Phase-Separating Peptides Recruiting Aggregation-Induced Emission Fluorogen for Rapid E. coli Detection

Area of Science:

• Biomolecular engineering
• Biosensing technology
• Microbiology

Background:

• Biomolecular condensates are primarily used in drug delivery due to efficient molecular trapping.
• Their application in biosensing remains largely unexplored.
• Rapid and sensitive detection of bacteria like E. coli is crucial for public health.

Purpose of the Study:

• To develop a simple, rapid, and sensitive assay for detecting E. coli.
• To explore the use of phase-separating peptide condensates in biosensing.
• To leverage aggregation-induced emission (AIE) fluorogens for visual detection.

Main Methods:

• Designed phase-separating peptide condensates incorporating an E. coli outer membrane protease (OmpT) recognition site.
• Recruited an AIE-fluorogen into the peptide condensates, which fluoresces upon aggregation.
• Utilized OmpT's enzymatic activity to cleave peptides, disrupting condensate formation and quenching fluorescence.

Main Results:

• Demonstrated successful detection of OmpT activity using recombinant enzyme and subsequently with E. coli K-12.
• Achieved detection of 10⁸ CFU/mL E. coli K-12 within 2 hours in spiked water samples.
• Showed potential for detecting 1-10 CFU/mL with a pre-culture step, significantly faster than commercial kits.

Conclusions:

• The developed assay provides a rapid and visually detectable method for E. coli identification.
• The assay platform is adaptable for detecting other Gram-negative bacteria and diagnostically relevant proteases.
• Further optimization of peptide sequences can enhance detection limits and assay speed.