Advanced liquid handling for hydrogen/deuterium exchange mass spectrometry

点击翻译按钮获取中文摘要

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.