Structural differences between active forms of plasminogen activator inhibitor type 1 revealed by conformationally sensitive ligands.
Plasminogen activator inhibitor type 1 (PAI-1) is a serine protease inhibitor (serpin) in which the reactive center loop (RCL) spontaneously inserts into a central beta-sheet, beta-sheet A, resulting in inactive inhibitor. Available x-ray crystallographic studies of PAI-1 in an active conformation relied on the use of stabilizing mutations. Recently it has become evident that these structural models do not adequately explain the behavior of wild-type PAI-1 (wtPAI-1) in solution. To probe the structure of native wtPAI-1, we used three conformationally sensitive ligands: the physiologic cofactor, vitronectin; a monoclonal antibody, 33B8, that binds preferentially to RCL-inserted forms of PAI-1; and RCL-mimicking peptides that insert into beta-sheet A. From patterns of interaction with wtPAI-1 and the stable mutant, 14-1B, we propose a model of the native conformation of wtPAI-1 in which the bottom of the central sheet is closed, whereas the top of the beta-sheet A is open to allow partial insertion of the RCL. Because the incorporation of RCL-mimicking peptides into wtPAI-1 is accelerated by vitronectin, we further propose that vitronectin alters the conformation of the RCL to allow increased accessibility to beta-sheet A, yielding a structural hypothesis that is contradictory to the current structural model of PAI-1 in solution and its interaction with vitronectin.
Li
Shih-Hon
SH
Department of Biochemistry, University of Illinois, Urbana, IL 61801, USA.
Gorlatova
Natalia V
NV
Lawrence
Daniel A
DA
Schwartz
Bradford S
BS
D000911:Antibodies, Monoclonal / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008954:Models, Biological; D008968:Molecular Conformation; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D017395:Plasminogen Activator Inhibitor 1 / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D013997:Time Factors; D019096:Vitronectin / Q000737:chemistry
D000911:Antibodies, Monoclonal; D008024:Ligands; D010455:Peptides; D017395:Plasminogen Activator Inhibitor 1; D019096:Vitronectin