RESISTANCE TO INHIBITION BY ALPHA-1-ANTI-TRYPSIN AND SPECIES-SPECIFICITY OF A CHIMERIC HUMAN BOVINE PROTEIN-C/

Human activated protein C (APC) has been shown to be physiologically s usceptible to inhibition by the abundant serpin inhibitor alpha-1-anti -trypsin (AAT). Studies on the inactivation by AAT [Heeb, M. J., and G riffin, J. H. (1988) J. Biol. Chem. 263, 11613-11616] have shown that the calculated rate of this inactivation matches that of the observed half-life of APC in vivo [Wydro, R., Oppenheimer, C., Rodger, R., and Miemi, S. (1988) Clin. Res. 36, 329A] and complex formation therefore probably represents a physiologic regulation process for APC. In this study we observed that bovine APC, in contrast to human APC, is nearly completely resistant to inactivation by human AAT. An additional diff erence between human and bovine APC is that human APC is a potent anti coagulant in human plasma, whereas bovine APC is only minimally active in human plasma. These functional differences exist despite considera ble structural similarity between the human and bovine molecules. In o rder to identify specific molecular regions responsible for function, a chimeric molecule consisting of the light chain of human protein C ( PC) and the heavy chain of bovine PC was constructed, expressed, and c haracterized. The activated chimeric PC is similar to human APC in hav ing potent anticoagulant activity in human plasma, but displays nearly identical resistance to AAT inhibition with the bovine molecule. The similarity between the chimeric and bovine molecules in resistance to AAT inhibition indicates that the structural determinants for inhibito r interactions reside within the heavy chain (serine protease) domain. The conservation of anticoagulant activity in human plasma between th e chimeric and human molecules further indicates that structural deter minants which confer species specificity to APC reside within the ligh t chain of the molecule.