Antimicrobial Mechanism of “Fatty Acyl Glutamic Acid” Biosurfactant against Escherichia coli O157:H7 and Listeria monocytogenes Involves Disruption of the Cytoplasmic Membrane

Abstract

We report the antimicrobial mechanism of fatty acyl glutamic acid (FA-glu) biosurfactant against the foodborne pathogens L. monocytogenes (Gram-positive bacteria) and E. coli O157:H7 (Gram-negative bacteria). Growth of both of the bacterium types was inhibited by 50% and 100% with 5 and 25 mg/L FA-glu in the bacterial broth medium, respectively. Bacterial inhibition involved rupturing and perforation of the cell envelopes respectively for E. coli and L. monocytogenes. Mass spectrometric and the nuclear magnetic resonance spectroscopic assessment of the cell-free broth of the FA-glu treated bacteria indicated the release of phospholipids (PLs) from the lipid bilayer (LBL) to the broth, which according to a model proposed by Helenius and Simons, suggested cell envelope damage resulting upon mixing of the aqueous-soluble FA-glu with the LBL PLs. Alterations in the LBL fluidity upon FA-glu and PL mixing, a plausible cause for cell envelope damage, were assessed in terms of the changes in the phase-transition temperature (ΔH 1/2 ) and the enthalpy (ΔH) for selected PLs representing these bacterium types. The PLs considered were 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) found in Gram-positive and Gram-negative bacteria and 1,2dimyristoyl-sn-glycero-3-phosphorylglycerol sodium salt (DMPG) and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) found predominately in L. monocytogenes and E. coli, respectively. Using a differential scanning calorimeter, the ΔH values for DPPE, DMPC, and DMPG upon mixing with FA-glu (% 12 mol/mol) were determined as −9 J/mol, −3 J/g, and −3 J/g respectively, with corresponding ΔT 1/2 values as ≈4 °C for DMPG, ≈1 °C for DPPE, and ≈2 °C for DMPC. Accordingly, we proposed that selective interactions of FA-glu with zwitterionic DMPC, than with anionic DMPG, of L. monocytogenes led to the formation of FA-glu-DMPC mixed micelle regions that served as defects for cell envelope perforation. Relatively less selective interactions for FA-glu with zwitterionic DPPE and DMPC of the E. coli resulted in an overall change in the LBL fluidity that led to cell envelope rupturing. Thus, alterations in the LBL fluidity due to FA-glu perturbation resulted in the rupturing/perforation of the cell envelope that inhibited bacterial growth.