Abstract: Pyruvate oxidase, having multi subunits, plays an important role in food and fermentation industry. In recent years, genetic technology help to greatly increase the yield of pyruvate oxidase. However, the stability of pyruvate oxidase needs to be further improved. In this study, pyruvate oxidase was encapsulated by sol-gel technology. Firstly, single factor experiments were carried out to obtain the best value of each factor. Then an orthogonal experiment was used to optimize the conditions for pyruvate oxidase encapsulation by sol-gel technology. In this study, the optimized condition is: shaking in 5.0 mmol/L dioctyl sulfosuccinate sodium solution for 10 min; adding 0.672 mmol/L tetramethoxysilane for 5 h adsorption and then 24 h for encapsulation. The results showed that the final encapsulated pyruvate oxidase residual activity increased by 14% compared with control sample. And two to ten times increased in thermal stability of the encapsulated pyruvate oxidase than control sample at 20-30 ℃. These results provided a reliable methodology for pyruvate oxidase immobilization and application.
IKEBUKURO K, WAKAMURA H, KARUBE I, et al. Phosphate sensing system using pyruvate oxidase and chemiluminescence detection [J]. Biosensors & Bioelectronics, 1996, 11(10): 959-965.
NAKAMURA H, TANAKA H, HASEGAWA M, et al. An automatic flow-injection analysis system for determining phosphate ion in river water using pyruvate oxidase G (from Aerococcus viridans) [J]. Talanta, 1999, 50(4): 799-807.
KEHR S, SCHUBERT F. Pyruvate oxidase based mono- and multienzyme sensors for pyruvate determination and for ADP determination using analyte recycling [C]. The Third World Congress on Biosensors Abstracts, 1994, 164.
MIZUTANI F, YABUKI S, SATO Y, et al. Amperometric determination of pyruvate, phosphate and urea using enzyme electrodes based on pyruvate oxidase-containing poly(vinyl alcohol)/polyion complex-bilayer membrane [J]. Electrochimica Acta, 2000, 45(18): 2 945-2 952.
SPELLERBERG B, CUNDELL D R, SANDROS J, et al. Pyruvate oxidase, as a determinant of virulence in Streptococcus pneumoniae [J]. Molecular Microbiology, 2010, 19(4): 803-813.
MISAKI H,MATSUURA K,HARADA S. Process for the manufacture of pyruvate oxidase, and analytical method and kit for the use of the same:U.S.Patent 4,246,234[P].1981-1-20.
ZHAO J, WANG Y, CHU J, et al. Statistical optimization of medium for the production of pyruvate oxidase by the recombinant Escherichia coli[J]. Journal of Industrial Microbiology & Biotechnology, 2008, 35(4): 257-262.
LIANG J G, ZHAO J, WANG Z J, et al. Temperature gradient-based high-cell density fed-batch fermentation for the production of pyruvate oxidase by recombinant E.coli[J]. Preparative Biochemistry & Biotechnology, 2018, 48(2): 188-193.
LU J, ZHAO Y, ZHANG J. High-level expression of Aerococcus viridans pyruvate oxidase in Escherichia coli by optimization of vectors and induction conditions [J]. Letters in Applied Microbiology, 2018, 67(3): 262-269.
ZHANG J, LU J, SU E. Soluble recombinant pyruvate oxidase production in Escherichia coli can be enhanced and inclusion bodies minimized by avoiding pH stress [J]. Journal of Chemical Technology & Biotechnology, 2019, 94: 2 661-2 670.
PIERRE A C. The sol-gel encapsulation of enzymes [J]. Biocatalysis and Biotransformation, 2004, 22(3): 145-170.
WANG G, ZHANG L. A biofriendly sol–gel route to new hybrid gels for enzyme encapsulation [J]. Journal of Sol-Gel Science and Technology, 2014, 72(1): 85-91.
ZAHARESCU M, PREDOANA L, PANDELE J. Relevance of thermal analysis for sol-gel-derived nanomaterials [J]. Journal of Sol-Gel Science and Technology, 2018, 86(1): 7-23.
HONG S G, KIM B C, NA H B, et al. Single enzyme nanoparticles armored by a thin silicate network: Single enzyme caged nanoparticles [J]. Chemical Engineering Journal, 2017, 322: 510-515.
DU Y, GAO J, ZHOU L, et al. Enzyme nanocapsules armored by metal-organic frameworks: A novel approach for preparing nanobiocatalyst [J]. Chemical Engineering Journal, 2017, 327:1 192-1 197.