Site‐Specific N‐Terminal Labeling of Peptides and Proteins using Butelase 1 and Thiodepsipeptide. (13th November 2015)

• Record Type: Journal Article
• Authors: Nguyen, Giang K. T.; Cao, Yuan; Wang, Wei; Liu, Chuan Fa; Tam, James P.
• Is Part Of: Angewandte Chemie. Volume 127:Number 52(2015)
• Page Start: 15920
• Page End: 15924
• Publication Date: 2015-11-13
• Subjects: Butelase 1 -- Ligationen -- Markierung -- Thiodepsipeptide; Chemistry -- Periodicals; 540
• DOI: 10.1002/ange.201506810
• Languages: English
• ISSNs: 0044-8249

Abstract

Abstract: 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. Abstract : Eine irreversible Ligation mit einem Thiodepsipeptid gelingt in >95 % Ausbeute mit einem minimalen Überschuss des Substrats und geringen Mengen an katalytisch wirkender Butelase 1. Mit dieser Methode wurde eine funktionelle Markierung („Tag"︁) an Ubiquitin und Grün fluoreszierendes Protein angeknüpft.