High-level expression, purification, and enzymatic characterization of truncated @entity1 plasminogen (Lys531-Asn791) in the methylotrophic yeast XXXX .

@entity1 @entity1715 @entity10101 @entity10056 @entity4461 @entity1351 @entity563 @entity715
BACKGROUND: @entity1351 is a @entity563 protease that plays a critical role in fibrinolysis, which is a process that prevents @entity4461 from growing and becoming problematic. Recombinant @entity1 microplasminogen (rh @entity1351 ) is a derivative of @entity1351 that solely consists of the catalytic domain of @entity1 @entity1351 and lacks the five kringle domains found in the native protein. Developing an industrial production method that provides high yields of this protein with high purity, quality, and potency is critical for preclinical research. RESULTS: The @entity1 microplasminogen gene was cloned into the pPIC9K vector, and the recombinant plasmid was transformed into @entity10101 . The concentration of @entity1351 reached 510.1  mg/L of culture medium. Under fermentation conditions, the yield of rh @entity1351 was 1.0  g/L. We purified rh @entity1351 to 96  % purity by gel-filtration and cation-exchange chromatography. The specific activity of rh @entity1351 reached 23.6 U/mg. The K m of substrate hydrolysis by recombinant @entity1 microplasmin was comparable to that of @entity1 @entity1351 , while rh Plm had higher k cat /Km values than @entity1351 . The high purity and activity of the rh @entity1351 obtained here will likely prove to be a valuable tool for studies of its application in @entity715 and @entity1715 . CONCLUSIONS: Reliable rh @entity1351 production (for use in therapeutic applications) is feasible using genetically modified @entity10056 as a host strain. The successful expression of rh @entity1351 in @entity10056 lays a solid foundation for its downstream application.
