N. Sargianos et al., STUDIES ON THE AUTOLYSIS OF M-CALPAIN FROM THE SKELETAL-MUSCLE OF THEAMPHIBIAN RANA-RIDIBUNDA, The Journal of experimental zoology, 271(2), 1995, pp. 82-94
The autolytic mechanisms responsible for the regulation of m-calpain p
urified from the skeletal muscle of the amphibian Rana ridibunda were
examined. Both subunits of the calpain molecule were found to undergo
autolysis in the presence of Ca2+. Various divalent cations were exami
ned for their ability to induce calpain autolysis. The concentrations
of these cations required for the complete calpain autolysis were: 500
mu M Ca2+, 800 mu M Mn2+, 2 mM Sr2+, 10 mM Ba2+, whereas Mg2+, even a
t 10 mM did not induce any autolysis. Calpain autolysis induced by the
above divalent cations is a temperature dependent process. Presence o
f Mn2+ or Sr2+ reduces the Ca2+ requirement of calpain for autolysis.
The rate of autolysis depends on the protease concentration; protease
inhibitors such as E-64, leupeptin, antipain, and iodoacetic acid inhi
bit the autolysis of calpain; E-64 inhibits irreversibly while leupept
in inhibits reversibly the autolysis; and irreversibly inactivated by
E-64 calpain is fully digested by native calpain. Autolysis of calpain
in the presence of alkali denatured casein increases the Ca2+ sensiti
vity of the protease for its half maximal and maximal caseinolytic act
ivity. Limited autolysis of calpain is also induced in the presence of
the endogenous substrate G-actin, and the rate of autolysis is slower
than that obtained in the absence of substrates. (C) 1995 Wiley-Liss,
Inc.