Expression of streptokinase in Pichia pastoris yeast

Streptokinase is a potent plasminogen activator with widespread clinical use as a thrombolytic agent. It is naturally secreted by several strains of hemolytic streptococci. We report here the expression of streptokinase in the methylotrophic yeast Pichia pastoris. The structural gene encoding streptokinase, minus the bacterial signal sequence, was fused to the methanol-inducible alcohol oxidase promoter from P. pastoris. Upon induction on methanol, strains of P. pastoris transformed with the recombinant plasmid pHTskc25 produced a 47-kD protein (the size of native streptokinase) which was reactive with polyclonal antibodies raised against native streptokinase. Lysates of transformed cells induced on methanol activate plasminogen, as determined by casein degradation on agar plates and by hydrolysis of a synthetic ester substrate. During continuous fermentation on methanol, the streptokinase gene was stably maintained and expressed and no adverse effects on cell growth were observed.