Covalent and non‐covalent interaction of chymotrypsin with α2‐macroglobulin

Abstract

The pattern of covalent crosslinking between human α₂-macroglobulin (α₂M) and chymotrypsin has been investigated by chromatography and polyacrylamide gel electrophoresis in denaturing medium. Reaction with a single mol of chymotrypsin per mol α₂M results in the formation of a 95% covalent 1:1 chymotrypsin-α₂M complex and in the proteolytic cleavage of both 180 kDa monomers in one α₂M subunit. Proteolytic cleavage in the other α₂M subunit requires the presence of a second mol of chymotrypsin; part (20%) of the protease in the 2:1 chymotrypsin-α₂M complex thus formed appears to be non-covalently bound to the α₂M chains. Covalent binding is abolished when the reaction of α₂M with the protease is carried out in the presence of hydroxylamine. A single mol of the protease is then able to cleave all four 180 kDa monomers in α₂M.