Structural Studies On Urokinase-type Plasminogen Activator And Its New Inhibitor

Urokinase-type plasminogen activator(uPA)is a trypsin-type serine protease.Together with its cell surface receptor(uPAR)and inhibitor(PAI-1),uPA mediates a variety of physiological rhythms on cell surface,including plasminogen activation,growth-factor activation,extracellular matrix remodeling and the initiation of intracellular signaling.As a consequence,uPA plays an important role in the occurrence and development of many diseases.The development of uPA inhibitors has been one of the research focuses,while the alkaline pharmacophores of typical uPA inhibitors lead to their poor bioavailability.As a member of flavonoids,quercetin is a flavonol and has been shown to inhibit uPA.There are no basic groups in quercetin,which indicates that quercetin may employ a different binding mode against typical uPA inhibitors,but the detailed molecular mechanism remains unknown.In this study,we determined the crystal structure of serine protease domain of uPA in complex with quercetin at a resolution of 3.0 angstroms.Structural analysis shows that quercetin binds to the major substrate binding pocket(S1 pocket)of uPA,forming a hydrogen bond between a hydroxy of quercetin B ring and Asp189 of uPA.The binding mode of quercetin differs from the typical uPA inhibitors which use the guanidino or amidine group to interact with the Asp 189 of uPA.Such distinct binding mode and unique hydroxyl-containing pharmacophore other than classical small-molecule inhibitors of serine proteases provide a new idea and direction to the development of small molecular inhibitors of uPA and other serine proteases,which would have potential applications for the serine protease-related indications,such as tumor and cardiovascular diseases.