One strategy to study folding intermediates is to identify protein fragments or peptide models that fold autonomously. Fragments of proteins having lost some of the tertiary interactions that hold them in contact with other amino acids in the uncleaved protein could mimic the conformation of the early intermediates of folding.
The initial aim of this project is to use a variety of techniques, including hydrogen exchange labelling coupled to NMR, and electrospray mass spectrometry, and combined with stopped-flow methods and NMR experiments, to examine the folding of a variety of protein fragments from two fibrinolytic multidomain proteins: urokinase and streptokinase. Urokinase-type plasminogen activator (u-PA), is a serine protease whose principal function appears to be the activation of plasminogen to plasmin by specific proteolytic cleavage. Streptokinase (SK) is a bactoferial secretory protein and also a potente activator of plasminogen. Both proteins are being widely used as a thrombolytic agents in treatment of acute myocardial infarction. A detailed understanding of domain interactions in these proteins will aid the design of novel therapeutic agents.
Virtually nothing is known about the folding of multidomain proteins. Once the work on fragments of the proteins is complete, studies on the intact proteins can be initiated to explore the mean in which such proteins are assembled.