Pregnancy zone protein, a proteinase-binding macroglobulin. Interactions with proteinases and methylamine

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Abstract
Human pregnancy zone protein (PZP) is a major pregnancy-associated plasma protein, strongly related to .alpha.2-macroglobulin (.alpha.2M). Its properties and its reactions with a number of enzymes, particularly chymotrypsin, and with methylamine have been investigated. It is concluded that native PZP molecules are dimers of disulfide-bridged 180-kDa subunits and that proteinase binding results in covalent 1:1 (tetrameric)PZP-enzyme complexes. Native PZP is unstable, and storage should be avoided, but when kept unfrozen at 0.degree.C most PZP preparations stay native 1-3 months. The reaction of PZP with chymotrypsin involves (i) proteolysis of bait regions, (ii) cleavage of .beta.-cysteinyl-.gamma.-glutamyl thiol ester groups, (iii) some change of the conformation and quaternary structure of PZP, and (iv) the formation of covalent 1:1 chymotrypsin-PZP(tetramer) complexes in which chymotrypsin is active but shows less activity than free chymotrypsin. The emission spectra of intrinsic fluorescence show significant differences between the PZP-chymotrypsin complex and its native components, whereas no differences are observed between methylamine-reacted PZP and native PZP. Methylamine reacts with the .beta.-cysteinyl-.gamma.-glutamyl thiol ester groups of PZP in a second-order process with k = (13.6 .+-. 0.5) M-1 s-1, pH 7.6, 25.degree.C. The reaction product is PZP(dimers); no PZP(tetramers) are formed. The proteinase-binding specificity of PZP is far more restricted than that .alpha.2M. Certain chymotrypsin-like and trypsin-like enzymes are bound much less efficiently than is chymotrypsin itself. It is suggested that one reason for this is the importance of the covalent binding of the enzyme to PZP, which is mainly determined by the content of lysine residues of the enzyme. Another reason is the importance of bait region cleavage, which is determined by the substrate specificity of the enzyme. PZP is analogous to .alpha.2-M, but also distinct in several respects. (i) The native molecule is a dimer. (ii) The tap mechanism is not at work. Only covalently bound, sterically inhibited proteinase-(tetramer)PZP complexes are formed. (iii) No apparent major change of conformation is triggered by cleavage of the .beta.-cysteinyl-.gamma.-glutamyl thiol esters alone. (iv) Change of conformation requires interaction with a proteinase.