Poonsiri Thanalai, Stransky Jan, Demitri Nicola, Haas Hubertus, Cianci Michele, Benini Stefano
Bioorganic Chemistry and Bio-Crystallography Laboratory (B2Cl) Faculty of Agricultural, Environmental and Food Sciences, Libera Università di Bolzano, Piazza Università, 1, 39100 Bolzano, Italy.
Institute of Biotechnology, AS CR, Centre of Molecular Structure, Průmyslová 595, 252 50 Vestec, Czech Republic.
J Struct Biol X. 2024 Dec 26;11:100119. doi: 10.1016/j.yjsbx.2024.100119. eCollection 2025 Jun.
Siderophore-mediated iron acquisition is essential for the virulence of , a fungus causing life-threatening aspergillosis. Drugs targeting the siderophore biosynthetic pathway could help improve disease management. The transacetylases SidF and SidL generate intermediates for different siderophores in . has a yet unidentified transacetylase that complements SidL during iron deficiency in SidL-lacking mutants. We present the first X-ray structure of SidF, revealing a two-domain architecture with tetrameric assembly. The N-terminal domain contributes to protein solubility and oligomerization, while the C-terminal domain containing the GCN5-related N-acetyltransferase (GNAT) motif is crucial for the enzymatic activity and mediates oligomer formation. Notably, AlphaFold modelling demonstrates structural similarity between SidF and SidL. Enzymatic assays showed that SidF can utilize acetyl-CoA as a donor, previously thought to be a substrate of SidL but not SidF, and selectively uses N5-hydroxy-L-ornithine as an acceptor. This study elucidates the structure of SidF and reveals its role in siderophore biosynthesis. We propose SidF as the unknown transacetylase complementing SidL activity, highlighting its central role in siderophore biosynthesis. Investigation of this uncharacterized GNAT protein enhances our understanding of fungal virulence and holds promise for its potential application in developing antifungal therapies.
铁载体介导的铁摄取对于烟曲霉的毒力至关重要，烟曲霉是一种可引发危及生命的曲霉病的真菌。靶向铁载体生物合成途径的药物有助于改善疾病管理。转乙酰酶SidF和SidL为烟曲霉中不同的铁载体生成中间体。烟曲霉有一种尚未鉴定的转乙酰酶，在缺乏SidL的突变体缺铁时可补充SidL的功能。我们展示了SidF的首个X射线结构，揭示了其具有四聚体组装的双结构域架构。N端结构域有助于蛋白质的溶解性和寡聚化，而包含GCN5相关N - 乙酰转移酶（GNAT）基序的C端结构域对于酶活性至关重要，并介导寡聚体形成。值得注意的是，AlphaFold建模显示SidF和SidL之间存在结构相似性。酶活性测定表明，SidF可以利用乙酰辅酶A作为供体，乙酰辅酶A以前被认为是SidL而非SidF的底物，并且SidF选择性地使用N5 - 羟基 - L - 鸟氨酸作为受体。这项研究阐明了SidF的结构，并揭示了其在铁载体生物合成中的作用。我们提出SidF就是补充SidL活性的未知转乙酰酶，突出了其在烟曲霉铁载体生物合成中的核心作用。对这种未表征的GNAT蛋白的研究增进了我们对真菌毒力的理解，并有望将其应用于开发抗真菌疗法。
