Izadi Shiva, Vavra Ulrike, Melnik Stanislav, Grünwald-Gruber Clemens, Föderl-Höbenreich Esther, Sack Markus, Zatloukal Kurt, Glössl Josef, Stöger Eva, Mach Lukas, Castilho Alexandra, Strasser Richard
Institute of Plant Biotechnology and Cell Biology, Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
Core Facility Mass Spectrometry, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
Front Bioeng Biotechnol. 2023 May 3;11:1180044. doi: 10.3389/fbioe.2023.1180044. eCollection 2023.
SARS-CoV-2 infects human cells via binding of the viral spike glycoprotein to its main cellular receptor, angiotensin-converting enzyme 2 (ACE2). The spike protein-ACE2 receptor interaction is therefore a major target for the development of therapeutic or prophylactic drugs to combat coronavirus infections. Various engineered soluble ACE2 variants (decoys) have been designed and shown to exhibit virus neutralization capacity in cell-based assays and models. Human ACE2 is heavily glycosylated and some of its glycans impair binding to the SARS-CoV-2 spike protein. Therefore, glycan-engineered recombinant soluble ACE2 variants might display enhanced virus-neutralization potencies. Here, we transiently co-expressed the extracellular domain of ACE2 fused to human Fc (ACE2-Fc) with a bacterial endoglycosidase in to produce ACE2-Fc decorated with N-glycans consisting of single GlcNAc residues. The endoglycosidase was targeted to the Golgi apparatus with the intention to avoid any interference of glycan removal with concomitant ACE2-Fc protein folding and quality control in the endoplasmic reticulum. The deglycosylated ACE2-Fc carrying single GlcNAc residues displayed increased affinity to the receptor-binding domain (RBD) of SARS-CoV-2 as well as improved virus neutralization activity and thus is a promising drug candidate to block coronavirus infection.
严重急性呼吸综合征冠状病毒2（SARS-CoV-2）通过病毒刺突糖蛋白与其主要细胞受体血管紧张素转换酶2（ACE2）结合来感染人类细胞。因此，刺突蛋白与ACE2受体的相互作用是开发治疗或预防药物以对抗冠状病毒感染的主要靶点。已设计出各种工程化的可溶性ACE2变体（诱饵），并在基于细胞的试验和模型中显示出具有病毒中和能力。人ACE2高度糖基化，其一些聚糖会损害与SARS-CoV-2刺突蛋白的结合。因此，糖基工程化的重组可溶性ACE2变体可能具有增强的病毒中和效力。在此，我们将与人类Fc融合的ACE2胞外域（ACE2-Fc）与一种细菌内切糖苷酶在[具体细胞或系统]中瞬时共表达，以产生用由单个N-乙酰葡糖胺残基组成的N-聚糖修饰的ACE2-Fc。该内切糖苷酶靶向高尔基体，目的是避免聚糖去除对内质网中ACE2-Fc蛋白折叠和质量控制的任何干扰。携带单个N-乙酰葡糖胺残基的去糖基化ACE2-Fc对SARS-CoV-2的受体结合域（RBD）表现出增加的亲和力以及改善的病毒中和活性，因此是一种有前景的阻断冠状病毒感染的候选药物。
