Optimizing E. coli as a formatotrophic platform for bioproduction via the reductive glycine pathway

Area of Science:

• Synthetic biology
• Microbial biotechnology
• Metabolic engineering

Background:

• Microbial C1 fixation offers a sustainable circular economy pathway.
• Engineered microorganisms can fix C1 substrates, but C1-based bioproduction is underexplored.
• Formatotrophic Escherichia coli strains have been developed for C1 substrate utilization.

Purpose of the Study:

• Optimize a previously engineered formatotrophic Escherichia coli strain for enhanced growth and bioproduction.
• Investigate the genetic basis for improved growth performance.
• Demonstrate the strain's potential for industrial applications and formate-based bioeconomy.

Main Methods:

• Adaptive laboratory evolution (ALE) for strain optimization.
• Genome sequencing and analysis to identify genetic modifications.
• Reverse engineering to validate findings.
• Bioreactor fed-batch cultivation for scale-up assessment.
• Metabolic engineering for lactate production from formate.

Main Results:

• ALE significantly enhanced biomass yield (3.3 g-CDW/mol-formate) and reduced doubling time (6 h) in formatotrophic E. coli.
• Acetate metabolism manipulation was identified as key to improved growth, confirmed by reverse engineering.
• The optimized strain reached an OD600 of 22 in bioreactor experiments.
• Engineered lactate production from formate achieved 1.2 mM (10% theoretical maximum).

Conclusions:

• Optimized formatotrophic E. coli exhibits robust growth and high biomass yield, suitable for industrial bioprocesses.
• The study provides the first proof-of-concept for using the reductive glycine pathway for bioproduction from formate.
• This work advances the development of a sustainable, formate-based bioeconomy.