Engineering the Outer Membrane Could Facilitate Better Bacterial Performance and Effectively Enhance...

Area of Science:

• Microbial biotechnology and synthetic biology.
• Metabolic engineering and cell factory optimization.
• Polymer biosynthesis and biomanufacturing.

Background:

• Poly-3-hydroxybutyrate (PHB) is an eco-friendly biopolymer produced in Escherichia coli.
• Overexpression of the phaCAB gene cluster facilitates PHB biosynthesis.
• Outer membrane (OM) composition impacts cellular metabolism and nutrient consumption.

Purpose of the Study:

• To engineer the E. coli outer membrane to enhance PHB production.
• To investigate the effects of disrupting OM components on PHB accumulation and fermentation characteristics.
• To elucidate the metabolic and cellular mechanisms underlying OM engineering benefits.

Main Methods:

• Genetic disruption of E. coli W3110 gene clusters involved in lipopolysaccharide (LPS), colanic acid (CA), flagella, and fimbria.
• Quantification of PHB content (wt% of dry cell weight).
• Analysis of fermentation parameters, cell morphology, and metabolic flux.

Main Results:

• Disrupting OM components significantly increased PHB production, reaching 83.0 wt% with complete OM disruption.
• LPS truncation to Kdo₂-lipid A was a key factor, yielding 82.0 wt% PHB and a 25-fold increase in cell volume.
• OM engineering improved fermentation by reducing acetate (69.1%), increasing autoaggregation (550%), decreasing biofilm (69.1%), and enhancing carbon flux to acetyl-CoA and cofactors.

Conclusions:

• Engineering the E. coli outer membrane is a highly effective strategy for enhancing PHB production.
• OM remodeling influences cellular metabolism, increasing precursor availability and cofactor supply for biopolymer synthesis.
• This approach offers a promising avenue for improving microbial cell factories for various bioproducts.