Selection of Optimum Expression System for Production of Kringle Fragment of Human Apolipoprotein(a) in Saccharomyces cerevisiae
Recombinant Saccharomyces cerevisiae expression systems were developed to produce a novel human anti-angiogenic protein called LK8, an 86 amino-acid kringle fragment protein with three disulfide linkages. Galactose-inducible LK8 expression plasmid was constructed, and LK8 production levels by four S. cerevisiae strains were compared in order to select an optimal host strain. S. cerevisiae 2805 was the most efficient among the strains tested. Elevating the LK8 gene copy number through multiple integration using 8-sequences as target sites resulted in more than a two-fold increase in the LK8 production level compared with the plasmidbased expression system. The maximum LK8 protein concentration of 25 mg/L was obtained from batch cultivation of the yeast transformant that harbors 16 copies of the LK8 gene. In conclusion, the strain integrated with the multiple LK8 gene secreted the protein with relatively high yield, although, the increased LK8 gene dosage over 11 copies did not lead to further enhancement in batch cultivations.
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