351. Engineering the Outer Membrane Could Facilitate Better Bacterial Performance and Effectively Enhance Poly-3-Hydroxybutyrate Accumulation.
- Author
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Jianli Wang, Wenjian Ma, Yu Fang, Hailing Zhang, Hao Liang, Haili Liu, Tingwei Wang, Shangwei Chen, Jian Ji, and Xiaoyuan Wang
- Subjects
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ACETYLCOENZYME A , *POLY-beta-hydroxybutyrate , *GENETIC overexpression , *CELL morphology , *CELL size , *FLAGELLA (Microbiology) , *LIPOPOLYSACCHARIDES - Abstract
Poly-3-hydroxybutyrate (PHB) is an environmentally friendly polymer and can be produced in Escherichia coli cells after overexpression of the heterologous gene cluster phaCAB. The biosynthesis of the outer membrane (OM) consumes many nutrients and influences cell morphology. Here, we engineered the OM by disrupting all gene clusters relevant to the polysaccharide portion of lipopolysaccharide (LPS), colanic acid (CA), flagella, and/or fimbria in E. coli W3110. All these disruptions benefited PHB production. Especially, disrupting all these OM components increased the PHB content to 83.0 wt% (PHB content percentage of dry cell weight), while the wild-type control produced only 1.5 wt% PHB. The increase was mainly due to the LPS truncation to Kdo2 (3-deoxy-D-manno-octulosonic acid)-lipid A, which resulted in 82.0 wt% PHB with a 25-fold larger cell volume, and disrupting CA resulted in 57.8 wt% PHB. In addition, disrupting LPS facilitated advantageous fermentation features, including 69.1% less acetate, a 550% higher percentage of autoaggregated cells among the total culture cells, 69.1% less biofilm, and a higher broken cell ratio. Further detailed mechanism investigations showed that disrupting LPS caused global changes in envelope and cellular metabolism: (i) a sharp decrease in flagella, fimbria, and secretions; (ii) more elastic cells; (iii) much greater carbon flux toward acetyl coenzyme A (acetyl-CoA) and supply of cofactors, including NADP, NAD, and ATP; and (iv) a decrease in by-product acids but increase in g-aminobutyric acid by activating s E factor. Disrupting CA, flagella, and fimbria also improved the levels of acetyl-CoA and cofactors. The results indicate that engineering the OM is an effective strategy to enhance PHB production and highlight the applicability of OM engineering to increase microbial cell factory performance. [ABSTRACT FROM AUTHOR]
- Published
- 2021
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