Finding the Right (Bioorthogonal) Chemistry

Abstract

Bioorthogonal chemistries can be used to tag diverse classes of biomolecules in cells and other complex environments. With over 20 unique transformations now available, though, selecting an appropriate reaction for a given experiment is challenging. In this article, we compare and contrast the most common classes of bioorthogonal chemistries and provide a framework for matching the reactions with downstream applications. We also discuss ongoing efforts to identify novel biocompatible reactions and methods to control their reactivity. The continued expansion of the bioorthogonal toolkit will provide new insights into biomolecule networks and functions and thus refine our understanding of living systems.

Strategies for cell membrane functionalization

“…15,45 Metabolic labeling and bio-orthogonal strategies still suffer from limitations associated with tightly regulated biosynthetic pathways (more of an issue for amino acids than glycans), interference from specific metabolic pathways (a particular issue with ketone labeling), cytotoxicity arising from certain mediators (such as copper ions in certain azide-alkyne reactions) and the necessity to include a compatible, ''clickable'' functional group on the secondary reactive species. 53 Most of these issues can be overcome with careful experimental design, and metabolic labeling coupled with bio-orthogonal bioconjugate chemistry remains an elegant approach to cell Metabolic labeling can be used to present reactive groups that can bind a secondary species (yellow). This is usually mediated by orthogonal click chemistry, in this example, an alkynated secondary species is bound to a cell metabolically labeled with azide groups.…”

Dual Strain-Promoted Alkyne–Nitrone Cycloadditions for Simultaneous Labeling of Bacterial Peptidoglycans

“…[18][19][20][21]23 However, combinations of different reactions can increase systematic error due to potential side reactions or variations in reagent stabilities. 13 Dual labeling with only nitrones and strained alkynes via the strain-promoted alkyne nitrone cycloaddition (SPANC) has potential to avoid such limitations, due to the unique tunability and increased stability of the endocyclic nitrone group (Scheme 1). 25,26 Stereoelectronically tuned alkyne−nitrone pairs display potential for promising selectivity while maintaining reactivity, 25 which suggests similar pairs could allow for detection of multiple biomolecular targets.…”

The potential of bioorthogonal chemistry for correlative light and electron microscopy: a call to arms

“…Its mechanism [7] relies on the introduction of a small abiotic chemical group (one that is non-reactive with other chemical functionality found in the cell) into a biomolecule of interest which can be specifically reacted with a detection moiety using a so-called bioorthogonal chemical reaction, a reaction of the tag with a detectable group that is essentially background-free in biological systems (Fig. 1a) [8].…”

“…Currently, a wide-ranging chemical toolkit is available of both tags for incorporation into biomolecules and reactions for subsequent labelling of these tags [10]. The choice of tag and modification chemistry can therefore be optimized and tailored for the specific biological hypothesis [7].…”

“…To the best of our knowledge, bioorthogonal reactions have not been combined with CLEM imaging up to now. This is surprising, since fluorescent imaging of bioorthogonal tags has become a commonplace over the last decade and a half [7]. In this review, we will highlight some of the inroads that have been made towards the CLEM imaging of bioorthogonal reactions.…”