Single-Cell, Time-Resolved Antimicrobial Effects of a Highly Cationic, Random Nylon-3 Copolymer on Live XXXX .

@entity1 @entity231 @entity526
UNASSIGNED: Synthetic random copolymers based on the nylon-3 (b-peptide) backbone show promise as inexpensive antimicrobial agents resistant to proteolysis. We present a time-resolved observational study of the attack of a particular copolymer MM63:CHx37 on single, live @entity526 cells. The composition and chain length of MM63:CHx37 (63% cationic subunits, 37% hydrophobic subunits, 35-subunit average length) were optimized to enhance antibacterial activity while minimizing lysis of @entity1 red blood cells. For @entity526 cells that export GFP to the periplasm, we obtain alternating phase-contrast and green fluorescence images with a time resolution of 12 s over 60 min following initiation of copolymer flow. Within seconds, cells shrink and exhibit the same plasmolysis spaces that occur following abrupt external osmotic upshift. The osmoprotection machinery attempts to replenish cytoplasmic water, but recovery is interrupted by permeabilization of the @entity231 ( @entity231 ) to GFP. Evidently, the highly cationic copolymer and its counterions rapidly translocate across the outer membrane without permeabilizing it to GFP. The @entity231 permeabilization event is spatially localized. Cells whose @entity231 has been permeabilized never recover growth. The minimum inhibitory concentration (MIC) for cells lacking the osmolyte importer ProP is 4-fold smaller than for normal cells, suggesting that osmoprotection is an important survival strategy. In addition, at the time of @entity231 permeabilization, we observe evidence of oxidative stress. The MIC under anaerobic conditions is at least 8-fold larger than under aerobic conditions, further implicating oxidative damage as an important bacteriostatic effect. Once the copolymer reaches the periplasm, multiple growth-halting mechanisms proceed in parallel.
