Garrigues Sandra, Gandía Mónica, Castillo Laia, Coca María, Marx Florentine, Marcos Jose F, Manzanares Paloma
Department of Biotechnology, Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas, Valencia, Spain.
Centre for Research in Agricultural Genomics (CRAG, CSIC-IRTA-UAB-UB), Barcelona, Spain.
Front Microbiol. 2018 Oct 5;9:2370. doi: 10.3389/fmicb.2018.02370. eCollection 2018.
Antifungal proteins of fungal origin (AFPs) are small, secreted, cationic, and cysteine-rich proteins. Filamentous fungi encode a wide repertoire of AFPs belonging to different phylogenetic classes, which offer a great potential to develop new antifungals for the control of pathogenic fungi. The fungus is one of the few reported to encode three AFPs each belonging to a different phylogenetic class (A, B, and C). In this work, the production of the putative AFPs from was evaluated, but only the representative of class A, PeAfpA, was identified in culture supernatants of the native fungus. The biotechnological production of PeAfpB and PeAfpC was achieved in with the -based expression cassette, which had been proved to work efficiently for the production of other related AFPs in filamentous fungi. Western blot analyses confirmed that only produces PeAfpA naturally, whereas PeAfpB and PeAfpC could not be detected. From the three AFPs from , PeAfpA showed the highest antifungal activity against all fungi tested, including plant and human pathogens. . was also sensitive to its self-AFPs PeAfpA and PeAfpB. PeAfpB showed moderate antifungal activity against filamentous fungi, whereas no activity could be attributed to PeAfpC at the conditions tested. Importantly, none of the PeAFPs showed hemolytic activity. Finally, PeAfpA was demonstrated to efficiently protect against fungal infections caused by in tomato leaves and in oranges. The strong antifungal potency of PeAfpA, together with the lack of cytotoxicity, and significant protection against phytopathogenic fungi that cause postharvest decay and plant diseases, make PeAfpA a promising alternative compound for application in agriculture, but also in medicine or food preservation.
真菌源抗真菌蛋白（AFPs）是一类小分子、分泌型、阳离子且富含半胱氨酸的蛋白质。丝状真菌编码了多种属于不同系统发育类别的AFPs，这为开发用于控制致病真菌的新型抗真菌剂提供了巨大潜力。该真菌是少数被报道编码三种分别属于不同系统发育类（A、B和C）的AFPs的真菌之一。在这项工作中，对该真菌假定的AFPs的产生进行了评估，但在天然真菌的培养上清液中仅鉴定出A类代表蛋白PeAfpA。利用基于载体的表达盒在该真菌中实现了PeAfpB和PeAfpC的生物技术生产，该表达盒已被证明能有效地用于丝状真菌中其他相关AFPs的生产。蛋白质免疫印迹分析证实，该真菌仅天然产生PeAfpA，而未检测到PeAfpB和PeAfpC。在该真菌的三种AFPs中，PeAfpA对所有测试真菌，包括植物和人类病原体，均表现出最高的抗真菌活性。该真菌对其自身的AFPs PeAfpA和PeAfpB也敏感。PeAfpB对丝状真菌表现出中等抗真菌活性，而在测试条件下PeAfpC未表现出活性。重要的是，所有PeAFPs均未表现出溶血活性。最后，证明PeAfpA能有效保护番茄叶片免受该真菌以及橙子免受该真菌引起的真菌感染。PeAfpA强大的抗真菌效力，加上缺乏细胞毒性，以及对导致采后腐烂和植物病害的植物病原真菌具有显著的保护作用，使得PeAfpA成为农业、医学或食品保鲜领域应用的一种有前景的替代化合物。
