Optimization of fermentation conditions of recombinant Pichia pastoris that can overexpress β-fructofuranosidase


Xihao Zhao;Qin Xiulin;Jia-Xun Feng;


Background(3-fructofuranosidase is an important enzyme that can be hydrolyzed and transferred fructosyl residues,generating fructooligosaccharides(FOS).The use of enzyme hydrolysis and fermentation of sugar transfer activity,large-scale production of FOS.Methods A process for efficient production of the p-fructofuranosidase from Penicillium oxalicum GXU20(PoFF32A)was established by secretory expression using Pichia pastoris.From Penicillium oxalicum cloning PoFF32A,were ligated into the plasmid pPIC9K to construct the recombintant plasmid p9K-BF.Then the plasmid p9K-BF was linearized with Sac I for the transformation into Pichia pastoris GS115.In order to improveβ-fructofuranosidase activity,four factors,including liquid volume in flask,induction temperature,amount of methanol,methanol and the amount of sorbitol,are supposed to have an impact on it.Results Signal-factor-at-a-time optimization was shown as follows:liquid volume in flask,20mL,the activity ofβ-fructofuranosidase up to 27.36 U/mL,induction temperature,28℃,the activity of pfructofuranosidase up to 3.14 U/mL,methanol,0.6%,the activity ofβ-fructofuranosidase up to 4.3U/mL,methanol and sorbitol,0.8%and 1.5%,the activity of p-fructofuranosidase up to 9.49 U/mL.Then the conditions for the activity ofβ-fructofuranosidase will be optimized by RSM.Conclusion The results indicate that the recombinant strain G/9K-BF can overexpressβ-fructofuranosidase under the conditions of liquid volume in flask,20mL,induction temperature,28℃,methanol,0.6%,methanol and sorbitol,0.8%and 1.5%.


β-fructofuranosidase;;Penicillium oxalicum;;Pichia pastoris


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