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Antimicrobial resistance among Gram-negative bacteria is a growing problem, fueled by the paucity of new antibiotics that target these microorganisms. One novel family of macrocyclic β-hairpin-shaped peptidomimetics was recently shown to act specifically against Pseudomonas spp. by a novel mechanism of action, targeting the outer membrane protein LptD, which mediates lipopolysaccharide transport to the cell surface during outer membrane biogenesis. Here we explore the mode of binding of one of these β-hairpin peptidomimetics to LptD in Pseudomonas aeruginosa , by examining the effects on antimicrobial activity following N-methylation of individual peptide bonds. An N -methyl scan of the cyclic peptide revealed that residues on both sides of the β-hairpin structure at a non-hydrogen bonding position likely mediate hydrogen-bonding interactions with the target LptD. Structural analyses by NMR spectroscopy further reinforce the conclusion that the folded β-hairpin structure of the peptidomimetic is critical for binding to the target LptD. Finally, new NMe analogues with potent activity have been identified, which opens new avenues for optimization in this family of antimicrobial peptides. [ABSTRACT FROM AUTHOR]

Abstract: We report structural studies in aqueous solution on backbone cyclic peptides that possess potent antimicrobial activity specifically against Pseudomonas sp. The peptides target the β-barrel outer membrane protein LptD, which plays an essential role in lipopolysaccharide transport to the outer membrane. The peptide L27-11 contains a 12-residue loop (T1W2L3K4K5R6R7W8K9K10A11K12) linked to a DPro–LPro template. Two related peptides were also studied, one with various Lys to ornithine or diaminobutyric acid substitutions as well as a DLys6 (called LB-01), and another containing the same loop sequence, but linked to an LPro–DPro template (called LB-02). NMR studies and MD simulations show that L27-11 and LB-01 adopt β-hairpin structures in solution. In contrast, LB-02 is more flexible and importantly, adopts a wide variety of different backbone conformations, but not β-hairpin conformations. L27-11 and LB-01 show antimicrobial activity in the nanomolar range against Pseudomonas aeruginosa , whereas LB-02 is essentially inactive. Thus the β-hairpin structure of the peptide is important for antimicrobial activity. An alanine scan of L27-11 revealed that tryptophan side chains (W2/W8) displayed on opposite faces of the β-hairpin represent key groups contributing to antimicrobial activity. [Copyright &y& Elsevier]