Song Jing, Zhang Shanshan, Xing Junya, Zhang Licong, Wang Jiajun, Shan Anshan
College of Animal Science and Technology, Northeast Agricultural University, Harbin 150030, Heilongjiang, PR China.
College of Animal Science and Technology, Northeast Agricultural University, Harbin 150030, Heilongjiang, PR China.
J Adv Res. 2024 Sep 23. doi: 10.1016/j.jare.2024.09.017.
Antifungal peptides (AFPs) have the potential to treat antifungal-resistant infections; however, their structure-function relationship remains unknown, hindering their rapid development. Therefore, it is imperative to investigate and clarify the structure-function relationships of AFPs.
This study aimed to investigate the impact of end-tagging single hydrophobic amino acids and capping the N-terminus with glycine (Gly) on the antifungal activity of peptide W4.
The antifungal efficacy of the engineered peptides was initially assessed by determining the minimum inhibitory concentration (MIC) /minimal fungicidal concentration (MFC), killing kinetics, and drug resistance induction, in addition to evaluating the biocompatibility and stability. Subsequently, the antifungal mechanism was investigated using fluorescence labeling, electron microscopy, reactive oxygen species (ROS) detection, and measurement of mitochondrial membrane potential and apoptosis. The impact of the engineered peptides on Candida albicans (C. albicans) biofilm and their potential application in the scratch keratomycosis model were investigated.
The antifungal activity of W4 was significantly enhanced by capping Gly at the N-terminus, resulting in a decrease in average activity from 11.86 μM to 6.25 μM (GW4) and an increase in TI values by 1.9-fold (TI = 40.99). Mechanistically, GW4 exerted its antifungal effect by disrupting the cellular membrane structure in C. albicans, forming pores and subsequent leakage of intracellular contents. Concurrently, it facilitated intracellular ROS accumulation while decreasing the mitochondrial membrane potential. Additionally, GW4 demonstrated an excellent ability to inhibit and eliminate biofilms of C. albicans. Notably, GW4 demonstrated significant therapeutic potential in a C. albicans-associated keratitis model.
Capping Gly at the N-terminus increased residue length while significantly enhancing the helical propensity of W4, thereby augmenting its antifungal activity. Our exploratory study demonstrated the potential strategies and avenues for optimizing the structure-function relationships of AFPs and developing highly effective antifungal drugs.
抗真菌肽（AFP）具有治疗耐药性真菌感染的潜力；然而，它们的结构-功能关系仍不清楚，这阻碍了它们的快速发展。因此，研究和阐明AFP的结构-功能关系势在必行。
本研究旨在探讨在肽W4的末端标记单个疏水氨基酸以及用甘氨酸（Gly）封闭N端对其抗真菌活性的影响。
除了评估生物相容性和稳定性外，还通过测定最低抑菌浓度（MIC）/最低杀菌浓度（MFC）、杀菌动力学和耐药性诱导，初步评估工程化肽的抗真菌效果。随后，使用荧光标记、电子显微镜、活性氧（ROS）检测以及线粒体膜电位和细胞凋亡的测量来研究抗真菌机制。研究了工程化肽对白色念珠菌（C. albicans）生物膜的影响及其在划痕性角膜真菌病模型中的潜在应用。
在N端封闭Gly显著增强了W4的抗真菌活性，平均活性从11.86 μM降至6.25 μM（GW4），治疗指数（TI）值增加了1.9倍（TI = 40.99）。从机制上讲，GW4通过破坏白色念珠菌的细胞膜结构、形成孔并导致细胞内内容物泄漏来发挥其抗真菌作用。同时，它促进了细胞内ROS的积累，同时降低了线粒体膜电位。此外，GW4表现出优异的抑制和消除白色念珠菌生物膜的能力。值得注意的是，GW4在白色念珠菌相关角膜炎模型中显示出显著的治疗潜力。
在N端封闭Gly增加了残基长度，同时显著增强了W4的螺旋倾向，从而增强了其抗真菌活性。我们的探索性研究展示了优化AFP结构-功能关系和开发高效抗真菌药物的潜在策略和途径。
