Engineering a Silk Protein-Mediated Customizable Compartment for Modular Metabolic Synthesis

Area of Science:

• Synthetic biology
• Metabolic engineering
• Biotechnology

Background:

• Microbial cell factories are valuable for chemical synthesis but suffer from low efficiency due to enzyme dispersion and metabolite accumulation.
• Optimizing enzyme localization and interaction is crucial for improving metabolic pathway efficiency.

Purpose of the Study:

• To design and construct a synthetic cellular microcompartment for isolating enzyme reactions and optimizing modular metabolic synthesis.
• To enhance the production of 2 -fucosyllactose (2 -FL) using this novel microcompartment system.

Main Methods:

• Engineered spider silk proteins to form self-assembling protein condensates (microcompartments) within microbial cells.
• Recruited enzymes to these compartments using peptide interactions or direct fusion to improve catalytic efficiency.
• Implemented and optimized a de novo synthesis pathway for 2 -FL within the engineered microcompartments.

Main Results:

• Demonstrated the formation and function of synthetic microcompartments for enzyme aggregation.
• Achieved a significant increase in 2 -FL titer compared to free enzymes and modular-optimized pathways.
• Successfully aggregated the 2 -FL synthesis pathway within the synthetic compartment, creating multienzyme aggregates.

Conclusions:

• Synthetic cell microcompartments offer an effective strategy for enzyme aggregation and enhancing microbial metabolic pathway efficiency.
• This approach provides a novel avenue for improving the production of valuable chemicals and drugs using engineered microbes.