Generation of an E. coli platform strain for improved sucrose utilization using adaptive laboratory evolution.

Area of Science:

• Microbiology
• Synthetic Biology
• Metabolic Engineering

Background:

• Sucrose is an abundant and inexpensive carbon source, but its utilization by Escherichia coli is limited.
• Existing sucrose-metabolizing E. coli strains exhibit slow growth or pathogenic traits, hindering industrial applications.

Purpose of the Study:

• To develop engineered Escherichia coli K-12 MG1655 and E. coli W platform strains with enhanced sucrose utilization capabilities.
• To identify genetic mutations conferring improved growth rates and sucrose uptake in E. coli.

Main Methods:

• Adaptive laboratory evolution (ALE) was employed to evolve engineered E. coli strains expressing sucrose utilization genes (cscB, cscK, cscA).
• Whole genome sequencing was used to identify genetic mutations in evolved strains.
• Reverse engineering was performed to validate the causal advantages of identified mutations.

Main Results:

• Evolved E. coli K-12 clones showed significant increases in growth (1.72-fold) and sucrose uptake (1.40-fold) on sugarcane juice.
• Evolved E. coli W clones exhibited a 1.4-fold increase in sucrose uptake rate.
• Key mutations were identified in global transcription regulators (rpoB, rpoC) and a pyrimidine biosynthetic pathway (pyrE) in K-12 strains, and in the cscR gene in W strains.

Conclusions:

• The developed E. coli K-12 and W platform strains possess enhanced sucrose utilization phenotypes.
• Specific genetic mutations identified are crucial for enabling efficient sucrose metabolism in E. coli.
• These engineered strains and mutation data serve as valuable resources for sucrose-based industrial bioproduction.