Structure function studies on staphylokinase based plasminogen activators

Abstract

Staphylokinase (SAK), is a potent fibrin-specific plasminogen activator secreted by S. aureus that offers some distinct advantages over the other existing thrombolytic agents. As SAK is of bacterial origin, its cost of production is quite low when compared to other plasminogen activators of mammalian origin. It is the smallest known Pg-activator, existing as a single domain 15 kDa protein and therefore, it is less immunogenic than other enzymes. It acts only on the surface of the clot to form a plasmin-staphylokinase complex, only activating plasminogen trapped in the thrombus. All these above requirements, has encouraged studies with SAK regarding its ability to function as a potential thrombolytic agent. Present study on SAK and SAK based Pg activators was undertaken to gain new insights into the cofactor mediated Pg-activation mechanism by bacterial Pg activators. In-detailed characterization of S. scuiri secreted Pg-activator revealed that despite sharing sequence similarity with SAK, Pg-activation properties of SAK variants from different staphylococcal species may varies significantly and therefore, requires a wide range of experimentation and analysis to determine their host specificity and molecular mechanism of protein function. Further, our site-directed mutagenesis on SAK N-terminal region showed that Ile can be a functional substitute of N-terminal Lys in SAK with comparable catalytic efficiency. However, having SK-like Ile exposed directly at N-terminus of SAK does not imply to confer SK-like mechanism in it, there must be some other factors involved apart from N-terminal Ile that constitute to determine its Pg-activation mechanism. Structure-based analysis of SAK C-terminal region helped to generate better understanding on how C-terminal lysine residues are involved in establishing vital interactions of SAK with partner Pm during Pg activation mechanism. Lastly, through this study we developed a recombinant SAK molecule by integrating Pg-binding domain to SAK.