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. 2025 Dec 9;48(1):11.
doi: 10.1007/s10529-025-03680-7.

Enhancement of phytase production in recombinant Escherichia coli by medium components modification and fed-batch cultivation process optimization

Il Kim  1 , Yong Won Ho  2 , Ju Song Kim  2 , Jung Hyok Jon  2
Affiliations

Enhancement of phytase production in recombinant Escherichia coli by medium components modification and fed-batch cultivation process optimization

Il Kim et al. Biotechnol Lett. .

Abstract

This study focused on phytase overproduction in recombinant E. coli BL21(DE3)/pET-appA expressing E. coli phytase(APPA). A modified mineral salt medium was investigated for the strain growth and phytase production in shake-flask, which included 30 mM NH4OH and 17 g L-1 KH2PO4 as sole nitrogen(N) and phosphorous(P) sources, respectively. After that, fed-batch cultivation process, especially induction strategy feeding lactose intermittently was optimized in 30L bioreactor by Taguchi orthogonal experiments (L18(21 × 36)) at an induction cell density of OD600 25, followed by research on effect of induction cell densities. The optimized fed-batch cultivation process parameters included glucose starvation time of 15 min just before induction, induction cell density of OD600 45, induction temperature of 25 °C, predetermined specific growth rate of 0.04 h-1, DO level of 15%, lactose concentration of 60 mM, lactose feed times of 6 times, and pH of 7.2-7.6. In the optimized condition, the highest soluble phytase activity of 2 152U mL-1 ever reported was obtained at 33.0 h, indicating the enhancements of 31.6-fold and 3.7-fold compared to batch cultivation process in shake flask and pre-optimized fed-batch cultivation process in the bioreactor, respectively. We propose that the defined medium, the induction strategy and fed-batch process parameters could be applied to overproduce soluble proteins in E. coli.

Keywords: E. coli BL21(DE3); AppA; Fed-batch cultivation; Mineral salt medium; Phytase.

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Conflict of interest statement

Declarations. Conflict of interest: The authors declare no financial or commercial conflict of interest.

References

    1. Berkmen M, Boyd D, Beckwith J (2005) The nonconsecutive disulfide bond of Escherichia coli Phytase (AppA) renders it dependent on the protein-disulfide isomerase, DsbC*. J Biol Chem 280(12):11387–11394 - DOI - PubMed
    1. Chen CC, Wu PH, Huang CT, Cheng KJ (2004) A Pichia pastoris fermentation strategy for enhancing the hetetologous expression of an Escherichia coli phytase. Enzyme Microb Technol 35:15–320. https://doi.org/10.1016/j.enzmictec.2004.05.007 - DOI
    1. Cheng LC, Hor LI, Wu JY, Chen TL (2003) Effect of specific growth rate on the production of a recombinant nuclease by Escherichia coli. Biochem Eng J 14:101–107 - DOI
    1. Farliahati MR, Ramanan RN, Mohamad R, Puspaningsih NNT et al (2010) Enhanced production of xylanase by recombinant Escherichia coli DH5α through optimization of medium composition using response surface methodology. Ann Microbiol 60:279–285. https://doi.org/10.1007/s13213-010-0038-z - DOI
    1. Fruchtl M, Sakon J, Beitle R (2016) Alternate carbohydrate and nontraditional inducer leads to increased productivity of a collagen binding domain fusion protein via fed-batch fermentation. J Biotechnol 226:65–73. https://doi.org/10.1016/j.jbiotec.2016.03.016 - DOI - PubMed

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