Effect of calpastatin on degradation of myofibrillar proteins by mu-calpain under postmortem conditions.
To improve our understanding of the regulation of mu-calpain activity in situ during postmortem storage of muscle, the effect of different calpastatin levels on proteolysis of myofibrillar proteins by mu-calpain in a system closely mimicking postmortem conditions was studied. Increasing the amount of calpastatin in the incubations limited both the rate and extent of proteolysis of myofibrillar proteins and autolysis of mu-calpain. Excess calpastatin (i.e., a mu-calpain:calpastatin ratio of 1:4) did not inhibit proteolysis completely. Western blot analysis revealed that proteolysis of myofibrillar proteins virtually ceased after 7 d of incubation, despite the presence of partly autolyzed, therefore seemingly active, mu-calpain. A series of incubations of autolyzed mu-calpain revealed that the autolyzed form of this enzyme is unstable at an ionic strength observed in postmortem muscle. The possible significance of these results in terms of the regulation of mu-calpain activity in postmortem muscle is discussed.
Geesink
G H
GH
Roman L. Hruska US Meat Animal Research Center, ARS, USDA, Clay Center, NE 68933-0166, USA.
Koohmaraie
M
M
D000818:Animals; D015153:Blotting, Western / Q000662:veterinary; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002154:Calpain / Q000037:antagonists & inhibitors*; D002417:Cattle; D015853:Cysteine Proteinase Inhibitors / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000037:antagonists & inhibitors*; D009124:Muscle Proteins / Q000378:metabolism*; D009210:Myofibrils / Q000378:metabolism*; D011180:Postmortem Changes
D002135:Calcium-Binding Proteins; D015853:Cysteine Proteinase Inhibitors; D007527:Isoenzymes; D009124:Muscle Proteins; C033668:calpastatin; D002154:Calpain