CHAPTER 24: ²H Solid-State NMR Study of Peptide–Membrane Interactions in Intact Bacteria

A key aspect of the antibacterial activity of membrane-active peptides and molecules is their interaction with lipid bilayers. Thus solid-state NMR (SS-NMR) studies of antimicrobial peptides (AMPs) in complex with model membrane bilayers have provided a wealth of valuable information regarding their mechanism of action. However, such simplified systems only approximate the lipid bilayers found in the much more complex cell envelopes that AMPs encounter when they are functioning in vivo. Microbe cell envelopes contain a number of additional components, such as lipopolysaccharides, peptidoglycans, membrane proteins, and complex lipid compositions, that likely affect how AMPs function. Consequently, there is a need to bridge the gap between the model membrane experiments we rely upon to reveal details of AMP mechanism and the complex environment AMPs actually encounter in vivo. We have thus developed novel approaches to accomplish ²H SS-NMR experiments with intact bacteria. By manipulating the growth conditions of either wild-type or mutated Escherichia coli, it is possible to achieve high levels of lipid deuteration in the bacteria and thus study phospholipid–AMP interactions within the context of intact bacteria. The novel techniques have been applied to the antimicrobial peptide MSI-78 and antimicrobial marennine pigments in order to reveal aspects of their mechanisms not addressable by other types of studies.