Abstract
The stoichiometry of the individual steps, i.e. polypeptide chain cleavage, hydrolysis of the putative thioester bond and conformational change, of the reaction between alpha 2-macroglobulin and trypsin or chymotrypsin was analysed. The chain cleavage was monitored by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, the thioester hydrolysis by both a spectroscopic and a fluorimetric technique and the conformational change by tryptophan fluorescence. A stoichiometry of close to 2:1 was obtained for all reactions. This finding indicates that the alpha 2-macroglobulin half-molecule is an independent functional unit of the inhibitor, within which co-operativity between the two subunits may occur.