A Comprehensive Protein Expression, Purification and Crystallization (PEPC) System

蛋白质是所有细胞的主力，确定它们的功能和结构对理解生物体至关重要。蛋白质的表达和纯化使我们能够分离这些生物分子，并询问它们的功能并确定它们的三维结构。x射线晶体学是获得蛋白质和其他生物分子的原子分辨率结构的最可靠和有效的工具。总之，结构/功能研究有助于我们对生物过程的理解，以及如何操纵这些过程以造福制造业、食品生产、农业和医药。蛋白质的表达和纯化需要大容量的培养箱来培养细菌或哺乳动物细胞，以及复杂的色谱机来将感兴趣的蛋白质从其他细胞成分中分离出来。蛋白质结晶是一个费力、昂贵和耗时的过程，结晶机器人可以大大加快这一过程。这样的基础设施是现代生物化学实验室必不可少的。然而，申请人以及西蒙弗雷泽大学（SFU）的分子生物学和生物化学（MBB）部门缺乏有效表达、纯化和结晶蛋白质的基础设施，这影响了我们的HQP培训，扰乱了我们的研究进展，限制了我们的竞争力。NSERC-RTI提案的资金将允许申请人创建模块化，可扩展和尖端的蛋白质表达，纯化和结晶（PEPC）系统，该系统将在未来几年内支持SFU的生物化学研究。这个集成包将包括两个摇床孵化器，一个快速液相色谱仪，一个结晶机器人，生长室和显微镜。通过现代化和简化蛋白质和晶体的生成，PEPC系统将：(1)创造大量的劳动力和试剂成本节约，(2)显著加强HQP培训，(3)扩大我们nserc资助项目的研究潜力。申请人、MBB和SFU理学院承诺提供约13.7万美元的配套资金，突出了社区对这些资源的需求和支持。拟议的PEPC系统将满足nserc资助申请人的多样化和迫切需求。领导这个提议，克雷格实验室将利用这个基础设施来解决IV型细菌菌毛和其他蛋白质参与菌毛组装和分泌的结构。Larijani实验室将探索来自进化上遥远物种的DNA/ rna编辑酶的结构/功能关系和新的生化特性。最后，Audas实验室将检查与功能性和病理性淀粉样蛋白聚集相关的蛋白质构象变化。除了列出的申请人外，该资源将向所有SFU用户开放，并将大大扩展我们的研究能力，生产力和HQP培训。

Proteins are the workhorses of all cells, and determining their functions and structures is critical to understanding living organisms. Protein expression and purification allow us to isolate these biological molecules and interrogate their functions and determine their 3-dimensional structures. X-ray crystallography is the most reliable and effective tool to obtain atomic resolution structures of proteins and other biomolecules. Together, structure/function studies contribute to our understanding of biological processes and how these processes might be manipulated for the benefit of manufacturing, food production, agriculture and medicine. Protein expression and purification require large capacity incubators in which to grow bacterial or mammalian cells, and a sophisticated chromatography machine to separate the proteins of interest from other cellular components. Protein crystallization is a laborious, costly and time-consuming process that can be greatly expedited with a crystallization robot. Such infrastructure is essential to a modern biochemistry laboratory. Yet the applicants, and the Molecular Biology and Biochemistry (MBB) department at Simon Fraser University (SFU), lack infrastructure to efficiently express, purify and crystallize proteins, impacting our HQP training, disrupting our research progress and limiting our competitiveness. Funding of this NSERC-RTI proposal will allow the applicants to create a modular, expandable, and cutting-edge Protein Expression, Purification, and Crystallization (PEPC) system that will support biochemistry research at SFU for years to come. This integrated package will include two shaker incubators, a fast liquid phase chromatography instrument, and a crystallization robot, growth chamber and microscope. By modernizing and streamlining the generation of proteins and crystals, the PEPC system will: (1) create substantial labour and reagent cost-savings, (2) significantly enhance HQP training, and (3) expand the research potential of our NSERC-funded programs. The applicants, MBB, and the Faculty of Science at SFU have committed to providing approximately $137K in matching funds, highlighting the need and support for these resources within the community. The proposed PEPC system will serve the diverse and urgent needs of the NSERC-funded applicants. Leading this proposal, the Craig lab will use this infrastructure to solve the structures of Type IV bacterial pilins and other proteins involved in pilus assembly and secretion. The Larijani lab will explore structure/function relationships and novel biochemical properties of DNA/RNA-editing enzymes from evolutionarily distant species. Finally, the Audas lab will examine protein conformational changes associated with functional and pathological amyloid aggregation. Beyond the listed applicants, this resource will be open to all SFU users and will significantly expand our research capacity, productivity and HQP training.