Stabilizing the prefusion SARS-CoV-2 spike The development of therapeutic antibodies and vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is focused on the spike (S) protein that decorates the viral surface. A version of the spike ectodomain that includes two proline substitutions (S-2P) and stabilizes the prefusion conformation has been used to determine high-resolution structures. However, even S-2P is unstable and difficult to produce in mammalian cells. Hsieh et al. characterized many individual and combined structure-guided substitutions and identified a variant, named HexaPro, that retains the prefusion conformation but shows higher expression than S-2P and can also withstand heating and freezing. This version of the protein is likely to be useful in the development of vaccines and diagnostics. Science, this issue p. 1501 The design of stabilizing mutations in the SARS-CoV-2 spike protein allows for high-yield production of a critical vaccine antigen. The coronavirus disease 2019 (COVID-19) pandemic has led to accelerated efforts to develop therapeutics and vaccines. A key target of these efforts is the spike (S) protein, which is metastable and difficult to produce recombinantly. We characterized 100 structure-guided spike designs and identified 26 individual substitutions that increased protein yields and stability. Testing combinations of beneficial substitutions resulted in the identification of HexaPro, a variant with six beneficial proline substitutions exhibiting higher expression than its parental construct (by a factor of 10) as well as the ability to withstand heat stress, storage at room temperature, and three freeze-thaw cycles. A cryo–electron microscopy structure of HexaPro at a resolution of 3.2 angstroms confirmed that it retains the prefusion spike conformation. High-yield production of a stabilized prefusion spike protein will accelerate the development of vaccines and serological diagnostics for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

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基于结构的预融合稳定 SARS-CoV-2 刺突设计

稳定融合前的 SARS-CoV-2 刺突 针对严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2) 的治疗性抗体和疫苗的开发主要集中在修饰病毒表面的刺突 (S) 蛋白上。包含两个脯氨酸取代 (S-2P) 并稳定融合前构象的尖峰胞外域版本已被用于确定高分辨率结构。然而，即使是 S-2P 也不稳定，难以在哺乳动物细胞中产生。谢等人。表征了许多单独和组合的结构引导替换，并鉴定了一种名为 HexaPro 的变体，它保留了融合前的构象，但表现出比 S-2P 更高的表达，并且还可以承受加热和冷冻。这个版本的蛋白质很可能在疫苗和诊断学的开发中有用。科学，这个问题p。1501 SARS-CoV-2 刺突蛋白中稳定突变的设计允许高产量生产关键疫苗抗原。2019 年冠状病毒病 (COVID-19) 大流行已导致加快开发治疗方法和疫苗的努力。这些努力的一个关键目标是刺突 (S) 蛋白，它是亚稳态且难以重组生产的。我们对 100 种结构导向的尖峰设计进行了表征，并确定了 26 种增加蛋白质产量和稳定性的单独替代。有益替代组合的测试导致鉴定出 HexaPro，这是一种具有六个有益脯氨酸替代的变体，表现出比其亲本构建体更高的表达（10 倍）以及承受热应激、室温储存和三个冻融循环。分辨率为 3.2 埃的 HexaPro 低温电子显微镜结构证实它保留了融合前的尖峰构象。稳定的融合前刺突蛋白的高产量生产将加速严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）疫苗和血清学诊断的开发。