ISOLATION AND CHARACTERIZATION OF CIRCULATING COMPLEX BETWEEN HUMAN PREGNANCY-ASSOCIATED PLASMA PROTEIN-A AND PREFORM OF EOSINOPHIL MAJOR BASIC-PROTEIN

The plasma protein previously known as pregnancy associated plasma pro tein-A (PAPP-A) and believed to contain only one kind of polypeptide c hain has recently been shown to be a complex containing two different chains in equimolar amounts. One of the chains is now defined as the P APP-A subunit, and the other has been identified as the preform of eos inophil major basic protein (proMBP) (Oxvig et al. (1993) J. Biol. Che m. 268, 12243-12246). A procedure for large scale preparation of the c irculating complex (PAPP-A/proMBP) from pooled pregnancy serum is desc ribed. The amino acid and carbohydrate compositions of the isolated re duced and carboxymethylated PAPP-A (199 kDa) and proMBP subunits (38 k Da), 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 gl ycosaminoglycan, previously found among plasma proteins only in inter- alpha-trypsin inhibitor and pre-alpha-trypsin inhibitor. It is shown t hat 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 inhibiti on is negligible at ionic strengths greater than 0.15. Human cathepsin G is also competitively inhibited (K-I approx. 1.10(-6) M). The inhib ition of both enzymes is most likely due to interactions with the glyc osaminoglycan moiety of PAPP-A/proMBP. It is concluded that PAPP-A/pro MBP is neither a potent nor a specific inhibitor of human leucocyte el astase.