EAGER: Collaborative Research: Design of Inhibitors for ORF7a and ORF7b Oligomerization in COVID-19

EAGER：合作研究：COVID-19 中 ORF7a 和 ORF7b 寡聚化抑制剂的设计

• 批准号: 2029900
• 负责人: Jeffery Klauda
• 金额: $ 15万
• 依托单位: University of Maryland, College Park
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-15 至 2022-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2029900&HistoricalAwards=false
• 关键词: EAGER; Collaborative; Research; Design; Inhibitors

With this award, the Chemistry of Life Processes Program in the Chemistry Division and the Chemical and Biochemical Engineering Program in the Chemical, Bioengineering, Environmental and Transport Systems Division are funding Dr. Bryan Berger (University of Virginia) and Dr. Jeffery Klauda (University of Maryland) to investigate two proteins named ORF7a and OR7b from the COVID19 virus that have been implicated in how harmful the virus is to its host, e.g. the human cells. The research will focus on how these two proteins form larger protein complexes that in turn affect the interactions between the virus and the infected cells and influence the immune response of the host. The research informs the development of peptides that could be used to probe the viral propagation. The research is based on the use of a combination of computational and experimental methods. Dr. Berger and Dr. Klauda distribute to the scientific community free of charge through Addgene the plasmids and associated protocols developed for this project, thus enabling the global scientific community that works on finding a solution to the current pandemic and to minimizing the possibility of future outbreaks to quickly use the outcomes of their research. This work will provide training for post-doctoral fellows working on critical challenges using state-of-the-art experimental and computational methods. The results of the research will be disseminated by the team to the greater community through conferences and workshops at University of Virginia and University of Maryland and through publications. The researchers also plan to inform and educate students on possible mechanisms of virus transmission and prevention by participation in existing outreach programs at their Institutions.This research project seeks to understand the basis of specificity for transmembrane and juxtamembrane oligomerization of ORF7a with BST-2 and for homooligomerization of ORF7b. Using bacterial transcriptional assays for membrane protein dimerization based on the E. coli AraC protein (AraTM and DN-AraTM assays), the researches determine specific amino acid residues and structural motifs responsible for the protein oligomerization in bacterial membranes. This knowledge informs computational models for formation of BST-2/ORF7a heterooligomers and ORF7b homooligomers. In turn, the computational models are used to make critical new predictions of sequences for transmembrane peptides that could influence protein-protein interactions involving ORF7a and ORF7b. These predictions and the properties of the peptides are tested by synthesizing peptide libraries and using AraTM, DN-AraTM, and mammalian, cell-based fluorescence resonance energy transfer assays. Validation of candidate sequences are achieved using mammalian cell-based assays for BST-2 function and apoptosis. The results of these studies could provide high-resolution, experimentally validated models for OR7a and ORF7b homo and heterooligomerization, as well as peptide sequences that can be used to probe the roles of ORF7a and ORF7b in viral propagation in vivo.This grant is being awarded using funds made available by the Coronavirus Aid, Relief, and Economic Security (CARES) Act supplement allocated to MPS and ENG.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

获得这一奖项的是化学、生物工程、环境与运输系统部的生命过程化学项目和化学、生物工程、环境与运输系统部门的化学与生化工程项目，他们资助弗吉尼亚大学的Bryan Berger博士和马里兰大学的Jeffery Klauda博士研究COVID19病毒的两种名为ORF7a和OR7b的蛋白质，它们与病毒对宿主，例如人类细胞的危害有关。研究的重点将是这两种蛋白质如何形成更大的蛋白质复合体，进而影响病毒与感染细胞之间的相互作用，并影响宿主的免疫反应。这项研究为可用于探测病毒传播的多肽的开发提供了信息。这项研究采用了计算和实验相结合的方法。Berger博士和Klauda博士通过Addgene免费向科学界分发为该项目开发的质粒和相关方案，从而使致力于找到解决当前流行病的解决方案并将未来爆发的可能性降至最低的全球科学界能够迅速使用他们的研究成果。这项工作将为使用最先进的实验和计算方法处理关键挑战的博士后研究员提供培训。研究小组将通过弗吉尼亚大学和马里兰大学的会议和研讨会以及出版物向更大的社区传播研究结果。研究人员还计划通过参加所在学院现有的推广计划来告知和教育学生有关病毒传播和预防的可能机制。本研究项目旨在了解ORF7a与BST-2跨膜和膜旁齐聚以及ORF7b同源齐聚的特异性基础。利用基于大肠杆菌AraC蛋白的细菌转录实验(AraTM和DN-AraTM实验)，确定了导致细菌膜蛋白齐聚的特定氨基酸残基和结构基序。这一知识为BST-2/ORF7a杂低聚物和ORF7b纯低聚物的形成提供了计算模型。反过来，计算模型被用来对可能影响涉及ORF7a和ORF7b的蛋白质-蛋白质相互作用的跨膜多肽序列进行关键的新预测。这些预测和多肽的性质通过合成肽文库和使用AraTM、dN-AraTM和哺乳动物的基于细胞的荧光共振能量转移分析来验证。候选序列的验证是通过基于哺乳动物细胞的BST-2功能和细胞凋亡检测来实现的。这些研究的结果可以为OR7a和ORF7b的同源和异寡聚提供高分辨率的实验验证模型，以及可以用来探索ORF7a和ORF7b在活体病毒传播中的作用的肽序列。这项拨款是使用分配给MPS和ENG的冠状病毒援助、救济和经济安全(CARE)法案附录提供的资金授予的。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

All-Atom Modeling of Complex Cellular Membranes

• DOI: 10.1021/acs.langmuir.1c02084
• 发表时间: 2021-12-28
• 期刊: LANGMUIR
• 影响因子: 3.9
• 作者: Hsieh, Min-Kang;Yu, Yalun;Klauda, Jeffery B.
• 通讯作者: Klauda, Jeffery B.

Variational embedding of protein folding simulations using Gaussian mixture variational autoencoders

• DOI: 10.1063/5.0069708
• 发表时间: 2021-11-21
• 期刊: JOURNAL OF CHEMICAL PHYSICS
• 影响因子: 4.4
• 作者: Ghorbani, Mahdi;Prasad, Samarjeet;Brooks, Bernard R.
• 通讯作者: Brooks, Bernard R.