Interactions of SARS-CoV-2 N-protein

为了让我们的调查立即开始，我们已经检查了N-蛋白从几个商业来源，并确定了合适的蛋白质结构。与此同时，我们已经开始与尤金瓦尔科夫博士的实验室联合开发内部蛋白表达。我们目前正在优化溶液条件，并正在探索和实施纯化策略。 我们已经进行了初始沉降速度分析超离心实验，以评估寡聚状态，并发现它是二聚体，类似于SARS冠状病毒N-蛋白。我们探索了不同的荧光标记策略，以更好地测量自缔合的亲和力。我们还进行了小寡核苷酸的初步结合实验，这将有助于我们优化组装反应的生物物理测定。与Greg Piszczek博士合作，我们已经开始表征蛋白质及其复合物的热性质。我们正在评估第一批结果。

To allow an immediate start of our investigations we have examined N-protein from several commercial sources and identified suitable protein constructs. Simultaneously we have started to develop in-house protein expression, jointly with the laboratory of Dr. Eugene Valkov. We are currently optimizing solution conditions, and are exploring and implementing purification strategies. We have carried out initial sedimentation velocity analytical ultracentrifugation experiments to assess the oligomeric state and found it to be dimeric, similar to SARS-CoV N-protein. We explored different fluorescent labeling strategies to better measure the affinity of self-association. We have also carried out initial binding experiments with small oligonucleotides, which will help us to optimize the biophysical assays for assembly reactions. In collaboration with Dr. Greg Piszczek we have begun to characterize the thermal properties of the protein and its complexes. We are evaluating the first results.