Data_Sheet_1_Assessing the Mobility of Severe Acute Respiratory Syndrome Coronavirus-2 Spike Protein Glycans by Structural and Computational Methods.pdf

Two years after its emergence, the coronavirus disease-2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) remains difficult to control despite the availability of several vaccines. The extensively glycosylated SARS-CoV-2 spike (S) protein, which mediates host cell entry by binding to the angiotensin converting enzyme 2 (ACE2) through its receptor binding domain (RBD), is the major target of neutralizing antibodies. Like to many other viral fusion proteins, the SARS-CoV-2 spike protein utilizes a glycan shield to thwart the host immune response. To grasp the influence of chemical signatures on carbohydrate mobility and reconcile the cryo-EM density of specific glycans we combined our cryo-EM map of the S ectodomain to 4.1 Å resolution, reconstructed from a limited number of particles, and all-atom molecular dynamics simulations. Chemical modifications modeled on representative glycans (defucosylation, sialylation and addition of terminal LacNAc units) show no significant influence on either protein shielding or glycan flexibility. By estimating at selected sites the local correlation between the full density map and atomic model-based maps derived from molecular dynamics simulations, we provide insight into the geometries of the α-Man-(1→3)-[α-Man-(1→6)-]-β-Man-(1→4)-β-GlcNAc(1→4)-β-GlcNAc core common to all N-glycosylation sites.

新型冠状病毒肺炎（COVID-19）大流行由严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）引发，在其出现两年后，尽管已有多款疫苗获批使用，疫情仍难以得到有效管控。广泛糖基化的SARS-CoV-2刺突（S）蛋白通过其受体结合域（RBD）结合血管紧张素转换酶2（ACE2）介导宿主细胞入侵，是中和抗体的主要靶标。与多数其他病毒融合蛋白类似，SARS-CoV-2刺突蛋白借助聚糖屏障逃避宿主免疫系统的攻击。为阐明化学特征对聚糖流动性的影响，并解析特定聚糖的冷冻电镜（cryo-EM）密度，我们将由有限数量颗粒重构得到的、分辨率达4.1 Å的S蛋白胞外域冷冻电镜图谱，与全原子分子动力学模拟相结合。在代表性聚糖上建模的化学修饰——去岩藻糖基化、唾液酸化以及末端LacNAc单元的添加——对蛋白屏障作用或聚糖柔性均无显著影响。通过估算选定位点处完整密度图谱与基于分子动力学模拟得到的原子模型图谱之间的局部相关性，我们揭示了所有N-糖基化位点共有的α-Man-(1→3)-[α-Man-(1→6)-]-β-Man-(1→4)-β-GlcNAc-(1→4)-β-GlcNAc核心结构的几何特征。