A calcium-activated protease which preferentially degrades the 160-kDa component of the neurofilament triplet.
A calcium-dependent protease fully active with 0.2 mM Ca2+ was found associated with the neurofilament-enriched cytoskeleton of the rat spinal cord prepared by the treatment with Triton X-100. The enzyme preferentially degrades the 160-kDa component of the neurofilament triplet. In addition, a soluble calcium-dependent protease activity was found in the supernatant from the spinal cord, which degraded a variety of cytoskeletal proteins including the neurofilament triplet, glial fibrillary acidic (GFA) protein, actin, tubulin, and a high molecular weight protein associated with microtubules. The possibility that the cytoskeleton-bound activity is an artefactual association of the soluble enzyme to the cytoskeleton seems to be negated on the basis of the following dissociation and reassociation experiments. The protease activity remained associated with the cytoskeleton in the physiological ionic strength, and was not completely dissociated from it until the KC1 concentration was raised to 0.6 M. When the 0.6 M KCl-extract was dialysed against salt-free buffer to remove KC1, and added back to the protease-free cytoskeletal pellet, proteolytic activity was partially restored. Full activity returned only when the extract and the protease-free cytoskeletal pellet were first combined in the presence of 0.6 M KC1, and then slowly reassociated by dialysis against salt-free buffer. Dissociated enzyme was rapidly inactivated at 37 degrees C in the presence of Ca2+. These results suggest the structural association of the protease with the cytoskeleton under the physiological condition.
D000818:Animals; D002154:Calpain; D003599:Cytoskeleton / Q000201:enzymology*; D010450:Endopeptidases / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D008970:Molecular Weight; D010455:Peptides / Q000378:metabolism; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D013116:Spinal Cord / Q000201:enzymology
D010455:Peptides; D008345:Manganese; D011189:Potassium Chloride; D010450:Endopeptidases; D002154:Calpain; D008274:Magnesium