Covalent interaction between proform of eosinophil major basic protein (proMBP) and pregnancy-associated plasma protein-A (PAPP-A) is a cell-mediated event and required for proMBP inhibition of the catalytic activity of PAPP-A

This study was undertaken to determine the mechanism by which proform of eosinophil major basic protein (proMBP) inhibits the IGFBP-4 proteolytic activity of pregnancy-associated plasma protein (PAPP)-A. Co-overexpression of PAPP-A with proMBP in 293T cells, or co-incubation of 293T cells, respectively, overexpressing proMBP and PAPP-A resulted in the formation of a covalent proMBP-PAPP-A complex and inhibition of IGFBP-4 proteolysis. Similar results were obtained when recombinant proMBP and PAPP-A were incubated in the presence of U2 osteosarcoma cells or when recombinant proMBP was added to the U2 cells overexpressing PAPP-A. In contrast, no formation of covalent proMBP-PAPP-A complex or inhibition of IGFBP-4 proteolysis was observed when recombinant proMBP and PAPP-A were incubated under cell-free conditions, although proMBP was able to interact with PAPP-A in a non-covalent manner. These new findings suggest that formation of covalent proMBP-PAPP-A complex is a cell-mediated event and is required for proMBP to inhibit the catalytic activity of PAPP-A.