Characterization of Brain Calpains

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Abstract
A new, simple one‐step procedure [Karlsson et al. Biochem. J. 231, 201–204 (1985)] for the separation of calpains I and II was used prior to the characterization of these enzymes from rabbit brain, using alkali‐denatured casein as the substrate. Enzyme activity was dependent on Ca2+ ions and free ‐SH groups and was maximal around pH 7.4. Incubation of calpains I and II with Ca2+ in the absence of substrate led to a rapid loss of enzyme activity. Enzyme activity was linear at room temperature and millimolar Ca2+ concentrations. However, when incubation ofcalpain I was performed with micromolar Ca2+ concentrations at room temperature proteolytic activity exhibited a lag period of approximately 10 min. This activation period was not as evident with calpain II.