Spectrofluorimeter for protein and peptide research

蛋白质和多肽以及酶-多肽复合体的三维构象和折叠构象的确定是蛋白质结构功能评价和发现新的生物活性多肽的关键要求。为了进行这些结构研究，需要测量蛋白质和多肽的本征荧光，因为所提供的信息定义了微观和宏观环境，使得能够在各种实验条件下准确地解释蛋白质和多肽的三维结构。光谱荧光仪是唯一能够提供本征荧光数据的仪器，因此是结构-功能研究所需基础设施的关键组成部分。在马尼托巴大学食品和人类营养科学系，只有一台荧光仪，这是我在2005年通过NSERC RTI赠款购买的。然而，这台已有17年历史的仪器不再适用，因为该型号已停产，马尼托巴大学校园内也没有合适的替代系统。因此，这项提议的目的是用一个新的荧光计取代不能使用的17年的系统，这将为我的NSERC和加拿大研究主席资助的生物活性多肽研究项目的继续运行提供关键支持，并加强行业研究合作。在生物活性多肽存在和不存在的情况下，需要该仪器来测量酶的构象。通过对酶-多肽复合体性质的准确分析，可以确定多肽上的氨基酸残基，这些氨基酸残基对酶活性具有强大的抑制作用。这一点很重要，因为我的NSERC资助的发现计划专注于阐明新型食品蛋白质衍生生物活性多肽的结构-功能特性。由于酶-多肽相互作用对分子结构的影响是确定多肽功能的关键步骤，因此荧光仪是我的Discovery研究工作和成功培训高素质人员(HQP)所必需的组件。该荧光仪将用于阐明生物活性多肽的氨基酸序列与其与目标酶结合的亲和力之间的关系。荧光仪的产出将提高蛋白质作物(生物活性多肽生产的底物)的附加值利用率，并加强加拿大食品和营养业的经济价值。该荧光仪将由研究生和本科生、博士后研究员/研究助理和技术人员每天使用，以研究酶-肽复合体的结构性质，这可能会导致新的生物活性多肽的发现。因此，该工具对于成功培训HQP至关重要，为他们提供有助于加拿大食品和生物技术行业全球竞争力的适销对路的技能。

Determination of the 3-dimensional and folding conformation of proteins and peptides as well as enzyme-peptide complexes is a critical requirement for structure-function evaluation of proteins and the discovery of novel bioactive peptides. In order to perform these structural studies, measurement of the intrinsic fluorescence of proteins and peptides is a requirement because the information provided defines both the micro and macro environments, enabling accurate interpretation of the 3-dimensional structure of proteins and peptides under various experimental conditions. The spectrofluorimeter is the only instrument capable of providing intrinsic fluorescence data and is, therefore a critical component of the infrastructure needed for structure-function studies. At the Department of Food and Human Nutritional Sciences, University of Manitoba, there is only one fluorimeter, which I purchased in 2005 through an NSERC RTI grant. However, the 17-year-old instrument is no longer serviceable because the model has been discontinued and there is no suitable alternative system on the University of Manitoba campuses. Therefore, the aim of this proposal is to replace the non-serviceable 17-year-old system with a new fluorimeter that will provide critical support for continued operation of my NSERC and Canada Research Chair funded research programs on bioactive peptides and also strengthen industry research collaborations. The instrument is needed to measure the conformation of enzymes in the presence and absence of bioactive peptides. Accurate analysis of the properties of enzyme-peptide complexes enables identification of amino acid residues on peptides that provide potent inhibition of enzyme activity. This is important because my NSERC-funded Discovery program focuses on elucidation of the structure-function properties of novel food protein-derived bioactive peptides. Because the consequences to molecular structure of enzyme-peptide interactions represent a critical step in defining peptide function, the fluorimeter is a required component of my Discovery research work and for successful training of highly qualified personnel (HQP). The fluorimeter will be used to elucidate the relationships between amino acid sequence of bioactive peptides and their binding affinity to target enzymes. The output from the fluorimeter will increase value-added utilization of protein crops (substrates for bioactive peptides production) and strengthen the economic value of the Canadian food and nutrition industry. The fluorimeter will be used by graduate and undergraduate students, postdoctoral fellows/research associates, and technicians on a daily basis to study structural properties of enzyme-peptide complexes, which could lead to the discovery of novel bioactive peptides. Therefore, the instrument is critical to the successful training of HQPs to provide them with marketable skills that contribute to the global competitiveness of Canadian food and biotech industries.