Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01

Halomonas bluephagenesis TD01 is one of the ideal chassis for low-cost industrial production based on “Next Generation Industrial Biotechnology,” yet the limited genetically regulatory parts such as transcriptional terminators, which are crucial for tuned regulations on gene expression, have hampere...

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Main Authors: Mengmeng Xu, Yue Chang, Yuyan Zhang, Weizhe Wang, Jingyi Hong, Jiping Zhao, Xiaoyun Lu, Dan Tan
Format: Article
Language:English
Published: Frontiers Media S.A. 2022-06-01
Series:Frontiers in Microbiology
Subjects:
Online Access:https://www.frontiersin.org/articles/10.3389/fmicb.2022.941306/full
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author Mengmeng Xu
Yue Chang
Yuyan Zhang
Weizhe Wang
Jingyi Hong
Jiping Zhao
Xiaoyun Lu
Dan Tan
author_facet Mengmeng Xu
Yue Chang
Yuyan Zhang
Weizhe Wang
Jingyi Hong
Jiping Zhao
Xiaoyun Lu
Dan Tan
author_sort Mengmeng Xu
collection DOAJ
description Halomonas bluephagenesis TD01 is one of the ideal chassis for low-cost industrial production based on “Next Generation Industrial Biotechnology,” yet the limited genetically regulatory parts such as transcriptional terminators, which are crucial for tuned regulations on gene expression, have hampered the engineering and applications of the strain. In this study, a series of intrinsic Rho-independent terminators were developed by either genome mining or rational design, and seven of them proved to exhibit higher efficiencies than the canonical strong T7 terminator, among which three terminators displayed high efficiencies over 90%. A preliminary modeling on the sequence-efficiency relationship of the terminators suggested that the poly U sequence regularity, the length and GC content of the stem, and the number and the size of hairpin loops remarkably affected the termination efficiency (TE). The rational and de novo designs of novel synthetic terminators based on the sequence-efficiency relationship and the “main contributor” engineering strategy proved to be effective, and fine-tuned polyhydroxylkanoates production was also achieved by the regulation of these native or synthetic terminators with different efficiencies. Furthermore, a perfectly positive correlation between the promoter activity and the TE was revealed in our study. The study enriches our knowledge of transcriptional termination via its sequence–strength relationship and enables the precise regulation of gene expression and PHA synthesis by intrinsic terminators, contributing to the extensive applications of H. bluephagenesis TD01 in the low-cost production of various chemicals.
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spelling doaj.art-545904abb185425b862e1283087484b52022-12-22T00:32:50ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2022-06-011310.3389/fmicb.2022.941306941306Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01Mengmeng XuYue ChangYuyan ZhangWeizhe WangJingyi HongJiping ZhaoXiaoyun LuDan TanHalomonas bluephagenesis TD01 is one of the ideal chassis for low-cost industrial production based on “Next Generation Industrial Biotechnology,” yet the limited genetically regulatory parts such as transcriptional terminators, which are crucial for tuned regulations on gene expression, have hampered the engineering and applications of the strain. In this study, a series of intrinsic Rho-independent terminators were developed by either genome mining or rational design, and seven of them proved to exhibit higher efficiencies than the canonical strong T7 terminator, among which three terminators displayed high efficiencies over 90%. A preliminary modeling on the sequence-efficiency relationship of the terminators suggested that the poly U sequence regularity, the length and GC content of the stem, and the number and the size of hairpin loops remarkably affected the termination efficiency (TE). The rational and de novo designs of novel synthetic terminators based on the sequence-efficiency relationship and the “main contributor” engineering strategy proved to be effective, and fine-tuned polyhydroxylkanoates production was also achieved by the regulation of these native or synthetic terminators with different efficiencies. Furthermore, a perfectly positive correlation between the promoter activity and the TE was revealed in our study. The study enriches our knowledge of transcriptional termination via its sequence–strength relationship and enables the precise regulation of gene expression and PHA synthesis by intrinsic terminators, contributing to the extensive applications of H. bluephagenesis TD01 in the low-cost production of various chemicals.https://www.frontiersin.org/articles/10.3389/fmicb.2022.941306/fullHalomonas bluephagenesisintrinsic terminatortermination efficiencyrational designpolyhydroxylkanoatesRNA-Seq
spellingShingle Mengmeng Xu
Yue Chang
Yuyan Zhang
Weizhe Wang
Jingyi Hong
Jiping Zhao
Xiaoyun Lu
Dan Tan
Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01
Frontiers in Microbiology
Halomonas bluephagenesis
intrinsic terminator
termination efficiency
rational design
polyhydroxylkanoates
RNA-Seq
title Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01
title_full Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01
title_fullStr Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01
title_full_unstemmed Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01
title_short Development and Application of Transcription Terminators for Polyhydroxylkanoates Production in Halophilic Halomonas bluephagenesis TD01
title_sort development and application of transcription terminators for polyhydroxylkanoates production in halophilic halomonas bluephagenesis td01
topic Halomonas bluephagenesis
intrinsic terminator
termination efficiency
rational design
polyhydroxylkanoates
RNA-Seq
url https://www.frontiersin.org/articles/10.3389/fmicb.2022.941306/full
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