(587b) Preferential Binding of Bacterial Membrane Lipids Influences the Free Energy of Membrane Binding and Refolding of the Antimicrobial Peptide CM15

With rising antimicrobial resistance, antimicrobial peptides (AMPs) are being explored as alternate therapeutics for bacterial infections. Despite extensive investigations on bacteria and model membrane systems, the underlying molecular mechanism of AMP activity is incompletely understood. It is known that CM15, an intrinsically disordered synthetic AMP, tends to fold into a helical state in the membrane environment; however, the transition mechanism from an unfolded state in solution to the helical state in the membrane is yet to be elucidated. Constructing the free energy landscape for peptide insertion in the membrane is challenging due to the associated secondary structure changes accompanying membrane binding and insertion, which require enhanced sampling molecular dynamics simulation techniques. Using a recently developed finite temperature string method with a two-dimensional collective variable space that accounts for the peptide position and the helical content, we study the CM15 insertion mechanism into the inner membrane of E. coli containing 5% cardiolipin. Our analysis reveals that the transition occurs in three main steps: membrane binding and insertion of unfolded CM15, folding of CM15 under the lipid headgroups to a membrane-parallel helical state, followed by the reorientation of the folded peptide to a partially tilted membrane inserted state. The largest free energy barrier of ~30 kJ/mol is associated with the insertion of the unfolded peptide into the phospholipid headgroups, wherein the largest dehydration of the peptide is observed. We identify that the cardiolipin interaction with the N-terminus of the CM15 plays a crucial role in influencing membrane insertion. Our study provides several novel insights into complex membrane association and refolding pathways associated with CM15 that can potentially allow for rational synthetic AMP design protocols.