September 25, 2008

Also appeared in print Oct. 6, 2008, p. 8

Chemical Biology

Rapid Tagging Of Biomolecules

Cycloaddition never before used biologically could help assemble sophisticated probes

Carmen Drahl

By adapting a cycloaddition reaction that has never been used in biological systems, chemists have developed an exceptionally fast method for labeling biomolecules such as proteins. The advance could aid the study of dynamic processes in living systems.

FAST TRACK The cycloaddition of a trans-cyclooctene to a tetrazine gives off nitrogen gas and rapidly labels a protein in a dilute solution.

Biomolecules inside cells usually exist at low concentrations. To get a glimpse of cellular goings-on, researchers label biomolecules with fluorescent or other tags. The tagging must be fast to be useful in living systems, says University of Delaware chemist Joseph M. Fox.

Together with Melissa L. Blackman and Maksim Royzen, Fox has developed a new labeling reaction based on the cycloaddition of biomolecules derivatized with trans-cyclooctene to a nitrogen-rich heterocycle called a tetrazine. The reaction needs no catalyst and is hundreds of times faster than click reactions, an established tagging method. The added speed could allow researchers to assemble more complex probes inside cells than before. The reaction is also bioorthogonal, meaning that the tetrazine and cyclooctene don't undergo side reactions with other molecules in biological systems.

Fox's method stands out for its selectivity and speed, says chemical biologist Carolyn R. Bertozzi of the University of California, Berkeley. The reaction's kinetics are unprecedented: "perhaps the fastest reported for any bioorthogonal ligation to date," she says.

The labeling reaction is an inverse-electron-demand Diels-Alder reaction, in which an electron-rich cyclooctene dienophile reacts with an electron-poor diene, the tetrazine. Although fast reactions of this sort between tetrazines and cyclooctenes are known, the tetrazines used in those methods react with water and are therefore unsuitable for labeling in cells.

Fox's team tuned the tetrazine's reactivity by adding pyridine moieties and showed that the revamped tetrazine reacts quickly and selectively with trans-cyclooctene in cell lysates. Moreover, the team labeled biologically relevant concentrations of a cyclooctene-tagged protein with low concentrations of their tetrazine within minutes, a result suggesting the reaction could be applicable in cellular environments. They are now initiating collaborations to further explore biological uses.

Fox's work "is an elegant demonstration of how classic chemistry from the physical organic literature can be artfully retrofitted for modern applications," Bertozzi says.

Chemical & Engineering News

ISSN 0009-2347

Copyright © 2009 American Chemical Society