Advanced liquid handling for hydrogen/deuterium exchange mass spectrometry

1. 需要支持的研究项目（预期用途）。申请人（Boraston, Boulanger, Burke）对理解生物分子相互作用如何介导基本细胞过程的分子基础有共同的兴趣。通过CFI和NSERC RTI之前的投资，他们积累了最先进的生物物理基础设施，包括x射线晶体学（XRC），质谱（MS）和生化分析设施。这种协作结构和生物物理设施是生物化学和微生物系的主要优势之一，也是加拿大最具生产力的设施之一。拟议的研究项目包括微生物发病机制（Boraston, Boulanger, and Burke），新型生物燃料的产生（Boraston）和癌症信号（Burke, Boulanger）。几乎所有这些研究的基础都是以高通量的方式测量蛋白质动力学和分子相互作用，特别是蛋白质-蛋白质、蛋白质-小分子和蛋白质-碳水化合物相互作用。2. 所需设备和成本（1套LEAP HDX机器人系统，216,000美元，NSERC-RTI要求150,000美元）。我们需要先进的LEAP机器人处理系统，用于HDX-MS分析的自动样品处理。这将大大提高我们的能力，并将加拿大最具生产力的HDX-MS设施之一提升到一个新的水平。以下规范是关键的：#1 -吞吐量。拟议的申请人正在研究bb50蛋白靶点，主要目标是描述蛋白质动力学。Burke博士还使用HDX-MS与学术和工业合作伙伴广泛合作。拟议的系统将使吞吐量增加至少4倍，允许每天24小时运行。#2 -复杂的系统。我们分析生物分子相互作用的关键是描述复杂系统结合的能力。所要求的基础设施具有处理复杂蛋白质膜组件的能力。3. 与其他资金的关系。所有的主要申请者（Boraston和Burke）目前都得到了NSERC发现基金和CIHR运营基金的支持。这笔资金不允许购买基础设施。所要求的LEAP机器人系统将为该研究组合提供关键支持。该基础设施对于支持当前和未来与学术、工业和政府合作伙伴的项目也至关重要。4. 所需要的。目前迫切需要该仪器进行蛋白质动力学的HDX-MS分析。目前过时的基础设施正在失效（大约80年），如果没有替换，将关闭维多利亚大学高产的HDX-MS设施。这个要求的基础设施对于维多利亚大学生物物理研究的持续成功至关重要。

1. Research programs to be supported (Intended use). The applicants (Boraston, Boulanger, Burke) share a common interest in understanding the molecular basis of how biomolecular interactions mediate fundamental cellular processes. Through previous investments from CFI and NSERC RTI they have amassed state of the art biophysical infrastructure, including facilities for X-ray crystallography (XRC), mass spectrometry (MS), and biochemical assays. This collaborative structural and biophysical facility is one of the major strengths of the Biochemistry and Microbiology Department, and is one of the most productive in Canada. The proposed research programs include microbial pathogenesis (Boraston, Boulanger, and Burke), generation of novel biofuels (Boraston), and cancer signalling (Burke, Boulanger). Fundamental to almost all of these studies is the measurement of protein dynamics and molecular interactions, specifically protein-protein, protein-small molecule, and protein-carbohydrate interactions in a high throughput manner. 2. Equipment requested and cost (1 x LEAP HDX Robotic system at $216,000, with $150,000 requested from NSERC-RTI). We are requesting a state of the art LEAP robotic handling system for automated sample processing for HDX-MS analysis. This will dramatically increase our capabilities, and will take one of the most productive HDX-MS facilities in Canada to the next level. The following specifications are critical: #1 - Throughput. The proposed applicants are working on >50 protein targets, with a major goal being the characterisation of protein dynamics. Dr Burke also extensively collaborates with academic and industrial partners using HDX-MS. The proposed system will increase the throughput at least 4 fold, allowing for 24 hr/day operation. #2 - Complex systems. Key to our analysis of biomolecular interactions is the capability to characterise binding of complicated systems. The requested infrastructure has capabilities to handle complex protein-membrane assemblies. 3. Relationship to other funding. All of the primary applicants (Boraston and Burke) are currently supported by both NSERC Discovery grants and CIHR operating grants. This funding does not allow for infrastructure purchase. The LEAP robotic system requested will provide critical support to this research portfolio. This infrastructure will also be essential to support current and future projects with both academic, industrial and governmental partners. 4. Need. There is an urgent need for the proposed instrumentation to carry out HDX-MS analysis of protein dynamics. The current outdated infrastructure is failing (>8 years old) and without replacement will shut down the highly productive HDX-MS facility at the University of Victoria. This requested infrastructure will be essential for the continued success of biophysical research at the University of Victoria.