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Site‐Specific N‐Terminal Labeling of Peptides and Proteins using Butelase 1 and Thiodepsipeptide
Author(s) -
Nguyen Giang K. T.,
Cao Yuan,
Wang Wei,
Liu Chuan Fa,
Tam James P.
Publication year - 2015
Publication title -
angewandte chemie
Language(s) - English
Resource type - Journals
eISSN - 1521-3757
pISSN - 0044-8249
DOI - 10.1002/ange.201506810
Subject(s) - dna ligase , chemistry , nucleophile , native chemical ligation , ubiquitin ligase , chemical ligation , combinatorial chemistry , stereochemistry , depsipeptide , functional group , intramolecular force , ubiquitin , cysteine , peptide , biochemistry , organic chemistry , enzyme , polymer , gene , catalysis
An efficient ligase with exquisite site‐specificity is highly desirable for protein modification. Recently, we discovered the fastest known ligase called butelase 1 from Clitoria ternatea for intramolecular cyclization. For intermolecular ligation, butelase 1 requires an excess amount of a substrate to suppress the reverse reaction, a feature similar to other ligases. Herein, we describe the use of thiodepsipeptide substrates with a thiol as a leaving group and an unacceptable nucleophile to render the butelase‐mediated ligation reactions irreversible and in high yields. Butelase 1 also accepted depsipeptides as substrates, but unlike a thiodesipeptide, the desipeptide ligation was partially reversible as butelase 1 can tolerate an alcohol group as a poor nucleophile. The thiodesipeptide method was successfully applied in N‐terminal labeling of ubiquitin and green fluorescent protein using substrates with or without a biotin group in high yields.