Enhancement of thermoalkaliphilic xylanase production by Pichia pastoris through novel fed-batch strategy in high cell-density fermentation

BMC Biotechnology

Research Article

Tingting Shang¹  Dongyan Zhang¹  Dayong Si¹  Xuhui Liu¹  Yu Fu¹  Cong Hu¹  Rijun Zhang¹  Longmei Zhao² 

[1] Laboratory of Feed Biotechnology, State Key Laboratory of Animal Nutrition, College of Animal Science & Technology, China Agricultural University, No.2 Yuanmingyuan West Road, 100193, Beijing, Haidian District, China;Laboratory of Feed Biotechnology, State Key Laboratory of Animal Nutrition, College of Animal Science & Technology, China Agricultural University, No.2 Yuanmingyuan West Road, 100193, Beijing, Haidian District, China;College of Animal Science and Technology, Henan University of Science and Technology, 471003, Luoyang, China;

关键词: Thermoalkaliphilic xylanase;    Pichia pastoris;    Fermentation parameter optimization;    Bioprocess;    Fed-batch strategy;    High cell-density fermentation;   

DOI  :  10.1186/s12896-017-0361-6

 received in 2017-01-16, accepted in 2017-04-20,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundXylanase degrades xylan into monomers of various sizes by catalyzing the endohydrolysis of the 1,4-β-D-xylosidic linkage randomly, possessing potential in wide industrial applications. Most of xylanases are susceptible to be inactive when suffering high temperature and high alkaline process. Therefore, it is necessary to develop a high amount of effective thermoalkaliphilic xylanases. This study aims to enhance thermoalkaliphilic xylanase production in Pichia pastoris through fermentation parameters optimization and novel efficient fed-batch strategy in high cell-density fermentation.ResultsRecombinant xylanase activity increased 12.2%, 7.4%, 12.0% and 9.9% by supplementing the Pichia pastoris culture with 20 g/L wheat bran, 5 mg/L L-histidine, 10 mg/L L-tryptophan and 10 mg/L L-methionine in shake flasks, respectively. Investigation of nutritional fermentation parameters, non-nutritional fermentation parameters and feeding strategies in 1 L bioreactor and 1 L shake flask revealed that glycerol and methanol feeding strategies were the critical factors for high cell density and xylanase activity. In 50 L bioreactor, a novel glycerol feeding strategy and a four-stage methanol feeding strategy with a stepwise increase in feeding rate were developed to enhance recombinant xylanase production. In the initial 72 h of methanol induction, the linear dependence of xylanase activity on methanol intake was observed (R2 = 0.9726). The maximum xylanase activity was predicted to be 591.2 U/mL, while the actual maximum xylanase activity was 560.7 U/mL, which was 7.05 times of that in shake flask. Recombinant xylanase retained 82.5% of its initial activity after pre-incubation at 80 °C for 50 min (pH 8.0), and it exhibited excellent stability in the broad temperature (60–80 °C) and pH (pH 8.0–11.0) ranges.ConclusionsEfficient glycerol and methanol fed-batch strategies resulting in desired cell density and xylanase activity should be applied in other P. pastoris fermentation for other recombinant proteins production. Recombinant xylanases with high pH- and thermal-stability showed potential in various industrial applications.

【 授权许可】

CC BY   
© The Author(s). 2017