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KMID : 1094720120170051068
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Biotechnology and Bioprocess Engineering
2012 Volume.17 No. 5 p.1068 ~ p.1075
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Production of salmosin, a snake venom-derived disintegrin, in recombinant Pichia pastoris using high cell density fed-batch fermentation
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Seo Myung-Ji
Choi Hak-Jong
Chung Kwang-Hoe
Pyun Yu-Ryang
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Abstract
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Salmosin, a snake venom-derived disintegrin, was successfully expressed in the methylotrophic yeast Pichia pastoris and secreted into the culture supernatant, as a 6 kDa protein. High-cell density fermentation of recombinant P. pastoris was optimized for the mass production of salmosin. In a 5 L jar fermentor, recombinant P. pastoris was fermented in growth medium containing 5% (w/v) glycerol at the controlled pH of 5.0. After culturing for 21 h, glycerol feeding medium was fed at one time into the culture broth. After 7 h (a total of 28 h), induction medium that contained methanol was increasingly added until the culture time totaled 75 h. Finally, these optimized culture conditions produced a high cell density of recombinant P. pastoris (dry cell weight of 113.38 g/L) and led to the mass production of salmosin (a total protein concentration of 369.2 mg/L). The culture supernatant containing salmosin inhibited platelet aggregation, resulting in a platelet aggregation of 9% compared to that of 94% in the control experiment, without culture supernatant. These results demonstrate that recombinant salmosin in culture supernatant from high cell density fed-batch fermentation can serve as a platelet aggregation inhibitor.
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KEYWORD
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salmosin, platelet aggregation inhibitor, Pichia pastoris, high cell density fed-batch fermentation
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