Structure Function of Food Related Proteins

我们NSERC资助的研究项目的长期目标是了解解释蛋白质功能如何由蛋白质结构决定的基本过程。在食品系统中，蛋白质(包括酶)在食品安全/植物病理学中具有显著的质构、风味、气味、营养和预防作用(例如，抗微生物和植物防御蛋白)以及加工助剂(例如，凝乳酶在干酪生产中凝结牛奶)。这项提议的研究重点是一组被称为天冬氨酸蛋白酶的酶，它们具有非常相似的结构和作用模式，最好的代表是胃酶胃蛋白酶。尽管有这种一致性，但存在着广泛的不同功能，这使它们成为研究蛋白质结构如何决定功能的理想模型。在我们团队和其他人之前研究的基础上，拟议的研究计划解决了两个主要领域的基本知识空白：1)植物天冬氨酸蛋白酶的食物腐败相关生物活性蛋白片段；2)天冬氨酸蛋白酶结构的基础，对高温和碱性的耐受性，以及对特定目标的选择性。对于第一个研究领域，来自植物的天冬氨酸蛋白酶与来自非植物来源的天冬氨酸蛋白酶相比，具有额外的部分，称为植物特异性插入(PSI)。PSI是植物天冬氨酸蛋白酶与膜相互作用的关键，包括对植物病原体的活性，作为植物对入侵真菌的免疫反应的一部分。为了更好地了解PSI的作用模式，将在最高水平的分子细节上解决与膜的结构和特定接触点的问题。第二个研究领域将需要解决很少被研究的疟疾天冬氨酸蛋白酶(纤溶酶V)与已知抑制剂复合的结构，以缩小可能的结合位点范围，从而指导针对纤溶酶V的计算机辅助抑制剂设计。其他研究将有助于理解非典型天冬氨酸蛋白酶如何能够在极端温度(来自酸性温泉中的生物体的热蛋白酶)和非酸性pH(来自人类肾脏的肾素)下工作。Thermopsin和Renin将作为工程胃酶(不是温度或中性pH稳定的)的模板，提供一种测试关键结构特征的稳定效果的方法。拟议研究计划的影响将是提高关于AP结构和功能的知识，从而为干预与食品有关的挑战(如植物疾病和酶生物技术应用)提供新的战略。

The long-term goal of our NSERC-funded research program has been to understand the fundamental processes that explain how protein functionality is dictated by protein structure. In food systems, proteins (including enzymes) have prominent roles in texture, flavour, odour, nutrition and prophylaxis in food safety/plant pathology (e.g., antimicrobial and plant defense proteins) as well as being processing aids (e.g., chymosin coagulates milk in cheese production). The research focus of this proposal is on a group of enzymes called aspartic proteases which share highly similar structures and modes of action, that are best represented by the stomach enzyme pepsin. Despite this uniformity, a wide spectrum of different functions exist making them an ideal model to study how protein structure determines function. Building on previous research by our group and others, the proposed research program addresses fundamental knowledge gaps in two overarching areas: 1) food spoilage-related bioactive protein segments of aspartic proteases in plants; and 2) the underpinnings of aspartic protease structure tolerance to heat and alkalinity, and selectivity for particular targets. For the first research area, aspartic proteases from plants have an extra portion compared to those from non-plant sources called the plant-specific insert (PSI). The PSI is critical for plant aspartic proteases to interact with membranes, including activity against plant pathogens as part of plants’ immune responses to invading fungi. The structure and specific points of contact with membranes will be solved at the highest level of molecular detail in order to better understand the mode of action of the PSI. The second research area will entail solving the structures of a little-studied malarial aspartic protease (plasmepsin V) complexed with well-known inhibitors to narrow the possible range of binding sites, so as to direct computer-aided inhibitor design specific to plasmepsin V. Other studies will contribute to understanding how atypical aspartic proteases are able to work at extreme temperature (thermopsin from an organism found in acidic hot springs) and non-acidic pH (renin from human kidney). Thermopsin and renin will serve as templates for engineering pepsin (which is not temperature- or neutral pH-stable) by providing a means of testing the stabilizing effects of key structural features. The impact of the proposed research program will be to improve knowledge regarding AP structures and functions thereby enabling novel strategies for interventions in food-related challenges such as plant diseases and enzyme biotechnological applications.