Translocation of the thioesterase domain for the redesign of plipastatin synthetase

Non-ribosomal peptide synthetases (NRPSs) are large enzymatic complexes that catalyse the synthesis of biologically active peptides in microorganisms. Genetic engineering has recently been applied to reprogram NRPSs to produce lipopeptides with a new sequence. The carboxyl-terminal thioesterase (TE)...

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Main Authors:	Gao, Ling, Liu, Hongxia, Ma, Zhi, Han, Jinzhi, Lu, Zhaoxin, Dai, Chen, Lv, Fengxia, Bie, Xiaomei
Format:	Online
Language:	English
Published:	Nature Publishing Group 2016
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5180189/

id	pubmed-5180189
recordtype	oai_dc
spelling	pubmed-51801892016-12-29 Translocation of the thioesterase domain for the redesign of plipastatin synthetase Gao, Ling Liu, Hongxia Ma, Zhi Han, Jinzhi Lu, Zhaoxin Dai, Chen Lv, Fengxia Bie, Xiaomei Article Non-ribosomal peptide synthetases (NRPSs) are large enzymatic complexes that catalyse the synthesis of biologically active peptides in microorganisms. Genetic engineering has recently been applied to reprogram NRPSs to produce lipopeptides with a new sequence. The carboxyl-terminal thioesterase (TE) domains from NRPSs catalyse cleavage products by hydrolysis or complex macrocyclization. In this study, we modified plipastatin synthetase by moving the intrinsic TE region to the end of the internal thiolation (T) domains, thus generating Bacillus subtilis strains that could produce new truncated cyclic or linear peptides of the predicted sequence, which further provided an important insight into the regioselectivity of plipastatin TE. The TE was capable of recognizing and catalysing the lactone formation between L-Try3 with the last few residues L-Pro7 and L-Gln8 at the C-terminus. Additionally, the unmatched linkers connecting the TE region and T domain resulted in nonproduction strains, suggesting that the native T–TE linker is necessary and sufficient for the TE domain to release the products from the hybrid enzymes. This is the first report to demonstrate truncated cyclic lipopeptides production and module skipping by simply moving the TE domain forward in an NRPS system. Nature Publishing Group 2016-12-23 /pmc/articles/PMC5180189/ /pubmed/28009004 http://dx.doi.org/10.1038/srep38467 Text en Copyright © 2016, The Author(s) http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Gao, Ling Liu, Hongxia Ma, Zhi Han, Jinzhi Lu, Zhaoxin Dai, Chen Lv, Fengxia Bie, Xiaomei
spellingShingle	Gao, Ling Liu, Hongxia Ma, Zhi Han, Jinzhi Lu, Zhaoxin Dai, Chen Lv, Fengxia Bie, Xiaomei Translocation of the thioesterase domain for the redesign of plipastatin synthetase
author_facet	Gao, Ling Liu, Hongxia Ma, Zhi Han, Jinzhi Lu, Zhaoxin Dai, Chen Lv, Fengxia Bie, Xiaomei
author_sort	Gao, Ling
title	Translocation of the thioesterase domain for the redesign of plipastatin synthetase
title_short	Translocation of the thioesterase domain for the redesign of plipastatin synthetase
title_full	Translocation of the thioesterase domain for the redesign of plipastatin synthetase
title_fullStr	Translocation of the thioesterase domain for the redesign of plipastatin synthetase
title_full_unstemmed	Translocation of the thioesterase domain for the redesign of plipastatin synthetase
title_sort	translocation of the thioesterase domain for the redesign of plipastatin synthetase
description	Non-ribosomal peptide synthetases (NRPSs) are large enzymatic complexes that catalyse the synthesis of biologically active peptides in microorganisms. Genetic engineering has recently been applied to reprogram NRPSs to produce lipopeptides with a new sequence. The carboxyl-terminal thioesterase (TE) domains from NRPSs catalyse cleavage products by hydrolysis or complex macrocyclization. In this study, we modified plipastatin synthetase by moving the intrinsic TE region to the end of the internal thiolation (T) domains, thus generating Bacillus subtilis strains that could produce new truncated cyclic or linear peptides of the predicted sequence, which further provided an important insight into the regioselectivity of plipastatin TE. The TE was capable of recognizing and catalysing the lactone formation between L-Try3 with the last few residues L-Pro7 and L-Gln8 at the C-terminus. Additionally, the unmatched linkers connecting the TE region and T domain resulted in nonproduction strains, suggesting that the native T–TE linker is necessary and sufficient for the TE domain to release the products from the hybrid enzymes. This is the first report to demonstrate truncated cyclic lipopeptides production and module skipping by simply moving the TE domain forward in an NRPS system.
publisher	Nature Publishing Group
publishDate	2016
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5180189/
_version_	1613814568959606784

Translocation of the thioesterase domain for the redesign of plipastatin synthetase

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