Understanding enzyme-substrate recognition in catalysis and inhibition.

了解催化和抑制中的酶-底物识别。

• 批准号: RGPIN-2016-05765
• 负责人: Sanders, David
• 金额: $ 2.4万
• 依托单位: University of Saskatchewan
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=709991
• 关键词: Understanding; enzyme; substrate; recognition; catalysis

Research in my lab is focused on the broad theme of using protein X-ray crystallography to understand molecular recognition in enzyme systems. By combining crystallography with enzymology and other biophysical techniques, we can characterize the roles of individual amino acids in ligand recognition. We are using state of the art facilities, including the Canadian Light Source, the Saskatchewan Structural Sciences Centre, and the Protein Characterization and Crystallization Facility located at the University of Saskatchewan. My lab has established its reputation by using structural studies to enhance our understanding of how enzymes recognize and bind their ligands. By examining these interactions at the atomic level, we can predict the effects of alterations on binding. We have the ultimate goal of using the information to develop compounds that can more effectively interact with enzymes and to use structural information to predict substrates for new, poorly understood enzymes. We have been using the thioredoxin system as a model system for studying protein-protein interactions. Our current research objectives are to probe the interactions between thioredoxin (Trx) and thioredoxin reductase (TrxR), to characterize the interactions that are important for binding and to examine the way these enzymes are compensating for temperature differences and other evolutionary pressures to maintain their interaction. We have chosen this system because the chemistry is very well characterized, the Trx system is ubiquitous and oxidative stress (in which Trx plays a regulatory role) is of great current scientific interest. By using crystallography and enzymology we are characterizing the interactions of the thioredoxin system from extremophiles to determine the recognition characteristics of this system. A full understanding of the interactions that determine the recognition and binding of the TrxR/Trx complex will offer insights into other protein-protein interactions. Our work also involves understanding how small molecules are specifically recognized by enzymes. In past grants, we have studied protein-ligand interactions for a variety enzymes, including inositol dehydrogenase (IDH), dihydrodipicolinate synthase (DHDPS) and UDP-galactopyranose mutase (UGM). We have determined the structures of these enzymes complexed with substrates and inhibitors in order to determine the modes of interactions and substrate recognition. We have utilized this information for to give us insights into how these diverse enzymes catalyze their reactions. Additionally, the information gained from our structural studies have aided us in identifying new sub-classes of enzyme families and to design novel enzyme inhibitors. In this grant, we propose to continue our work in using protein x-ray crystallography and enzymology to examine the interactions between novel enzymes and their ligands.

我实验室的研究集中在一个广泛的主题上，即使用蛋白质X射线结晶学来理解酶系统中的分子识别。通过结合结晶学、酶学和其他生物物理技术，我们可以表征单个氨基酸在配体识别中的作用。我们正在使用最先进的设施，包括加拿大光源、萨斯喀彻温省结构科学中心，以及位于萨斯喀彻温省大学的蛋白质表征和结晶设施。 我的实验室通过使用结构研究来加强我们对酶如何识别和结合它们的配体的理解，从而建立了它的声誉。通过在原子水平上研究这些相互作用，我们可以预测变化对结合的影响。我们的最终目标是利用这些信息开发能够更有效地与酶相互作用的化合物，并利用结构信息预测新的、知之甚少的酶的底物。 我们一直使用硫氧还蛋白系统作为研究蛋白质-蛋白质相互作用的模型系统。我们目前的研究目标是探索硫氧还蛋白(Trx)和硫氧还蛋白还原酶(TrxR)之间的相互作用，表征对结合至关重要的相互作用，并研究这些酶如何补偿温差和其他维持它们相互作用的进化压力。我们之所以选择这个系统，是因为它的化学特性非常好，TRX系统无处不在，而且氧化应激(TRX在其中发挥调节作用)是当前非常有科学意义的。通过使用结晶学和酶学方法，我们表征了硫氧还蛋白系统与极端细菌的相互作用，以确定该系统的识别特性。充分了解决定TrxR/Trx复合体识别和结合的相互作用将为深入了解其他蛋白质-蛋白质相互作用提供依据。 我们的工作还包括了解小分子是如何被酶特异性识别的。在过去的资助中，我们研究了多种酶的蛋白质-配体相互作用，包括肌醇脱氢酶(IDH)、二氢二氢吡啶甲酸合成酶(DHDPS)和UDP-半乳糖变位酶(UGM)。我们已经确定了这些酶与底物和抑制剂的络合结构，以确定相互作用的模式和底物识别。我们利用这些信息来让我们深入了解这些不同的酶是如何催化它们的反应的。此外，从我们的结构研究中获得的信息有助于我们识别酶家族的新亚类和设计新的酶抑制剂。在这笔赠款中，我们建议继续使用蛋白质X射线结晶学和酶学来研究新型酶与其配体之间的相互作用。