Integrated base editing and microfluidics boost microbial lipid production from lignin

Area of Science:

• Biotechnology and Synthetic Biology
• Microbial Engineering
• Sustainable Chemistry

Background:

• Lignin is an abundant, renewable aromatic biopolymer with potential for sustainable chemical production.
• Efficient microbial conversion of lignin into valuable products remains a challenge due to its recalcitrant nature.

Purpose of the Study:

• To investigate the transcriptional response of Curvularia clavata J1 to alkali lignin (AL).
• To develop and apply a novel genome-wide base editing system for fungal strain improvement.
• To enhance lignin biotransformation capabilities for increased lipid and enzyme production.

Main Methods:

• Transcriptomic analysis to understand cellular responses to AL.
• Development of a copper-inducible MCM5-AID base editor for targeted mutagenesis.
• Ultrahigh-throughput screening using droplet microfluidics to identify improved mutants.
• Bioreactor cultivation to validate enhanced strain performance.

Main Results:

• AL induced upregulation of electron transport, lipid catabolism, and iron homeostasis in C. clavata J1.
• Engineered mutant M6 exhibited a 36% increase in lipid content (49% of dry cell weight) and a 75% increase in laccase activity (228.58 U/L).
• Validated performance in bioreactors showed 33% increased lipid (51% of DCW) and 87% increased laccase activity (240.43 U/L).
• Genome sequencing confirmed C-to-T mutations, and transcriptomics revealed enhanced biosynthesis pathways redirecting carbon flux towards oil-enriched single-cell protein.

Conclusions:

• The study presents an integrated framework for lignin biotransformation using engineered fungi.
• Synthetic biology and microfluidics integration enables precise fungal engineering for enhanced bioproduct formation.
• This approach offers a sustainable strategy for valorizing lignin into valuable biochemicals and biofuels.