Data_Sheet_1_Effects of LPS Composition in Escherichia coli on Antibacterial Activity and Bacterial Uptake of Antisense Peptide-PNA Conjugates.docx

The physical and chemical properties of the outer membrane of Gram-negative bacteria including Escherichia coli have a significant impact on the antibacterial activity and uptake of antibiotics, including antimicrobial peptides and antisense peptide-peptide nucleic acid (PNA) conjugates. Using a defined subset of E. coli lipopolysaccharide (LPS) and envelope mutants, components of the LPS-core, which provide differential susceptibility toward a panel of bacterial penetrating peptide (BPP)-PNA conjugates, were identified. Deleting the outer core of the LPS and perturbing the inner core only sensitized the bacteria toward (KFF)3K-PNA conjugates, but not toward conjugates carrying arginine-based BPPs. Interestingly, the chemical composition of the outer LPS core as such, rather than overall hydrophobicity or surface charge, appears to determine the susceptibility to different BPP-PNA conjugates thereby clearly demonstrating the complexity and specificity of the interaction with the LPS/outer membrane. Notably, mutants with outer membrane changes conferring polymyxin resistance did not show resistance toward the BPP-PNA conjugates, thereby eliminating one possible route of resistance for these molecules. Finally, envelope weakening, through deletion of membrane proteins such as OmpA as well as some proteins previously identified as involved in cationic antimicrobial peptide uptake, did not significantly influence BPP-PNA conjugate activity.

革兰氏阴性菌，尤其是大肠杆菌的细胞外膜之物理与化学特性，对抗菌活性及抗生素（包括抗菌肽和反义肽-肽核酸（PNA）偶联物）的摄取具有显著影响。通过使用定义明确的E. coli脂多糖（LPS）亚群及其包膜突变体，识别了LPS核心的组分，这些组分对一系列细菌穿透肽（BPP）-PNA偶联物表现出不同的易感性。仅删除LPS的外核，扰动内核，则使细菌对（KFF）3K-PNA偶联物敏感，而对携带精氨酸基BPP的偶联物则无此效应。有趣的是，外LPS核心的化学组成，而非整体的疏水性和表面电荷，似乎决定了其对不同BPP-PNA偶联物的敏感性，从而清晰地展示了与LPS/细胞外膜相互作用的复杂性和特异性。值得注意的是，那些赋予多粘菌素耐药性的细胞外膜变化突变体，对BPP-PNA偶联物并不表现出耐药性，从而排除了这些分子耐药的可能途径。最终，通过删除如OmpA等膜蛋白以及一些先前已识别的参与阳离子抗菌肽摄取的蛋白，包膜的削弱并未显著影响BPP-PNA偶联物的活性。