Animal Nutrition Institute, Chongqing Academy of Animal Science, Chongqing 402460, China.
Institute of Nutrition and Feed, National Center of Technology Innovation for Pigs, Chongqing 402460, China.
Int J Mol Sci. 2024 Sep 29;25(19):10508. doi: 10.3390/ijms251910508.
The interaction forces and mechanical properties of the interaction between melittin (Mel) and lipopolysaccharide (LPS) are considered to be crucial driving forces for Mel when killing Gram-negative bacteria (GNB). However, how their interaction forces perform at the single-molecule level and the dissociation kinetic characteristics of the Mel/LPS complex remain poorly understood. In this study, the single-molecule-level interaction forces between Mel and LPSs from K-12, O55:B5, O111:B4, and O128:B12 were explored using atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS). AFM-based dynamic force spectroscopy (DFS) and an advanced analytical model were employed to investigate the kinetic characteristics of the Mel/LPS complex dissociation. The results indicated that Mel could interact with both rough (R)-form LPS ( K-12) and smooth (S)-form LPSs ( O55:B5, O111:B4, and O128:B12). The S-form LPS showed a more robust interaction with Mel than the R-form LPS, and a slight difference existed in the interaction forces between Mel and the diverse S-form LPS. Mel interactions with the S-form LPSs showed greater specific and non-specific interaction forces than the R-form LPS ( < 0.05), as determined by AFM-based SMFS. However, there was no significant difference in the specific and non-specific interaction forces among the three samples of S-form LPSs ( > 0.05), indicating that the variability in the O-antigen did not affect the interaction between Mel and LPSs. The DFS result showed that the Mel/S-form LPS complexes had a lower dissociation rate constant, a shorter energy barrier width, a longer bond lifetime, and a higher energy barrier height, demonstrating that Mel interacted with S-form LPS to form more stable complexes. This research enhances the existing knowledge of the interaction micromechanics and kinetic characteristics of Mel and LPS at the single-molecule level. Our research may help with the design and evaluation of new anti-GNB drugs.
蜂毒素（Mel）与脂多糖（LPS）相互作用的相互作用力和力学特性被认为是 Mel 杀死革兰氏阴性菌（GNB）的关键驱动力。然而，它们在单分子水平上的相互作用力以及 Mel/LPS 复合物的解离动力学特性仍知之甚少。在这项研究中，使用原子力显微镜（AFM）-基于单分子力谱（SMFS）研究了 Mel 与 K-12、O55:B5、O111:B4 和 O128:B12 的 LPS 之间的单分子级相互作用力。AFM 基于动态力谱（DFS）和先进的分析模型被用来研究 Mel/LPS 复合物解离的动力学特性。结果表明，Mel 可以与粗糙（R）形式 LPS（K-12）和光滑（S）形式 LPS（O55:B5、O111:B4 和 O128:B12）相互作用。S 形式 LPS 与 Mel 的相互作用比 R 形式 LPS 更牢固，而 Mel 与不同 S 形式 LPS 之间的相互作用力存在微小差异。通过 AFM 基于 SMFS 测定，Mel 与 S 形式 LPS 的相互作用表现出更大的特异性和非特异性相互作用力，比 R 形式 LPS 大（<0.05）。然而，三种 S 形式 LPS 的特异性和非特异性相互作用力没有显著差异（>0.05），这表明 O-抗原的变异性并不影响 Mel 与 LPS 之间的相互作用。DFS 结果表明，Mel/S 形式 LPS 复合物的解离速率常数较低，能量势垒宽度较短，键寿命较长，能量势垒高度较高，这表明 Mel 与 S 形式 LPS 相互作用形成更稳定的复合物。这项研究增强了现有关于 Mel 和 LPS 在单分子水平上的相互作用力学和动力学特性的知识。我们的研究可能有助于新的抗 GNB 药物的设计和评估。
