PURIFICATION, CHARACTERIZATION, AND CDNA CLONING OF A KUNITZ-TYPE PROTEINASE-INHIBITOR SECRETED BY THE PORCINE UTERUS

The porcine uterus synthesizes a proteinase inhibitor (M(r) 14,000) un der the influence of progesterone that is relatively specific for plas min and trypsin, but that also has weak affinity for chymotrypsin. Sev eral isoforms of this uterine plasmin/trypsin inhibitor were purified by a procedure whose final two steps involved affinity chromatography on immobilized chymotrypsin and cation exchange chromatography. Amino- terminal sequencing showed that at least three of the isoforms were cl osely related. An oligonucleotide probe based on the protein sequence was used to identify a cDNA that contained an open reading frame codin g for a mature protein (M(r) 10,295) of 98 amino acids. The inhibitor had a well defined, but unique, Kunitz domain of 64 residues at its am ino terminus that shared 67% sequence identity to bovine pancreatic tr ypsin inhibitor. Its P-1 residue was arginine rather than lysine. Nort hern analysis showed the presence of a single mRNA species (700 bases) that in adult female pigs appeared to be confined to the uterus. Duri ng pregnancy, UPTI mRNA expression was high until Day 30 and decreased significantly thereafter. By contrast, uteroferrin mRNA reached maxim al concentrations in late pregnancy. These data are consistent with an earlier hypothesis that the inhibitor serves to neutralize the activi ties of one or more serine proteinases generated by the proliferating trophoblast during the formation of the noninvasive placenta of the pi g.