Free calcium and calpain I activity.
Activation of purified calpain I proceeds through a Ca(2+)-induced autolysis from the 80 kDa catalytic subunit to a 76 kDa form via an intermediate 78 kDa form, and from a 30 kDa form to a 18 kDa form as the result of two autocatalytic processes (intra and intermolecular). The minimum Ca2+ requirements for autolysis and proteolysis have been determined by physico-chemical and electrophoretic methods in the presence or absence of a digestible substrate. According to our results the activation process needs less free Ca2+ than the proteolysis of a digestible substrate, which means that proteolysis is really subsequent to activation. For very low Ca2+ levels, a digestible substrate does not initiate the calpain I activation process. In the presence of phospholipid vesicles, such as PI, PS or a mixture of PI (20%), PS (20%) and PC (60%), the apparent kinetic constants of activation are greatly increased without any change in the initial velocity of the substrate proteolysis. Thus, enzyme activation and substrate proteolysis are observed as independent phenomena. These results obtained from experiments using low free Ca2+ concentrations enable us to propose a hypothesis for the mechanism of regulation by which the enzyme could be activated in the living cell.
Cottin
P
P
Laboratoire de Biochimie et Technologie des Aliments, ISTAB, Université Bordeaux I, Talence, France.
Poussard
S
S
Desmazes
J P
JP
Georgescauld
D
D
Ducastaing
A
A
D001329:Autolysis; D002118:Calcium / Q000494:pharmacology*; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D005914:Globins / Q000494:pharmacology; D007700:Kinetics; D010743:Phospholipids / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism; D014746:Vimentin / Q000494:pharmacology
D010743:Phospholipids; D011506:Proteins; D014746:Vimentin; D005914:Globins; D002154:Calpain; D002118:Calcium