Investigating Daptomycin-Membrane Interactions Using Native MS and Fast Photochemical Oxidation of Peptides in Nanodiscs

16 February 2023, Version 2
This content is a preprint and has not undergone peer review at the time of posting.


Daptomycin is a cyclic lipopeptide antibiotic that targets the lipid membrane of Gram-positive bacteria. Membrane fluidity and charge can affect daptomycin activity, but its mechanisms are poorly understood because it is challenging to study daptomycin interactions within lipid bilayers. Here, we combined native mass spectrometry (MS) and fast photochemical oxidation of peptides (FPOP) to study daptomycin-membrane interactions with different lipid bilayer nanodiscs. Native MS suggests that daptomycin incorporates randomly and does not prefer any specific oligomeric states when integrated into bilayers. FPOP reveals significant protection in most bilayer environments. Combining the native MS and FPOP results, we observed that stronger membrane interactions are formed with more rigid membranes, and pore formation may occur in more fluid membranes to expose daptomycin to FPOP oxidation. Electrophysiology measurements further sup-ported the observation of polydisperse pore complexes from the mass spectrometry data. Together, these results demonstrate the complementarity of native MS, FPOP, and membrane conductance experiments to shed light on how antibiotic peptides interact with and within lipid membranes.


Mass Spectrometry
Native MS

Supplementary materials

Supporting Information
Supplemental Methods, Figures, Tables, and References


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