Advanced liquid handling for hydrogen/deuterium exchange mass spectrometry

1.拟支持的研究项目(目标用途)。申请者(Boraston，Boulanger，Burke)在理解生物分子相互作用如何调节基本细胞过程的分子基础方面有着共同的兴趣。通过CFI和NSERC RTI之前的投资，他们已经积累了最先进的生物物理基础设施，包括X射线结晶学(XRC)、质谱仪(MS)和生化分析设施。这个协作的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，也是加拿大生产率最高的设施之一。拟议的研究计划包括微生物发病机制(Boraston、Boulanger和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-碳水化合物的相互作用。2.所需设备和费用(1个LEAP HDX机器人系统，216000美元，向NSERC-RTI申请150000美元)。我们要求最先进的LEAP机器人处理系统，用于HDX-MS分析的自动样品处理。这将极大地提高我们的能力，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。以下规格至关重要：#1-吞吐量。被提议的申请者正在研究&gt；50个蛋白质靶标，主要目标是描述蛋白质动力学。伯克博士还使用HDX-MS与学术界和工业界合作伙伴广泛合作。拟议的系统将使吞吐量增加至少4倍，允许24小时/天的运营。#2-复杂系统。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。所要求的基础设施有能力处理复杂的蛋白质-膜组件。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探索补助金和CIHR运营补助金的支持。这笔资金不允许购买基础设施。所要求的LEAP机器人系统将为这一研究组合提供关键支持。这一基础设施对于支持当前和未来与学术界、工业界和政府合作的项目也是至关重要的。4.需要。迫切需要拟议的仪器来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灵(已有8年历史)，如果不更换，维多利亚大学高生产率的HDX-MS设施将被关闭。所要求的基础设施将是维多利亚大学生物物理研究继续取得成功的关键。

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.