Kinetics and Thermodynamic Properties of Glucose Oxidase Obtained from Aspergillus fumigatus ASF4 doi.org/10.26538/tjnpr/v6i3.22
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Abstract
Heat instability is a major setback that prevents the broader use of glucose oxidase (GOx) in industries. This research explored the kinetic and thermodynamic parameters of Aspergillus fumigatus ASF4 GOx to determine its potential for biotechnological applications. Aspergillus fumigatus ASF4 GOx was purified 2.18-fold with a 6.25% yield after ammonium sulfate precipitation (60%), dialysis, ion-exchange chromatography, and gel filtration. The pH and temperature optima for GOx activity were 5.5 and 40°C, respectively. Metal ions, Ag2+ and Hg2+ had a remarkable inhibitory effect on GOx activity whereas Ca2+, Mg2+, and Mn2+ enhanced GOx activity. The maximum velocity (Vmax) and Michaelis constant (KM) were 2000 µmol/min, and 24 mM, respectively. The enzyme retained 85% and 90% of its initial activity at 40ᵒC and 30ᵒC, respectively after 120 min of incubation. At 50ᵒC and 45ᵒC, the enzyme retained more than 50% of its initial activity after 120 min of incubation. The k values at 37°C were the lowest (0.002) whereas that at 70°C was the highest (0.011). The Z-value was 0.3 and the activation energy (Ea) was 70.64 KJ/mol/K suggesting great sensitivity of GOx to temperature change. The D-value of Aspergillus fumigatus ASF4 GOx ranged between 115.5 to 208.4 min. The thermodynamic studies showed that glucose oxidation by Aspergillus fumigatus ASF4 GOx was reversible (ΔS<0), endothermic (ΔH>0), and non-spontaneous (ΔG>0) at all temperatures tested. The results on the optimum conditions for GOx activity and stability have shown that Aspergillus fumigatus ASF4 GOx can find application in the industrial production of gluconic acid.
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Lim HY and Dolzhenko AV. Gluconic acid aqueous solution: A bio-based catalytic medium for organic synthesis. Sustain Chem Pharm. 2021; 21(2021):1-15.
Leskovac V, Trivic S, Wohlfahrt G, Kandrac J, Pericin D. Glucose oxidase from Aspergillus niger: the mechanism of action with molecular oxygen, quinones, and one-electron acceptors. Int J Biochem. 2005; 37(45):731-750.
Bankar S, Bule M, Singhal R, Ananthanarayan L. Optimization of Aspergillus niger fermentation for the production of glucose oxidase. Food Bioproc Technol. 2008; 10(7):47-50.
Mu Q, Cui Y, Tian Y, Hu M, Tao Y, Wu B. Thermostability improvement of the glucose oxidase from Aspergillus niger for efficient gluconic acid production via computational design. Int J Biol Macromol. 2019; 136(2019):1060-1068.
Ashande CM, Masunda AT, Ngbolua K, Kilembe JT, Matondo A, Clément IL, Benjamin GZ, Emmanuel LM, Tshibangu DST, Tshilanda DD, Mpiana PT, Mudogo V. Glucose oxidase as a model enzyme for antidiabetic activity evaluation of medicinal plants: In vitro and in silico
evidence. Nat Resour Hum Health. 2022; 2(2):265-273.
Khatami SH, Vakili O, Ahmadi N, Fard ES, Mousavi P, Khalvati B, Maleksabet A, Savardashtaki A, TaheriAnganeh M, Movahedpour A. Glucose oxidase: Applications, sources, and recombinant production.Biotechnol Appl Biochem. 2021; 1(2021):1-12.
Pavankalyan U, Yasaswi A, Teja A, Kanuri G, Chanda C.Optimization and cost-effective production of fungal glucose oxidase using palm jaggery. Int J Sci Technol Res. 2020; 9(6):466-469.
Afjeh ME, Pourahmad R, Akbari-Adergani B, Azin M. Characteristics of glucose oxidase immobilized on magnetic chitosan nanoparticles. Food Sci Technol. 2020; 40(1):68-75.
Wang Y, Wang J, Leng F, Ma J, Bagadi A. Expression of Aspergillus niger glucose oxidase in Pichia pastoris and its antimicrobial activity against Agrobacterium and Escherichia coli. Peer J. 2020 4(8):1-17.
Bankar SB, Bule MV, Singhal RS, Ananthanarayan L. Optimization of A. niger fermentation for the production of GOx. Food Bioproc Tech. 2009; 2(4):344–352.
Sunoqrot S, Al-Hadid A, Manasrah A, Khnouf R, Ibrahim LH. Immobilization of glucose oxidase on bioinspired polyphenol coatings as a high-throughput glucose assay platform. RSC Adv. 2021; 11(2021):39582-39592.
Yuan M, Ning C, Yang S, Liang Q, Mou H, Liu Z. A new cold-active glucose oxidase from Penicillium: High-level expression and application in fish preservation. Front. Microbiol. 2020; 11(606007):1-14.
Tu T, Wang Y, Huang H, Wang Y, Jiang X, Wang Z, Yao B, Luo H. Improving the thermostability and catalytic efficiency of glucose oxidase from Aspergillus niger by molecular evolution. Food Chem. 2019; 281(2019):163-170.
Dubey MK, Zehra A, Aamir M, Meena M, Ahirwal L, Singh S, Shukla S, Upadhyay RS, Bueno-Mari R, Bajpai VK. Improvement strategies, cost effective production, and potential applications of fungal glucose oxidase (GOx): Current updates. Front Microbiol. 2017; 8(1032):10-32.
Xu G, Xu Y, Li A, Chen T, Liu J. Enzymatic bioactivity investigation of glucose oxidase modified with hydrophilic or hydrophobic polymers via in situ RAFT polymerization. J Polym Sci A Polym Chem. 2017; 55(8):1289-1293.
Dang DX, Hoque MR, Liu Y, Chen N, Kim IH. Dietary glucose oxidase supplementation improves growth performance, apparent nutrient digestibility, and serum antioxidant enzyme parameters in growing pigs. Ital J Anim Sci. 2021; 20(1):1568-1574.
Mao Y. Preparation of gluconic acid by oxidation of glucose with hydrogen peroxide. J Food Proc Preserv. 2016; 00(2016):1-5.
Ambarsari L, Maulana FA, Wahyudi ST, Kurniatin PA, Nurcholis W. Molecular dynamics analysis of glucose oxidase stability against temperature. Biointerf Res Appl Chem. 2022; 12(3):4062-4073.
Martin N, de Souza SR, da Silva R, Gomes E. Pectinase production of fungal strains in solid-state fermentation using agro-industrial bioproduct. Braz Arch Biol Technol. 2004; 47(5):813-819.
Park EH, Shin YM, Lim YY, Kwon TH, Kim DH, Yang MS. Expression of glucose oxidase by using recombinant yeast. J Biotechnol. 2000; 81(1):35-44.
Fiedurek J and Szezodrak J. Glucose Oxidase biosynthesis in relation to bio-chemical mutation in A. niger. Acta Biotechnol. 1995; 15(1):107-115.
Bergmeyer H, Gawehn K, Grassl M. Methods of Enzymatic Analysis. (2nd ed.). New York: Academic Press Inc; 1974; 457-458p.
Lowry OH, Rosebrough NJ, Farr AL, Randell RJ. Protein measurement with the folin pheniol reagent. J Biol Chem.1951; 193(1):265-275.
Chilaka F, Nwachukwu A, Uvere P. Thermal stability studies of β-galactosidase from germinating seeds of the brown beans, vignaunguiculata. Nig J Biochem Mol Biol. 2002; 17(1):51-56.
Singh J and Verma N. Glucose oxidase from Aspergillus niger: Production, characterization and immobilization for glucose oxidation. Adv Appl Sci Res. 2013; 4(2013):250-257.
Sandalli C, Saral A, Ulker S, Karaoglu H, Belduz AO, Cicek AC. Cloning, expression and characterization of a novel CTP synthase gene from Anoxybacillus gonensis G2. Turk J Biol. 2014; 37(1):111-117.
Yanmiş D, Karaoglu H, Colak ND, Sal FA, Canakci S, Belduz AO. Characterization of a novel xylose isomerase from Anoxybacillus gonensis G2T. Turk J Biol. 2014; 38(2014):586-592.
Simpson C, Jordaan J, Gardiner NS, Whiteley C. Isolation, purification and characterization of a novel glucose oxidase from Penicillium sp. CBS 120262 optimally active at neutral pH. Protein Expr Purif. 2007; 51(2):260-266.
Bai L, He L, Yu P, Luo J, Yang M, Xiangren A, Wei X. Molecular characterization of mycobiota and Aspergillus species from Eupolyphaga sinensis Walker Based on highthroughput sequencing of ITS1 and CaM. J Food Qual. 2020; 2020(1):1-7.
Zia MA, Ain Q, Iftikhar T, Abbas RZ, Khalil-ur-Rahman. Production of rabbit antibodies against purified glucose oxidase. Braz Arch Biol Technol. 2012; 55(1):69-74.
Jithendar T, Sairam K, Verma V. Purification, Characterization, Thermostability and Shelf Life Studies of Glucose Oxidase from Aspergillus niger PIL7. Res J Pharm Biol Chem Sci. 2015; 6(3):1666-1679.
Belyad F, Karkhanei AA, Raheb J. Expression, characterization and one step purification of heterologous glucose oxidase gene from Aspergillus niger ATCC 9029 in Pichia pastoris. Proteom. 2018; 19(2018):1-5.
Bhatti HN, Madeeha M, Asghr M, Batool N. Purification and thermodynamic characterization of glucose oxidase from a newly isolated strain of Aspergillus. Can JMicrobiol. 2006; 52(6):519-524.
Wilson L, Illanes A, Pessela BCC, Abian O, FernándezLafuente R, Guisán JM. Encapsulation of crosslinked penicillin G acylase aggregates in lentikats: Evaluation of a novel biocatalyst in organic media. Biotechnol. Bioeng. 2014; 86(5):558-562.
Sukhacheva MV, Davydova M.E, Netrusov AI. Production of Penicillium funiculosum 433 glucose oxidase and its properties. Appl Biochem Microbiol. 2004; 40(1):25–29.
Wu Y, Chu L, Liu W, Jiang L, Chen X, Wang Y, Zhao Y. The screening of metal ion inhibitors for glucose oxidase based on the peroxidase-like activity of nano-Fe3O4. RSC Adv. 2017; 7(75):47309-47315.
Okwuenu PC, Agbo KU, Ezugwu AL, Eze SO, Chilaka FC. Effect of divalent metal ions on glucoamylase activity of glucoamylase isolated from Aspergillus niger. Ferment Technol. 2017; 6(1):1-6.
Akor J, Ugwoke IF, Odu NM, Ogara AL, Ogbonna EK, Ogidigo OJ, Oluigbo CC, Aham CE, Parker E. Joshua PE, Eze SOO. Assessment of kinetic parameters of peroxidase isolated from maturing Solanum lycopersicum fruits for analytical and biotechnological applications. Trop J Nat
Prod Res. 2021; 5(12):2149-2153.
Ahmed SA, Saleh SAA, Abdel-Hameed SAM, Fayad AM. Catalytic, kinetic, and thermodynamic properties of free and immobilized caseinase on mica glass-ceramics. Heliyon. 2019; 5(5):1-13.
Omeje K, Eze SOO, Chilaka FC. Studies on the Energetics of Heat Inactivation of Peroxidase from Green Colored Solanum ethiopicum Fruit. J Thermodyn Cat. 2017; 8(3):1-3.
Zeyadi M and Almulaiky YQ. A novel peroxidase from Ziziphus jujuba fruit: purification, thermodynamics and biochemical characterization properties. Sci Rep. 2020; 10(8007):1-11.
Sant’Anna V, Cladera-Olivera F, Brandelli A. Kinetic and thermodynamic study of thermal inactivation of the antimicrobial peptide P34 in milk. Food Chem. 2012; 130(2012):84-89.
Gohel V and Naseby DC. Thermal-stabilization of chitinolytic enzymes of Pantoea dispersa. Biochem Eng J. 2007; 35(2007):150-157.
Wierzbicki AJ, Dala P, Cheatam PET, Knickelbein EJ, Haymet ADJ, Madura DJ. Antifreeze protein at the ice/water interphase: three calculated discrimination properties for orientation of type I proteins. Biophys J. 2007; 93(5):1442-1451.
Prajapati VA, Trivedi UB, Patel KC. Kinetic and thermodynamic characterization of glucoamylase from Colletotrichum sp. KCP1. Ind J Microbiol. 2014; 54(1):87–93.
Deylami MZ, Rahman RA, Tan CP, Bakar J, Olusegun L. Thermodynamics and kinetics of thermal inactivation of peroxidase from mangosteen (garcinia mangostana l.) pericarp. J Eng Sci Technol. 2014; 9(3):374-383.