How does macolacin evade colistin resistance conferred by lipid A modification

Abstract

Antimicrobial resistance (AMR) is a major global health threat, with estimates forecasting 10 million annual AMR-induced deaths by 2050. Antimicrobial peptides (AMP) are molecules that protect hosts from potential pathogens and are produced by most living organisms ranging from humans, animals, and plants to bacteria and fungi. AMPs also have the potential to combat multi-drug resistant (MDR) and pan-drug resistant (PDR) bacteria. One example is the polymyxin family of AMPs, which includes polymyxin B (PMB) and E (colistin), our last-resort treatment option against MDR Gram-negative bacterial infections. Colistin translocates through the bacterial outer membrane (OM) via self-promoted uptake, starting with the displacement of divalent cations that stabilise the bacterial OM, to reach the cytoplasmic membrane (CM), where polymyxin-membrane interaction leads to membrane permeabilization and cell death. However, in 2015, the discovery of a colistin resistance gene capable of horizontal gene transfer that jeopardizes colistin’s efficacy, mcr-1 (mobile colistin resistance 1), was reported. mcr-1 encodes a phosphoethanolamine (PEtN) transferase that adds a PEtN to the lipid A portion of the OM lipopolysaccharides, disrupting colistin binding to lipid A and the subsequent displacement of divalent cations that typically precedes membrane disruption. Recently, natural analogues of colistin, macolacin, and biphenyl-macolacin that displayed potent activity against mcr-1-mediated colistin-resistant bacteria were discovered. While the experimental efficacies of (biphenyl) macolacin were tested, their mechanism behind overcoming mcr-1-mediated colistin resistance remains unknown. Similarly, detailed and atomistic knowledge regarding the mechanism of action of polymyxins and mcr-1-mediated resistance is lacking. Therefore, the present project aimed to characterize these mechanisms using molecular dynamics (MD) simulations to model the interaction of polymyxin and (biphenyl) macolacin with colistin-susceptible (CSM) and resistant bacterial membranes (CRM). Results revealed two main findings. First, the PEtN in the CRM cross-links with neighbouring phosphate headgroups to stabilize the membrane through Coulombic interactions and hydrogen bonding, reducing the CRM’s reliance on magnesium ions for membrane stabilization. Secondly, deeply penetrating peptides exhibit an ‘unfolded’ structure, and the lack of the unfolding of the peptide was the main reason there was only one deeply penetrating peptide against the CRM. Through MD simulations, the present project contributes to understanding mcr-1 mediated colistin resistance and polymyxins’ mechanism of action and provides insight into the design of more effective AMPs. The limitations of the present thesis will hopefully contribute to better designs of polymyxin MD simulations.