Mechanisms and Integration of Signal Pathway: A Role for Calpains?

In order to survive cells must sense and respond to changes in their environment. Environmental cues trigger a variety of events within cells. The concentration and movements of calcium ions are essential regulators of many of these cellular responses. Proper control of intracellular calcium is essential because at thigh levels calcium can lead to cell damage or death. Calcium accomplishes it effects through binding to specific proteins such as calmodulin and calpain. Calmodulin, named for its ability to bind calcium and to modulate the activity of other cellular components, is an important mediator of calcium signals and its mechanism of action is relatively well understood. The calpains are proteolytic enzymes that are regulated by calcium binding. Proteolytic enzymes modify cellular proteins such that the target protein is destroyed or is altered in its function. Calpains can modify those proteins, called pumps or channels, that directly allow calcium ion movement and therefore calpains may be involved directly in helping cells to regulate calcium signals. Through its action on other key regulator proteins and enzymes, calpains might also help to integrate cellular responses that involve calcium with other events that do not. Although the activity of calpain is controlled by calcium its action may be prevented by another protein (calpastatin) that specifically inhibits these enzymes. Although calpastatin does not bind calcium directly it binds to calpain only when calpain has bound calcium. Thus the interactions between these two proteins is important in regulating calpain function. By comparison with calmodulin, there is little known about how calcium regulates calpain structure, function or its interactions with other proteins. Thus the immediate goals for this proposal are to understand more about the biochemical mechanisms that regulate calpain. This project aims to answer two major questions: 1) how does calcium binding to calpain regulate the enzymatic activity? 2) can a calpastatin insensitive calpain be generated by exploiting what is known about the interaction between calpain and calpastatin? An enzyme with this property will be useful, in future studies, for determining the role of calpains in cells. These goals will be accomplished through techniques of protein chemistry and expression of recombinant DNA molecules to generate modified enzymes or enzyme fragments for study. This knowledge will provide information on the role of calpain in critical cellular processes. %%% Calcium is important in the regulation of many essential intracellular processes. Many of the effecPts of calcium occur after it binds to specific proteins. One of these proteins is calpain. Calpains are proteolytic enzymes that are regulated by calcium binding, and in turn modify other cellular proteins in ways which often alter their function. Little is known of the mechanism by which calcium modifies calpain, and this project will study the biochemical mechanisms that regulate calpain and the role of calcium in these processes