Bacterial membrane pertubation by native versus dry-heated lysozyme

Antimicrobial drug resistance causes public health problems worldwide and stimulates research for novel antimicrobials.1 Particular attention is given to antimicrobial molecules that limit drug resistance development and have a large spectrum. Antimicrobial peptides and proteins are thus good candidates acting on the bacterial cell wall because of their physico-chemical properties 2These antimicrobial peptides and proteins can thus perturb the bacterial cell envelope, enter the cell, and/or act on intracellular targets. 2,3 Hen egg white lysozyme is largely studied and widely known for its muramidase activity against the peptidoglycan of Gram-positive bacteria. 4 However, lysozyme is also described as being active against some Gram-negative bacteria, especially because it permeabilizes the bacterial membranes. 4,5 But few is known about thisactivity, and the consequences remain quite limited. Since dry-heated lysozyme (DHL) is more positively charged, more hydrophobic and more tensio-active than native lysozyme (NL) 6 , dry-heating could be a way to increase lysozyme activity against the bacterial membranes, and especially against Gram-negative bacteria.To investigate the membrane activity of both NL and DHL, membrane permeability measurements using a mutantE. coli ML-35p and measurements of membrane potential perturbation were performed. These experiments revealed that NL and DHL induce different consequences on the bacterial membranes. NL permeabilizes both outer (OM) and inner (IM) membranes. Pore formation could be observed for OM, but not for IM. Moreover, the membrane potential of IM is modified in the presence of NL. DHL induces an overall stronger disturbance than NL, by giving rise to the formation of more and/or larger pores in OM, by permeabilizing more rapidly IM, and by leading to a stronger disturbance of membrane potentia