Structural Disruptions of the Outer Membranes of Gram-Negative Bacteria by Rationally Designed Amphiphilic Antimicrobial Peptides

Abstract

Gram-negative bacteria are covered by both an inner cytoplasmic membrane (IM) and an outer membrane (OM). Antimicrobial peptides (AMPs) must first permeate through the OM and cell wall before attacking the IM to cause cytoplasmic leakage and kill the bacteria. The bacterial OM is an asymmetric bilayer with the outer leaflet primarily composed of lipopolysaccharides (LPSs) and the inner leaflet composed of phospholipids (PLs). Two cationic α-helical AMPs were designed to target Gram-negative bacteria, a full peptide G­(IIKK)3I-NH2 (G3), and a hydrophobic lipopeptide C8-G­(IIKK)2I-NH2 (C8G2, with C8 denoting the octanoyl chain). LPS dominates OM functions as the first line of defense against antibiotics, thereby reducing drug susceptibility. This work explores how the two AMPs interact with LPS through several carefully chosen OM models that facilitated measurements from solid-state nuclear magnetic resonance (ss-NMR), small-angle neutron scattering (SANS), and neutron reflectivity (NR). The results revealed that G3 molecules bound preferably to the LPS head region and functioned as bridge molecules to reassemble the dislocated lipids into bilayer stacks. In contrast, C8G2 lipopeptides could quickly penetrate into the central region of the OM to cause direct removal of some membrane lipids. Different structural disruptions implicated different antimicrobial efficacies from these AMPs. The demonstration of the structural features underlying different susceptibilities of the OM to AMPs offers a useful route for the future development of strain-specific AMPs against antimicrobial-resistant pathogens.

Disclosure of a Promising Lead to Tackle Complicated Skin and Skin Structure Infections: Antimicrobial and Antibiofilm Actions of Peptide PP4-3.1

“…Rational design, along with chemical synthesis, offers a multiplicity of peptide-based constructs with no match in Nature, enabling a fine tuning of their pharmacodynamic and pharmacokinetic properties, thereby improving their fitness to reach the clinics [40][41][42]. De novo design has been delivering very promising HDP in the past couple of years such as, e.g., short amphiphilic peptides possessing the general structure K n F m K n [43] and activity against antibiotic-susceptible strains of E. coli and S. aureus (2 ≤ MIC ≤ 32 mg/mL), or a structure based on IIKK repeat units conferring selective activity against antibioticsusceptible E. coli (8 ≤ MIC ≤ 16 mg/mL) due to the ability to specifically interact with the outer membrane of this bacterium [44]. Computer-aided rational design of HDP is widening the chemical space around antimicrobial peptides [45], which, together with growing knowledge on specific features of bacterial cell surfaces, allows the tailoring of biomimetic peptides able to, e.g., overcome the extracellular polysaccharide capsule of Klebsiella pneumoniae [46] or specifically interact with the mycolic acid-rich envelope of Mycobacterium tuberculosis, killing this bacterium [47].…”

Unravelling the structural complexity of protein–lipid interactions with neutron reflectometry

“…Examples of this are in studies on peptide disruption of lipid monolayers [23] and venom toxin disruption of supported lipid bilayers [22] . Recently NR has been used to examine the antimicrobial activity of new peptide based antibiotics [25][59][60] [61] and provide a precision understanding of the activity of natural antimicrobial peptides [21][62][63] [64] . Biochemically relevant protein-lipid interactions are another area where NR is able to provide unique insights.…”

Highly selective performance of rationally designed antimicrobial peptides based on ponericin-W1

“…1,2 Based on current trends it is predicted that by 2050 nearly 10 million people would die from antimicrobial resistant bacterial infections worldwide and the global economic losses associated with AMR would cost $ 100 trillion. [1][2][3][4] Therefore, the development of new antimicrobial agents which are capable of overcoming the AMR is of utmost importance. The emergence of antimicrobial peptides (AMPs) has provided an alternative approach to the treatment of resistant bacteria.…”

“…Modification of amino acids, multimerization of AMPs, replacing the cationic or hydrophobic amino acids, substitution of L-amino acids with D-amino acids, C and N terminal modifications and shortening the peptide sequence are some of the approaches to develop new AMPs. 4,6,14,18,19 The biological activities of the AMPs are directed by their amino acid composition, length, net charge, secondary structure, amphiphilicity and hydrophobicity. 20 Therefore, these parameters must be taken into account for designing novel antimicrobial peptides.…”