Redesigning SARS CoV-2 spike protein to improve immunization - COVID-19

持续的COVID-19大流行继续带来巨大的健康和经济挑战，突出了对预防SARS-CoV-2进一步传播的疫苗的迫切需求。SARS-CoV-2的刺突蛋白S的受体结合域（RBD）是疫苗开发的靶点，因为它启动病毒进入宿主细胞。然而，RBD中的关键中和表位仅短暂暴露，这是由于“闭合”S构象的采样将这些表位隐藏于体液免疫系统之外。 我们的目标是（i）描绘导致SARS-CoV-2蛋白S的同源三聚体状态中关键表位暴露的结构机制，以及（ii）利用这些在S变体工程中的见解，通过保留中和敏感的构象和四级表位来提高免疫原性。 我们将通过多学科的方法来实现这一目标，该方法借鉴了研究团队和我们合作伙伴的专业知识。具体而言，我们将首先采用计算分子建模和模拟来设计变体，这些变体在RBD及其周围暴露抗原区域的情况下稳定“开放”状态（Gsponer Lab）。 然后，我们将在单粒子Cryo-EM（Subramaniam实验室）的帮助下表达，纯化和表征计算设计变体的所需结构效应。最后，与我们的生物技术合作伙伴AbCellera合作，我们将使用经过充分表征和结构验证的变体来免疫人源化小鼠，并测试诱导的抗血清和抗体的中和活性。 对拟议研究的支持将使我们能够建立一个管道，用于新S蛋白变体的工程和结构表征，以及这些设计变体的免疫原性的后续表征。

The ongoing COVID-19 pandemic continues to pose enormous health and economic challenges, highlighting the urgent need for a vaccine that prevents further spread of SARS-CoV-2. The receptor binding domain (RBD) of SARS-CoV-2' s spike protein S is a target for vaccine development because it initiates entry of the virus into host cells. However, key neutralizing epitopes in the RBD are only transiently exposed due to the samplings of "closed" S conformations that hide these epitopes from the humoral immune system. Our goals are to (i) delineate structural mechanisms leading to exposures of key epitopes in the homo-trimeric state of the SARS-CoV-2 protein S and (ii) exploit these insights in the engineering of S variants that improve immunogenicity through preservation of neutralization-sensitive conformational and quaternary epitopes. We will achieve this in a multidisciplinary approaches that draws from the expertise present in the research team and our partner. Specifically, we will first employ computational molecular modeling and simulations to design variants that stabilize "open'" states with exposed antigenic regions in RBD and its surroundings (Gsponer Lab). We will then express, purify, characterize the desired structural effects of the computationally designed variants with the help of single particle Cryo-EM (Subramaniam lab). Finally, in collaboration with our biotech partner AbCellera, we will use well-characterized and structurally-validated variants for immunization of humanized mice and test neutralizing activities of induced anti-sera and antibodies. Support for the proposed research will enable us to establish a pipeline for the engineering and structural characterization of new S protein variants and the subsequent characterization of the immunogenicity of these designed variants.