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. 2022 Nov 25;28(66):e202201843.
doi: 10.1002/chem.202201843. Epub 2022 Sep 19.

Fast Cysteine Bioconjugation Chemistry

Fa-Jie Chen  1 , Jianmin Gao  1
Affiliations

Fast Cysteine Bioconjugation Chemistry

Fa-Jie Chen et al. Chemistry. .

Abstract

Cysteine bioconjugation serves as a powerful tool in biological research and has been widely used for chemical modification of proteins, constructing antibody-drug conjugates, and enabling cell imaging studies. Cysteine conjugation reactions with fast kinetics and exquisite selectivity have been under heavy pursuit as they would allow clean protein modification with just stoichiometric amounts of reagents, which minimizes side reactions, simplifies purification and broadens functional group tolerance. In this concept, we summarize the recent advances in fast cysteine bioconjugation, and discuss the mechanism and chemical principles that underlie the high efficiencies of the newly developed cysteine reactive reagents.

Keywords: N-terminal cysteine; cysteine bioconjugation; fast kinetics; protein modification; reaction rate constant.

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Figures

Figure 1.
Figure 1.
Cysteine bioconjugation and some commonly used reagents.
Figure 2.
Figure 2.
Developing fast cysteine bioconjugation chemistries. (A) Illustration of fast cysteine conjugation on proteins and cells. E: electrophile. k2: secondary rate constant. SH: thiol group. (B) General considerations in addition to reaction rate.
Figure 3.
Figure 3.
Overview of reaction rates of fast cysteine conjugation protocols. SPAAC: strain-promoted alkyne−azide cycloaddition.
Figure 4.
Figure 4.
Fast bioconjugation chemistries of internal cysteines
Figure 5.
Figure 5.
Activated Michael acceptors for fast cysteine bioconjugation. aObserved reaction rate constant for N-Boc-Cys-OMe. bObserved reaction rate constant for N-Ac-Cys-NH2. ΔG≠:Gibbs free energy of activation
Figure 6.
Figure 6.
Activated heteroaromatic compounds for fast cysteine bioconjugation. Observed reaction rate constant for N-Ac-Cys-OMe.
Figure 7.
Figure 7.
Cation-activated electrophiles for fast cysteine bioconjugation
Figure 8.
Figure 8.
Strain-releasing reagents for fast cysteine bioconjugation
Figure 9.
Figure 9.
Hypervalent iodine reagents for fast cysteine bioconjugation. Observed reaction rate constant for Ellman’s reagent 2-nitro-5-thiobenzoic acid anion (TNB2-).
Figure 10.
Figure 10.
Fast bioconjugation chemistries of N-terminal cysteines
Figure 11.
Figure 11.
Activated aldehyde for fast N-terminal cysteine bioconjugation
Figure 12.
Figure 12.
Activated nitriles for fast N-terminal cysteine conjugation.
Figure 13.
Figure 13.
Activated Michael acceptors for fast N-terminal cysteine bioconjugation
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