In vitro Proteolytic Degradation of Bovine Brain Calcineurin by m-Calpain |
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Authors: | Lakshmikuttyamma Ashakumary Selvakumar Ponniah Ratan Sharma Anil Anderson Deborah H Sharma Rajendra K |
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Institution: | Department of Pathology, College of Medicine, University of Saskatchewan, Saskatchewan Cancer Agency, Saskatoon, Saskatchewan, Canada S7N 4H4. |
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Abstract: | A major cause of neuronal dysfunction is due to altered Ca2+ regulation. An increase in Ca2+ influx can activate Ca2+-dependent enzymes including calpains, causing the proteolysis of its specific substrates. In the present study, calcineurin (CaN) was found to be proteolysed by a Ca2+-dependent cysteine protease, m-calpain. In the presence of Ca2+, the 60 kDa subunit (CaN A) was degraded to a 46 kDa immunoreactive fragment, whereas in the presence of Ca2+ /calmodulin (CaM) immunoreactive fragments of 48 and 54 kDa were observed. The beta-subunit (CaN B) was not proteolysed in either condition. The proteolysis of CaN A increased its phosphatase activity and rendered it totally CaM-independent after 10 min of proteolysis. The molecular weight of the proteolytic fragments suggested that the m-calpain cleaved CaN A in the CaN B binding domain. A CaM-overlay experiment revealed that the CaM-binding site was present only in the 54 kDa fragment produced by CaN A proteolysis in the presence of Ca2+ /CaM. Thus, the increase in CaN A phosphatase activity observed in many neuronal disorders, may be due to the action of calpain. |
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Keywords: | Ca2+ calcineurin calmodulin calpain |
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