Off-Pathway-Sensitive Protein-Splicing Screening Based on a Toxin/Antitoxin System

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http://hdl.handle.net/10138/305079

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Beyer , H M & Iwai , H 2019 , ' Off-Pathway-Sensitive Protein-Splicing Screening Based on a Toxin/Antitoxin System ' , ChemBioChem : a European journal of chemical biology , vol. 20 , no. 15 , pp. 1933-1938 . https://doi.org/10.1002/cbic.201900139

Title: Off-Pathway-Sensitive Protein-Splicing Screening Based on a Toxin/Antitoxin System
Author: Beyer, Hannes M.; Iwai, Hideo
Contributor: University of Helsinki, Institute of Biotechnology
University of Helsinki, Hideo Iwai / Principal Investigator
Date: 2019-08-01
Language: eng
Number of pages: 6
Belongs to series: ChemBioChem : a European journal of chemical biology
ISSN: 1439-4227
URI: http://hdl.handle.net/10138/305079
Abstract: Protein-splicing domains are frequently used engineering tools that find application in the in vivo and in vitro ligation of protein domains. Directed evolution is among the most promising technologies used to advance this technology. However, the available screening systems for protein-splicing activity are associated with bottlenecks such as the selection of pseudo-positive clones arising from off-pathway reaction products or fragment complementation. Herein, we report a stringent screening method for protein-splicing activity in cis and trans, that exclusively selects productively splicing domains. By fusing splicing domains to an intrinsically disordered region of the antidote from the Escherichia coli CcdA/CcdB type II toxin/antitoxin system, we linked protein splicing to cell survival. The screen allows selecting novel cis- and trans-splicing inteins catalyzing productive highly efficient protein splicing, for example, from directed-evolution approaches or the natural intein sequence space.
Subject: antitoxins
intein
protein splicing
selection screen
toxins
NPU DNAE INTEIN
SPLIT INTEINS
HIGHLY EFFICIENT
DIRECTED EVOLUTION
POSITIVE-SELECTION
STRUCTURAL BASIS
CCDB
PURIFICATION
GYRASE
DESIGN
1182 Biochemistry, cell and molecular biology
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