, BONCAT) for purification or a fluorescent marker (blue, FUNCAT) for imaging.

1. Fluorescent Tagging and Molecular Imaging

One of the most exciting uses of bioorthogonal chemistry is its ability to help scientists visualize molecules inside living organisms. Techniques like BONCAT and FUNCAT allow researchers to tag newly made proteins in real time, offering insights into how cells grow, react to stress, or develop.1, 9 In one of Dr. Bertozzi’s known experiments, her team used bioorthogonal chemistry to track sugars in zebrafish embryos. After feeding the fish these modified sugars, her lab used fluorescent dyes to highlight where different sugars appeared during their development. This resulted in images that mapped the sugar patterns across the zebra’s living tissues.1,2

1. Delivering Drugs Precisely

Bioorthogonal chemistry is also changing how we deliver drugs. Antibody-drug conjugates (ADCs) use antibodies to guide chemotherapy drugs to cancer cells. The drug remains inactive until bioorthogonal reaction activates it at the tumor site. This helps protect healthy cells and reduce side effects.2, 10 Newer techniques, like click-to-release chemistry, take this one step further. Instead of needing light or heat to activate a drug, it uses a second chemical reaction inside the body. Companies like Shasqi and Palleon Pharmaceuticals are already exploring this in clinical trials, aiming to make treatments smarter and more targeted.2, 5

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*THE FUTURE OF BIOORTHOGONAL CHEMISTRY*

Figure 6: In situ drug assembly. In this mouse model, cancer drugs are activated only at the tumor site using a “click-to-release” reaction. A tetrazine or a strained molecule is first delivered to the tumor and then the drug is released through a bioorthogonal reaction.

As the field continues to grow, chemists are pushing the boundaries of bioorthogonal chemistry in both speed and precision. One ongoing challenge is improving reaction kinetics under biological conditions. For example, while tetrazines are fast, they tend to easily break in water. Chemists are working to develop more stable variants that remain effective in water-like environments, such as the plasma in blood.1, 2 Another exciting area is in situ drug assembly—a strategy where the components of a drug are delivered separately and only come together through a bioorthogonal reaction at the disease site. This would allow for ultra-targeted therapies that form only when needed, limiting toxicity or potential side-effects.4,5,9 Similarly, there is growing interest among chemists in multiplexed bioorthogonal labeling, where multiple reactions occur independently in the same cell or tissue. This technique would allow scientists to track several types of molecules simultaneously and get a richer picture of what’s happening inside the cell.1,4 While there is still a lot to learn, what’s happening in this field shows how much potential chemistry has to improve lives in ways chemists never expected. It’s a reminder that with collaboration and persistence, science can keep breaking boundaries. While the field of bioorthogonal chemistry is still relatively new, its impact on science is already clear. And for students, researchers, and future scientists, this is an exciting field to watch—and maybe even be a part of.\ \ ACKNOWLEDGMENTS

I’d like to thank Carolyn Bertozzi, Organic Chemistry Professor and Researcher at Stanford University, for generously reviewing my article and providing thoughtful feedback and grammar suggestions.\

REFERENCES

1. Bird, R. E., Lemmel, S. A., Yu, X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479. https://doi.org/10.1021/acs.bioconjchem.1c00461
2. Nobel Prize. (2022, December 8). Nobel Prize lecture: Carolyn Bertozzi, Nobel Prize in Chemistry 2022 [Video]. YouTube. https://www.youtube.com/watch?v=WyGoeQEHBEk
3. Devaraj, N. K. (2018). The future of bioorthogonal chemistry. ACS Central Science, 4(8), 952–959. https://doi.org/10.1021/acscentsci.8b00251
4. Mitry, M. M. A., Greco, F., & Osborn, H. M. I. (2023). In vivo applications of bioorthogonal reactions: chemistry and targeting mechanisms. Chemistry – a European Journal, 29(20). https://doi.org/10.1002/chem.202203942
5. Scinto, S. L., Bilodeau, D. A., Hincapie, R., Lee, W., Nguyen, S. S., Xu, M., Ende, C. W. A., Finn, M. G., Lang, K., Lin, Q., Pezacki, J. P., Prescher, J. A., Robillard, M. S., & Fox, J. M. (2021). Bioorthogonal chemistry. Nature Reviews Methods Primers, 1(1). https://doi.org/10.1038/s43586-021-00028-z
6. Science Communication Lab. (2010, May 2). Carolyn Bertozzi (UC Berkeley) Part 2: Imaging the Glycome [Video]. YouTube. https://www.youtube.com/watch?v=g17QmtZOyWc
7. Science Communication Lab. (2010a, March 24). Carolyn Bertozzi (UC Berkeley) Part 1: Chemical Glycobiology[Video]. YouTube: https://www.youtube.com/watch?v=WCbg-kOY_8E
8. Bertozzi, C. R., & Kiessling, A. L. L. (2001). Chemical glycobiology. Science, 291(5512), 2357–2364. https://doi.org/10.1126/science.1059820
9. Schauenburg, D., & Weil, T. (2025). Not so bioorthogonal chemistry. Journal of the American Chemical Society. https://doi.org/10.1021/jacs.4c15986
10. Knight, J. C., & Cornelissen, B. (2014, March 20). Bioorthogonal chemistry: implications for pretargeted nuclear (PET/SPECT) imaging and therapy. https://pmc.ncbi.nlm.nih.gov/articles/PMC3992206/
11. Saplakoglu, Y. (2022, October 5). Molecule-Building Innovators Win 2022 Chemistry Nobel Prize | Quanta Magazine. Quanta Magazine. https://www.quantamagazine.org/molecule-building-innovators-win-2022chemistry-nobel-prize-20221005/

IMAGE REFERENCES

1. Header Image: College of Chemistry, UC Berkeley. (2022). Chemistry Nobelist Carolyn Bertozzi on her years at UC Berkeley. https://chemistry.berkeley.edu/news/chemistry-nobelist-carolyn-bertozzi-years-at-ucberkeley
2. Figure 1: Bird, R. E., Lemmel, S. A., Yu, X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479 https://doi.org/10.1021/acs.bioconjchem.1c00461
3. Figure 2: Bird, R. E., Lemmel, S. A., Yu, X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479 https://doi.org/10.1021/acs.bioconjchem.1c00461
4. Figure 3: Bird, R. E., Lemmel, S. A., Yu,X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479 https://doi.org/10.1021/acs.bioconjchem.1c00461
5. Figure 4: Bird, R. E., Lemmel, S. A., Yu, X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479 https://doi.org/10.1021/acs.bioconjchem.1c00461
6. Figure 5: Bird, R. E., Lemmel, S. A., Yu, X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479 https://doi.org/10.1021/acs.bioconjchem.1c00461
7. Figure 6: Bird, R. E., Lemmel, S. A., Yu, X., & Zhou, Q. A. (2021). Bioorthogonal chemistry and its applications. Bioconjugate Chemistry, 32(12), 2457–2479 https://doi.org/10.1021/acs.bioconjchem.1c00461

December 6, 2025

Ruben Levya

2025, BSJ Features, Spring 2025

Features Department