Sugrue Ivan, Ade Carolin, O'Connor Paula M, Daniel Jan-Martin, Innocenti Paolo, Kirsch Nico, Martin Nathaniel I, Weindl Günther, Hill Colin, Schneider Tanja, Paul Ross R
APC Microbiome Ireland, University College Cork, Cork, Ireland.
School of Microbiology, University College Cork, Cork, Ireland.
NPJ Antimicrob Resist. 2025 Jul 22;3(1):66. doi: 10.1038/s44259-025-00135-x.
Antimicrobial peptides are defense molecules found across all domains of life holding promise for developing therapies against drug-resistant pathogens. Actifensin, from Actinomyces ruminicola DPC7226, exhibits potent activity against gram-positive bacteria and shares structural similarities with eukaryotic defensins. This study characterized actifensin's mechanism of action and therapeutic potential. The findings revealed that actifensin inhibits peptidoglycan synthesis by binding lipid II (Kd = 30 ± 20 nM). Unlike defensins, it also binds lipid I (Kd = 24 ± 27 nM) without significant difference, suggesting the N-acetyl glucosamine moiety of lipid II is not required for complexation. Membrane disruption was not observed with DiSC(5) fluorescence, or synthetic unilamellar liposomes, indicating indirect cell death via cell wall weakening, visualised by phase contrast microscopy. Actifensin showed no haemolytic activity or toxicity up to 128 µg/ml in human erythrocytes and Hep G2 cells. The peptide was not immunogenic, demonstrating no induction of LDH release in PBMCs or any effect on TLR-mediated signalling. Structural motif analysis identified actifensin as part of a conserved trans-kingdom defensin subfamily, GXGCP, distinct from XTCD peptides in more recently evolved arthropods. These findings emphasise the conserved structure-function relationship of antimicrobials across kingdoms, suggesting a shared evolutionary history of defensins and highlight the therapeutic potential for them or their variants.
抗菌肽是在生命的所有领域中发现的防御分子，有望开发出针对耐药病原体的疗法。来自反刍放线菌DPC7226的放线杀菌素对革兰氏阳性菌具有强大活性，并且与真核防御素具有结构相似性。本研究对放线杀菌素的作用机制和治疗潜力进行了表征。研究结果表明，放线杀菌素通过结合脂质II（解离常数Kd = 30±20 nM）来抑制肽聚糖合成。与防御素不同的是，它还能结合脂质I（Kd = 24±27 nM），且无显著差异，这表明脂质II的N-乙酰葡糖胺部分并非复合物形成所必需。用DiSC(5)荧光或合成单层脂质体未观察到膜破坏现象，表明通过相差显微镜观察到的细胞壁弱化导致间接细胞死亡。在人红细胞和Hep G2细胞中，放线杀菌素在高达128 μg/ml时未表现出溶血活性或毒性。该肽无免疫原性，在PBMC中未诱导LDH释放，对TLR介导的信号传导也无任何影响。结构基序分析确定放线杀菌素是保守的跨界防御素亚家族GXGCP的一部分，与最近进化的节肢动物中的XTCD肽不同。这些发现强调了跨界抗菌剂保守的结构-功能关系，表明防御素具有共同的进化历史，并突出了它们或其变体的治疗潜力。
