Inhibition of chicken calpain II by proteins of the cystatin superfamily and alpha 2-macroglobulin was investigated. Human liver cystatins A and B, human cystatin C, chicken cystatin and rat T-kininogen were found not to be inhibitory. Inhibition was, however, observed for bovine and rat kininogens, with Ki (inhibition constant) values of 0.8 nM and 30 nM respectively. alpha 2-Macroglobulin inhibits calpain with an initial rate constant of the order of 3 X 10(4) M-1.S-1. Calpain complexed with alpha 2-macroglobulin showed only limited reactivity towards azocasein, but reacted readily with the peptide substrate Suc-Leu-Tyr-4-methyl-7-coumarylamide and with L-3-carboxy-trans-2,3-epoxypropionyl-leucylamido-(4-guanidin o)butane (E-64). The calpain in the complexes was at least partially protected from loss of activity due to autolysis. The calpain-alpha 2-macroglobulin complexes contained both the calpain subunits.
