Submitted to: Journal of Animal Science
Publication Type: Peer reviewed journal
Publication Acceptance Date: 2/12/2001
Publication Date: N/A
Citation: Interpretive Summary: During refrigerated storage of cuts of meat or whole carcasses, meat will gradually tenderize. The reason for this tenderization is the breakdown of proteins that are responsible for structural integrity of meat. While postmortem tenderization occurs in most carcasses, it does not occur in all carcasses. The reason for inconsistent meat tenderness is the variation in the activity of the enzyme system that is responsible for meat tenderization. The enzyme system is called the calpain proteolytic system and it has three components, two enzymes (u-calpain, which is active at low calcium concentration, and m-calpain, which is active at high calcium concentration) and one inhibitor (calpastatin). There is disagreement within the scientific community on whether u-calpain, m-calpain, or both, are involved in postmortem meat tenderization. Because the two calpains are so similar, it has been very difficult to determine which one plays a key role in tenderization. In this study we used a more sensitive assay than the standard assay for calpain activity. The results from this more sensitive assay provide stronger evidence that u-calpain, not m-calpain, is responsible for postmortem meat tenderization. Hence, understanding the regulation of u-calpain in postmortem meat should be the focus of future studies.
Technical Abstract: Casein zymography was used to determine the effect of postmortem storage on the proteolytic activity of u-calpain and m-calpain in lamb longissimus. Casein zymography is a method to quantify calpain activity in non-denaturing casein-containing polyacrylamide gels. Casein zymography assays were conducted on crude muscle extracts (only one centrifugation). Six market weight crossbred lambs were slaughtered and a portion of the longissimus lumborum was removed at death (within 15 min of exsanguination) and after 3, 6, 9, 12, 24, 72, and 360 h postmortem. Muscle samples were snap frozen in liquid nitrogen and stored at minus 70 deg C. There was a gradual decline in u-calpain activity (P < 0.05) such that after 24 h and 72 h postmortem, u-calpain had lost 42% and 95% of its activity, respectively. After 360 h postmortem no u-calpain activity could be detected (under the conditions used in this study). Unlike u-calpain, postmortem storage had no effect on m-calpain (P > 0.05). When the calcium concentration of a muscle extract was increased to the level that induces m-calpain autolysis, m-calpain was autolyzed and its autolysis was readily detected by the in-gel casein assay. Collectively, these results demonstrate that calcium concentration in postmortem muscle is only high enough to activate u-calpain. These results support the widely believed conclusion that u-calpain-mediated proteolysis of key myofibrillar and cytoskeletal proteins is responsible for postmortem tenderization. Hence, understanding the regulation of u-calpain in postmortem muscle should be the focus of future studies.