Vampire bat salivary plasminogen activators (DSPAs) are thrombolytic agents that are under clinical investigation for the treatment of acute ischemic stroke. In, this study, the synthetic active salivary plasminogen activator alpha 2 (DSPA alpha 2) gene optimized for the preferred codons of Pichia pastoris was assembled from 48 oligonucleotides, and cloned into the yeast expression vector pPIC9 with a strong enhancer from human cytomegalovirus, (HCMV),. This system achieved. high expression of an active DSPA alpha 2 in P. pastoris yeast GS115. secreted active DSPA alpha 2 recombinant protein was purified from broth supernatant by a simple one-step procedure on Sephadex chromatography and was confirmed by SDS-PAGE and Western blot analysis. ELISA showed that 2.5 ring of recombinant protein could be obtained from 100-ml culture broth supernatant. The fibrinolytic activity of the recombinant DSPA alpha 2 was 1.28 x 10(5) IU/mg. (c) 2007 Published by Elsevier Inc.