Theranostic FRET Gate to Visualize and Quantify Bacterial Membrane Breaching

Abstract

Designing new antimicrobial-cum-probes to study real-time bacterial membrane breaching and concurrently developing inquisitorial image-based analytical tools is essential for the treatment of infectious diseases. An array of aggregation-induced emission (AIE) polymers (donor) consisting of neutral, anionic, and cationic charges were designed and employed as antimicrobial theranostic gatekeepers for the permeabilization of the peptidoglycan layer-adherable crystal violet (CV, acceptor). An AIE-active tetraphenylethylene (TPE)-tagged polycaprolactone biodegradable platform was chosen, and their self-assembled tiny amphiphilic nanoparticles were employed as a gatekeeper in the construction of bacterial membrane-reinforced fluorescent resonance energy transfer (FRET) probes. Electrostatic adhering of the cationic AIE polymer and subsequent gate opening aided fluorescent FRET probe activation on the membrane of Gram-negative bacteria, Escherichia coli. The selective photoexcitation energy transfer process in confocal microscopy experiments facilitated the building of a visualization-based FRET assay for the quantification of bactericidal activity. Nonantimicrobial AIE polymers (neutral and anionic) did not breach the bacterial membrane, resulting in no FRET signal. Detailed photophysical studies were done to establish the FRET probe mechanism, and a proof of concept was established.