Plasminogen activator inhibitor 1. Structure of the native serpin, comparison to its other conformers and implications for serpin inactivation

The crystal structure of a constitutively active multiple site mutant of pl asminogen activator inhibitor 1 (PAI-1) was determined and refined at a res olution of 2.7 Angstrom. The present structure comprises a dimer of two crystallographically indepen dent PAI-1 molecules that pack by association of the residues P6 to P3 of t he reactive centre loop of one molecule (A) with the edge of the main beta- sheet A of the other molecule (B). Thus, the reactive centre loop is ordered for molecule A by crystal packing forces, while for molecule B it is unconstrained by crystal packing contac ts and is disordered. The overall structure of active PAI-1 is similar to the structures of other active inhibitory serpins exhibiting as the major secondary-structural fea ture a five-stranded beta-sheet A and an intact proteinase-binding loop pro truding from the one end of the elongated molecule. No preinsertion of the reactive centre loop is observed in this structure. A comparison of the present structure with the previously determined crysta l structures of PAI-1 in its alternative conformations reveals that, upon c leavage of an intact form of PAI-1 or formation of latent PAI-1, the well-c haracterised rearrangements of the serpin secondary structural elements are accompanied by dramatic and partly unexpected conformational changes of he lical and loop structures proximal to beta-sheet A. The present structure explains the stabilising effects of the mutated resid ues, reveals the structural cause for the observed spectroscopic difference s between active and latent PAI-1, and provides new insights into possible mechanisms of stabilisation by its natural binding partner, vitronectin. (C ) 2000 Academic Press.