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Optimization of the pilot-scale production of an ice-binding protein by fed-batch culture of Pichia pastoris

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Title
Optimization of the pilot-scale production of an ice-binding protein by fed-batch culture of Pichia pastoris
Other Titles
피키아 발현시스템을 이용한 LeIBP 결빙방지단백질의 대량생산 최적화 연구 (공동 제1저자)
Authors
Lee, Jun Hyuck
Kim, Hak Jun
Han, Se Jong
Yong-Hoe Choe
Eunjung Kim
Jong Chan Park
Do, Hackwon
Lee, Sung Gu
Subject
Biotechnology & Applied Microbiology
Keywords
Fed-batch culture; Ice-binding protein; LeIBP; Leucosporidium sp.; Pichia pastoris
Issue Date
2013
Publisher
Springer
Citation
Lee, Jun Hyuck, et al. 2013. "Optimization of the pilot-scale production of an ice-binding protein by fed-batch culture of Pichia pastoris". APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 97(8): 3383-3393.
Abstract
Ice-binding proteins (IBPs) can bind to the ice crystal and inhibit its growth. Because this property of IBPs can increase the freeze-thaw survival of cells, IBPs has attracted the attention from industries for their potential use in biotechnoogical applications. However, their use was largely hampered by the lack of the large-scale recombinant production system. In this study, the codon-optimized IBP from Leucosporidium sp. (LeIBP) was constructed and subjected to high-level expression in methylotrophic Pichia pastoris sytem. In a laboratory-scale fermentation (7 L), the optimal induction temperature and pH were determined to be 25°C and 6.0, respectively. Further, employing glycerol fed-batch phase prior to methanol induction phase enhanced the production of recombinant LelBP (rLeIBP) by ~100 mg/L. The total amount of secreted proteins at these conditions (25°C, pH 6.0, and glycerol fed-batch phase) was ~443 mg/L, 60 % of which was rLeIBP, yielding ~272 mg/L. In the pilot scale fermentation (700 L) under the same conditions, the yield of rLeIBP was 300 mg/L. To our best knowledge, this result reports the highest production yield of the recombinant IBP. More importantly, the rLeIBP secreted into culture media was stable and active for six days of fermentation. The thermal hysteresis (TH) activity of rLeIBP was about 0.42°C, which is almost the same to those reported previously. The avaotechnoogical applications. However, their use was largely hampered by the lack of the large-scale recombinant production system. In this study, the codon-optimized IBP from Leucosporidium sp. (LeIBP) was constructed and subjected to high-level expression in methylotrophic Pichia pastoris sytem. In a laboratory-scale fermentation (7 L), the optimal induction temperature and pH were determined to be 25°C and 6.0, respectively. Further, employing glycerol fed-batch phase prior to methanol induction phase enhanced the production of recombinant LelBP (rLeIBP) by ~100 mg/L. The total amount of secreted proteins at these conditions (25°C, pH 6.0, and glycerol fed-batch phase) was ~443 mg/L, 60 % of which was rLeIBP, yielding ~272 mg/L. In the pilot scale fermentation (700 L) under the same conditions, the yield of rLeIBP was 300 mg/L. To our best knowledge, this result reports the highest production yield of the recombinant IBP. More importantly, the rLeIBP secreted into culture media was stable and active for six days of fermentation. The thermal hysteresis (TH) activity of rLeIBP was about 0.42°C, which is almost the same to those reported previously. The ava
URI
http://repository.kopri.re.kr/handle/201206/6137
DOI
http://dx.doi.org/10.1007/s00253-012-4594-y
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