Development Of Peptidomimetics Of Staphylokinase

Intravascular thrombosis is one of the main causes of a wild variety of cardiovascular diseases. The agents currently used in the clinics include tissue-type plasminogen activator (t-PA), streptokinase, urokinase and their variants. These thrombolytic agents can effectively reduce the mortality and disability, however, they still have the shortcomings such as incomplete recanalization, delayed reperfusion time, reocclusion, bleeding, allergic reaction etc. Staphylokinase (Sak), is a promising thrombolytic agent at least equipotent to t-PA for coronary artery recanalization, significantly more fibrin selective and obtainable with high yield in Escherichia coli. Unfortunately, the wt-Sak(wild type) may still cause quite severe allergic reactions in rats, so do the patients, which therefore hampered its clinical application.Plasminogen activation is a key event in the fibrinolytic system that results in the dissolution of blood clots, and also promotes cell migration and tissue remodelling. The recent structure determinations of microplasmin in complex with the bacterial plasminogen activators staphylokinase and streptokinase have provided novel insights into the molecular mechanisms of plasminogen activation and cofactor function. These bacterial proteins are cofactor molecules that contribute to exosite formation and enhance the substrate presentation to the enzyme. At the same time, they modulate the specificity of plasmin towards substrates and inhibitors, making a 'specificity switch' possible.To develop novel thrombolytic agent with reduced immunogenicity as well as an oral preventive of thrombosis, many efforts have been fulfilled: from the usage of aspirin to scu-PA and APSAC, etc. However, none of them gives satisfactory answer. Here we present the idea that by reducing the molecular weight of Sak while keeping its thrombolytic activity, we may find out novel agonist of Plasminogen(Plg).During our earlier works, we found that wt-Sak may reserve quite a lot of activities in the presence of denaturant such as 6M of carbamidine chloride or 8M of urea, suggesting there may be some functional epitopes within the molecule of Sak, which offer the dorminating contributions in the interaction of Sak and Plg.The most important natural sources of new leads are plant extracts, bacterialbroths, animal venoms and peptides isolated from living organisms. However, only the three first have been used extensively in the development of new therapeutic agents. This is probably due to the low pharmacological profile exhibited by peptides, that requires a lengthy transformation to make them suitable as new leads. In contrast, bioactive compounds isolated from the other sources are regularly closer to be used as lead compounds. Nevertheless, the sources for compounds of this category are nowadays scarce. In contrast, there are new bioactive peptides discovered quite often and reported as ligands for different receptors. Under these circumstances peptides appear as an attractive source of prospective new leads.Using the phage-displayed random peptide library, we obtained several peptide sequences after biopanning with uPlg as the target protein. Thse high affinity peptide was then synthesized using solid phase synthesis method, which followed with the purification by HPLC and then the identification by MS analysis, then, the peptides were used for activity assay. Based on the analysis of the acivity assay, the computer aided design was carried out to make modifications to the peptide in order to improve its thrombolytic potent.