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KMID : 1094720150200020315
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Biotechnology and Bioprocess Engineering
2015 Volume.20 No. 2 p.315 ~ p.323
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Enhancement of thermostable β-glucosidase production in a slow methanol utilization strain of Pichia pastoris by optimization of the specific methanol supply rate
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Charoenrat Theppanya
Sangprapai Kanyalak
Promdonkoy Peerada
Kocharin Kanokarn
Tanapongpipat Sutipa
Roongsawang Niran
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Abstract
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Control of the methanol supply during the production stage is a crucial parameter for maintaining cell growth and enhancing recombinant protein production by Pichia pastoris. In this study, optimization of the initial specific methanol supply rate was explored for high cell density fed-batch cultivation of recombinant fungal β- glucosidase in a slow methanol utilization strain (MutS), P. pastoris KM71. By varying the methanol feed rates for initiating the induction at a cell concentration of 60 g/L, the optimum initial specific methanol supply rate was determined to be 30.34 ± 0.34 mg/g·h. A methanol feed rate was proposed according to this optimum parameter and applied for the production of recombinant β-glucosidase at the higher cell concentrations of 80 and 100 g/L. The highest recombinant β-glucosidase activity obtained was 2851.7 ± 14.6 U/mL, which was four times higher than that obtained with the reference condition (40 g/L initial cell concentration). The success of this approach suggests that the strategy of optimizing the initial specific methanol supply rate could be adopted and applied for the production of other recombinant proteins in P. pastoris employing a methanol inducible system.
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KEYWORD
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β-glucosidase, Pichia pastoris, methanol feed rate, specific methanol supply rate, specific methanol consumption rate
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