Covid-19 pandemic outbreak is the reason of the current world health crisis. The development of effective antiviral compounds and vaccines requires detailed descriptive studies of the SARS-CoV-2 proteins. The SARS-CoV-2 spike (S) protein mediates virion binding to the human cells through its interaction with the ACE2 cell surface receptor and is one of the prime immunization targets. A functional virion is composed of three S1 and three S2 subunits created by furin cleavage of the spike protein at R682, a polybasic cleavage sites that differs from the SARS-CoV spike protein of 2002. We observe that the spike protein is O-glycosylated on a threonine (T678) near the furin cleavage site occupied by core-1 and core-2 structures. In addition, we have identified eight additional O-glycopeptides on the spike glycoprotein and we confirmed that the spike protein is heavily N-glycosylated. Our recently developed LC-MS/MS methodology allowed us to identify LacdiNAc structural motifs on all occupied N-glycopeptides and polyLacNAc structures on six glycopeptides of the spike protein. In conclusion, our study substantially expands the current knowledge of the spike proteins glycosylation and enables the investigation of the influence of the O-glycosylation on its proteolytic activation.

中文翻译：

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SARS-CoV-2 刺突蛋白的 N 和 O 糖基化。

Covid-19大流行的爆发是当前世界健康危机的原因。开发有效的抗病毒化合物和疫苗需要对 SARS-CoV-2 蛋白进行详细的描述性研究。SARS-CoV-2 刺突 (S) 蛋白通过与 ACE2 细胞表面受体的相互作用介导病毒粒子与人体细胞的结合，是主要的免疫靶点之一。一个功能性病毒体由三个 S1 和三个 S2 亚基组成，这些亚基由弗林蛋白酶切割 R682 处的刺突蛋白产生，这是一个不同于 2002 年 SARS-CoV 刺突蛋白的多元切割位点。我们观察到刺突蛋白在在核心 1 和核心 2 结构占据的弗林蛋白酶切割位点附近的苏氨酸 (T678)。此外，我们已经在刺突糖蛋白上鉴定出另外八种 O-糖肽，并且我们证实刺突蛋白是高度 N-糖基化的。我们最近开发的 LC-MS/MS 方法使我们能够识别刺突蛋白的六种糖肽上所有占据的 N-糖肽和 polyLacNAc 结构上的 LacdiNAc 结构基序。总之，我们的研究大大扩展了当前关于刺突蛋白糖基化的知识，并能够研究 O-糖基化对其蛋白水解活化的影响。