Fraser Bryan J, Wilson Ryan P, Ferková Sára, Ilyassov Olzhas, Lac Jackie, Dong Aiping, Li Yen-Yen, Seitova Alma, Li Yanjun, Hejazi Zahra, Kenney Tristan M G, Penn Linda Z, Edwards Aled, Leduc Richard, Boudreault Pierre-Luc, Morin Gregg B, Bénard François, Arrowsmith Cheryl H
Structural Genomics Consortium Toronto, Toronto, ON, Canada.
Department of Medical Biophysics, University of Toronto, Toronto, ON, Canada.
Nat Commun. 2025 May 10;16(1):4351. doi: 10.1038/s41467-025-59677-3.
Transmembrane Protease, Serine-2 (TMPRSS2) and TMPRSS11D are human proteases that enable SARS-CoV-2 and Influenza A/B virus infections, but their biochemical mechanisms for facilitating viral cell entry remain unclear. We show these proteases spontaneously and efficiently cleave their own zymogen activation motifs, activating their broader protease activity on cellular substrates. We determine TMPRSS11D co-crystal structures with a native and an engineered activation motif, revealing insights into its autocleavage activation and distinct substrate binding cleft features. Leveraging this structural data, we develop nanomolar potency peptidomimetic inhibitors of TMPRSS11D and TMPRSS2. We show that a broad serine protease inhibitor that underwent clinical trials for TMPRSS2-targeted COVID-19 therapy, nafamostat mesylate, was rapidly cleaved by TMPRSS11D and converted to low activity derivatives. In this work, we develop mechanistic insights into human protease viral tropism and highlight both the strengths and limitations of existing human serine protease inhibitors, informing future drug discovery efforts targeting these proteases.
跨膜丝氨酸蛋白酶2（TMPRSS2）和TMPRSS11D是人类蛋白酶，可促成严重急性呼吸综合征冠状病毒2（SARS-CoV-2）和甲型/乙型流感病毒感染，但其促进病毒进入细胞的生化机制仍不清楚。我们发现这些蛋白酶能自发且高效地切割自身的酶原激活基序，从而激活其对细胞底物的更广泛蛋白酶活性。我们确定了TMPRSS11D与天然和工程化激活基序的共晶体结构，揭示了其自切割激活以及独特的底物结合裂隙特征。利用这些结构数据，我们开发出了纳摩尔效力的TMPRSS11D和TMPRSS2拟肽抑制剂。我们发现，一种曾用于针对TMPRSS2的新冠肺炎治疗临床试验的广谱丝氨酸蛋白酶抑制剂甲磺酸萘莫司他，被TMPRSS11D迅速切割并转化为低活性衍生物。在这项研究中，我们深入了解了人类蛋白酶的病毒嗜性机制，并突出了现有人类丝氨酸蛋白酶抑制剂的优势和局限性，为未来针对这些蛋白酶的药物研发工作提供了参考。
