STRUCTURAL BASIS OF LATENCY IN PLASMINOGEN-ACTIVATOR INHIBITOR-1

HuMAN plasminogen activator inhibitor-1 (PAI-1) 1,2 is the fast-acting inhibitor of tissue plasminogen activator and urokinase 3 and is a member of the serpin family of protease inhibitors 4. Serpins normally form complexes with their target proteases that dissociate very slowly as cleaved species and then fold into a highly stable inactive state 5 in which the residues that flank the scissile bond (P1 and P1'; ref. 6) are separated by about 70 angstrom (refs 7-9). PAI-1 also spontaneously folds into a stable 10 inactive state without cleavage; this state is termed 'latent' because inhibitory activity can be restored through denaturation and renaturation 2,10. Here we report the structure of intact latent PAI-1 determined by single-crystal X-ray diffraction to 2.6 angstrom resolution. The three-dimensional structure reveals that residues on the N-terminal side of the primary recognition site are inserted as a central strand of the largest beta-sheet, in positions similar to the corresponding residues in the cleaved form of the serpin alpha-1-proteinase inhibitor (alpha-1-PI) 7. Residues C-terminal to the recognition site occupy positions on the surface of the molecule distinct from those of the corresponding residues in cleaved serpins 7-9 or in the intact inactive serpin homologue, ovalbumin 11, and its cleavage product, plakal-bumin 12. The structure of latent PAI-1 is similar to one formed after cleavage in other serpins, and the stability of both latent PAI-1 and cleaved serpins may be derived from the same structural features 13,14.