Structure-Function Relationships of Food-Related Enzymes: Aspartic Proteases

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The long-term goal of Dr. Rickey Yada's NSERC-funded research program has been to fundamentally understand how an enzyme's functionality is determined by its structure. In food systems, enzymes have wide-ranging functions including prevention of microbial spoilage, enhancing nutritional value of protein sources, industrial sanitisation, and acting as food processing aids in the manufacture of a variety of products (e.g., cheese making). This Discovery Grant proposal is focused on aspartic proteases, a group of enzymes which share highly similar structures and are best represented by the stomach enzyme pepsin. Despite this uniformity, minor structural differences result in a wide spectrum of diverse functions, making them an ideal model to study how enzyme structure determines function. Specifically, knowledge gaps in two overarching areas will be addressed: First, understanding the structural underpinnings of aspartic protease tolerance to environmental conditions (heat, acidity, and pressure) will involve the investigation of atypical aspartic proteases that are capable of functioning at high temperature (thermopsin from an organism in hot springs) and non-acidic conditions (renin from human kidney). Through protein engineering, the stabilising effects of suspected key structural features present in thermopsin and renin will be tested in pepsin (which is neither stable at high temperature nor non-acidic conditions). Additionally, the structure of an aspartic protease called plasmepsin V will be resolved so that lessons relevant to stability can be learned from its unique way of forming its active structure, potentially leading to the design of inhibitors. Lastly, the effects of High Pressure Processing (an emerging processing technology in the food industry) on the structures of several cheese making aspartic proteases will be characterised. A second research area will examine a portion of aspartic proteases from plants, termed swaposin', that inhibits the growth of food spoilage organisms. To better understand the swaposin mode of action, their structures and specific points of contact with membranes will be studied at the highest level of detail. The impact of the proposed research program will be to gain knowledge necessary for developing solutions to food-related challenges involving aspartic proteases such as food spoilage, microbial growth and food processing applications involving enzymes in varying environmental conditions.