Condensation
of 2‑((Alkylthio)(aryl)methylene)malononitrile
with 1,2-Aminothiol as a Novel Bioorthogonal Reaction for Site-Specific
Protein Modification and Peptide Cyclization
posted on 2020-03-06, 17:05authored byXiaoli Zheng, Zhuoru Li, Wei Gao, Xiaoting Meng, Xuefei Li, Louis Y. P. Luk, Yibing Zhao, Yu-Hsuan Tsai, Chuanliu Wu
Site-specific
modification of peptides and proteins has wide applications
in probing and perturbing biological systems. Herein we report that
1,2-aminothiol can react rapidly, specifically and efficiently with
2-((alkylthio)(aryl)methylene)malononitrile (TAMM) under biocompatible
conditions. This reaction undergoes a unique mechanism involving thiol-vinyl
sulfide exchange, cyclization, and elimination of dicyanomethanide
to form 2-aryl-4,5-dihydrothiazole (ADT) as a stable product. An 1,2-aminothiol functionality
can be introduced into a peptide or a protein as an N-terminal cysteine or an unnatural amino acid. The bioorthogonality
of this reaction was demonstrated by site-specific labeling of not
only synthetic peptides and a purified recombinant protein but also
proteins on mammalian cells and phages. Unlike other reagents in bioorthogonal
reactions, the chemical and physical properties of TAMM can be easily
tuned. TAMM can also be applied to generate phage-based ADT-cyclic
peptide libraries without reducing phage infectivity. Using this approach,
we identified ADT-cyclic peptides with high affinity to different
protein targets, providing valuable tools for biological studies and
potential therapeutics. Furthermore, the mild reaction conditions
of TAMM condensation warrant its use with other bioorthogonal reactions
to simultaneously achieve multiple site-specific modifications.