Recombinant Batroxobin Expressed Highly in Pichia pastoris

Methylotrophic yeast, Pichia pastoris, was used to express recombinant batroxobin, and a technology for producing a recombinant protein was finally established. We synthesized batroxobin gene artificially by means of recursive PCR. pPIC9-batroxobin was constructed and transformed into P. pastoris GS115 (his4). Recombinant batroxobin was expressed in yeast engineering strain and purified from the culture supernatant. 10 mg of recombinant batroxobin was purified from 1 L fermentation media, exhibiting specific activity of 238 NIH units/mg, and had molecular weight of 30.55 kD. The purified recombinant protein converted fibrinogen into fibrin clot in vitro and shortened bleeding time in vivo. The study laid a foundation for the development of hemostatic of recombinant snake venom thrombin-like enzyme (SVTLE).