Abstract

Heterodimeric calpain, consisting of the large (80 kDa) and the small (30 kDa) subunit, was isolated and purified from human erythrocytes by a highly reproducible fourstep purification procedure. Obtained material is more than 95% pure and has a specific activity of 6 7 mU/mg. Presence of contaminating proteins could not be detected by HPLC and sequence analysis. During storage at 80 C the enzyme remains fully activatable by Ca 2+ , although the small subunit is partially processed to a 22 kDa fragment. This novel autolysis product of the small subunit starts with the sequence 60 RILG and is further processed to the known 18 kDa fragment. Active forms and typical transient and stable autolysis products of the large subunit were identified by protein sequencing. In caseinzymograms only the activatable forms 80 kDa+30 kDa, 80 kDa+22 kDa and 80 kDa+18 kDa displayed caseinolysis.