Isolation and characterization of circulating complex between human pregnancy-associated plasma protein-A and proform of eosinophil major basic protein

The plasma protein previously known as pregnancy associated plasma protein-A (PAPP-A) and believed to contain only one kind of polypeptide chain has recently been shown to be a complex containing two different chains in equimolar amounts. One of the chains is now defined as the PAPP-A submit, and the other has been identified as the proform of eosinophil major basic protein (proMBP) (Oxvig et al. (1993) J. Biol. Chem. 268, 12243–12246). A procedure for large scale preparation of the circulating complex (PAPP-A/proMBP) from pooled pregnancy serum is described. The amino acid and carbohydrate compositions of the isolated reduced and carboxymethylated PAPP-A (199 kDa) and proMBP subunits (38 kDa), and of the intact PAPP-A/proMBP have been determined. The PAPP-A and proMBP subunits contain 13.4% (w/w) and 38.6% (w/w) carbohydrate, respectively, and the intact complex contains 17.4% (w/w) carbohydrate. The PAPP-A subunit contains N -bound carbohydrate groups. In contrast, the proMBP subunit contains both N - and O -bound groups as well as glycosaminoglycan, previously found among plasma proteins only in inter-α-trypsin inhibitor and pre-α-trypsin inhibitor. It is shown that PAPP-A/proMBP can competitively inhibit human leucocyte elastase ( K I = (5–10) · 10 −9 M) at an ionic strength of 0.075, but the inhibition is negligible at ionic strengths greater than 0.15. Human cathepsin G is also competitively inhibited ( K I approx. 1 · 10 −6 M). The inhibition of both enzymes is most likely due to interactions with the glycosaminoglycan moiety of PAPP-A/proMBP. It is concluded that PAPP-A/proMBP is neither a potent nor a specific inhibitor of human leucocyte elastase.