Design of a programmable biosensor-CRISPRi genetic circuits for dynamic and autonomous dual-control of metabolic flux in Bacillus subtilis
File(s)
Published version
Author(s)
Type
Journal Article
Abstract
Dynamic regulation is an effective strategy for fine-tuning metabolic pathways in order to maximize target product synthesis. However, achieving dynamic and autonomous up- and down-regulation of the metabolic modules of interest simultaneously, still remains a great challenge. In this work, we created an autonomous dual-control (ADC) system, by combining CRISPRi-based NOT gates with novel biosensors of a key metabolite in the pathway of interest. By sensing the levels of the intermediate glucosamine-6-phosphate (GlcN6P) and self-adjusting the expression levels of the target genes accordingly with the GlcN6P biosensor and ADC system enabled feedback circuits, the metabolic flux towards the production of the high value nutraceutical N-acetylglucosamine (GlcNAc) could be balanced and optimized in Bacillus subtilis. As a result, the GlcNAc titer in a 15-l fed-batch bioreactor increased from 59.9 g/l to 97.1 g/l with acetoin production and 81.7 g/l to 131.6 g/l without acetoin production, indicating the robustness and stability of the synthetic circuits in a large bioreactor system. Remarkably, this self-regulatory methodology does not require any external level of control such as the use of inducer molecules or switching fermentation/environmental conditions. Moreover, the proposed programmable genetic circuits may be expanded to engineer other microbial cells and metabolic pathways.
Date Issued
2020-01-24
Date Acceptance
2019-11-16
Citation
Nucleic Acids Research, 2020, 48 (2), pp.996-1009
ISSN
0305-1048
Publisher
Oxford University Press
Start Page
996
End Page
1009
Journal / Book Title
Nucleic Acids Research
Volume
48
Issue
2
Copyright Statement
© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
Identifier
https://www.ncbi.nlm.nih.gov/pubmed/31799627
PII: 5651338
Subjects
05 Environmental Sciences
06 Biological Sciences
08 Information and Computing Sciences
Developmental Biology
Publication Status
Published
Coverage Spatial
England
Date Publish Online
2019-12-04