Purification, characterization, kinetics and thermodynamic analysis of polygalacturonase from Aspergillus tamarii for industrial applications
Increasing industrial demand has urged the scientists to biosynthesize polygalacturonase (PG) that would be operative, stable, cost effective and can be used in industrial applications particularly in fruit juice clarification and poultry feed. Polygalacturonase purification and characterization from A. tamarii was the main focus of the current research work. Fractionation by ammonium sulphate along with ion exchange chromatography was used for purification and 2.27 purification fold with 51.25 percentage yield was attained. SDS-PAGE analysis showed the molecular mass of purified enzyme as 70kDa. Enzyme kinetic assessment i.e. Km 2.85mg/mL and Vmax 55.55 in addition with thermodynamic determinants such as Ea=-39.84 KJ/mol, ΔH=37.62 KJ/mol and ΔS=-38.05 KJ/mol proves industrial stability of this enzyme. Further, different parameters were characterized which revealed that enzyme remained stable up to 50℃ and pH 8. Furthermore, PG found to retain its activity in the presence of different metals whereas some inhibitors reduced its activity i.e. PMSF and EDTA. The enzyme was found to have a better shelf life of 30 days at -20 and 4oC compared to room temperature. The reduction in turbidity of fresh juice and increase in body weight of chicks feeding on feed treated with purified PG proves its efficiency in industrial applications.
Amid, M., Manap, Y. and Zohdi, K. (2014). Purifcation and characterisation of thermo-alkaline pectinase enzyme from hylocereus polyrhizus. European Food Research and Technology 239, 21–29. (https://doi.org/10.1007/s00217-014-2188-x).
Anand, G., Yadav, S. and Yadav, D. (2016). Purification and characterization of polygalacturonase from Aspergillus fumigatus MTCC 2584 and elucidating its application in retting of Crotalaria juncea fiber. 3 Biotech 6, 201. (https://doi.org/10.1007/s13205-016-0517-4).
Azar, R.I.L., Morales, D.L.M., Alfenas, M.G.P., Falkoski, D.L., Alfenas, R.F. and Guimarães, V.M. (2020). Apple juice clarification by a purified polygalacturonase from Calonectria pteridis. Food and Bioproducts Processing 119, 238-245. (https://doi.org/10.1016/j.fbp.2019.11.013).
Chaudhri, A. and Suneetha, V. (2012). Microbially derived pectinases: A review. Journal of Pharmacy and Biological Sciences 2, 1-5. (https://doi.org/10.9790/3008-0220105)
Dalagnol, L.M., Silveira, V.C., da Silva, H.B., Manfroi, V. and Rodrigues, R.C. (2017). Improvement of pectinase, xylanase and cellulase activities by ultrasound: effects on enzymes and substrates, kinetics and thermodynamic parameters. Process Biochemistry 61, 80-87. (https://doi.org/10.1016/j.procbio.2017.06.029)
Daniel, I.U., Chukwunonso, A.N., Arinze, L.E., Sabinus, O.O., Chukwudi, A., Peter, N. U. and Ferdinand, C.C. (2014). Extraction, partial purification and characterization of pectinases isolated from Aspergillus species cultured on mango (Mangifera indica) peels. African Journal of Biotechnology 13, 2445-2454.
Deng, Z., Wang, F., Zhou, B., Li, J., Li, B. and Liang, H. (2019). Immobilization of pectinases into calcium alginate microspheres for fruit juice application. Food hydrocolloids 89, 691-699. (https://doi.org/10.1016/j.foodhyd.2018.11.031).
Deshmukh, N., Talkal, R., Jha, K., Singh, P.G. and Prajapati, D.C. (2012). Production, Purification, Characterization and Comparison of Polygalacturonase from various strains of Aspergillus. International Journal of Scientific and Technology Resource 1, 85-91.
Fratebianchi, D., Cavello, I.A. and Cavalitto, S.F. (2017). Purification and biochemical and kinetic properties of an endo-polygalacturonase from the industrial fungus Aspergillus Sojae. Journal of Molecular Microbiology and Biotechnology 27, 102-109. (https://doi.org/10.1159/000460296).
Giacobbe, S., Pepe, O., Ventorino, V., Birolo, L., Viciguerra, R. and Faraco, V. (2014). Identification and characterization of a pectinolytic enzyme form Paenibacillus xylanolyticus. Bioresources 9, 4873-4887.
Jagadeesh B.P.E. and Viswanathan, R. (2010). Studies on the effect of pH, temperature and metal ions on the production of pectinase from tamarind kernel powder by submerged fermentation using Aspergillus foetidus (NCIM 505). Asian-Pacific Journal of Chemical Engineering 5, 396–400. (https://doi.org/10.1002/apj.296)
Khatri, B.P., Bhattarai, T., Shrestha, S. and Maharjan, J. (2015). Alkaline thermostable pectinase enzyme from Aspergillus niger strain MCAS2 isolated from Manaslu Conservation Area, Gorkha, Nepal. SpringerPlus 4, 488. (https://doi.org/10.1186/s40064-015-1286-y)
Kutlu, H.R., Saber, S.N., Kutay, H., Celik, L., Yusuf, U.Z.U.N., Nurten, T.O.Y. and Yavuz, B. (2019). Effect of Multi-enzyme Produced By a Single Fungus on Growth Performance and Some Carcass Parameters of Broiler Chicks Fed on Maize-Soya Based Diets. Kafkas Üniversitesi Veteriner Fakültesi Dergisi 25, 221-230. (DOI: 10.9775/kvfd.2018.20765).
Li, S., Yang, X., Yang, S., Zhu, M. and Wang, X. (2012). Technology prospecting on enzymes: application, marketing and engineering. Computational and Structural Biotechnology Journal 2, 1-11. (https://doi.org/10.5936/csbj.201209017).
Ma, Y., Sun, S., Hao, H. and Xu, C. (2016). Production, purification and characterization of an exopolygalacturonase from Penicillium janthinellum sw09. Proceedings of the Brazilian Acdemy of Sciences 88, 479-487. (http://dx.doi.org/10.1590/0001-3765201620150051)
Marquez, W.V.E., Martinez-Trujillo, M.A., Osorio, G.A., Patiño, F., Alvarez, M.S., Rodríguez, A., Sanromán, M.Á. and Deive, F.J. (2017). Scaling-up and ionic liquid-based extraction of pectinases from Aspergillus flavipes cultures. Bioresource technology 225, 326-335. (https://doi.org/10.1016/j.biortech.2016.11.067).
Martin, M.C., López, O.V., Ciolino, A.E., Morata, V.I., Villar, M.A. and Ninago, M.D. (2019). Immobilization of enological pectinase in calcium alginate hydrogels: A potential biocatalyst for winemaking. Biocatalysis and Agricultural Biotechnology 18, 101091. (https://doi.org/10.1016/j.bcab.2019.101091).
Martos, M.A., Butiuk, A.P., Rojas, N.L. and Hours, R.A. (2014). Purification and Characterization of a Polygalacturonase Produced by Wickerhamomyces anomalus. Brazilian Archives of Biology and Technology 57, 587-594. (http://dx.doi.org/10.1590/S1516-8913201402214)
Munir, M., Abdullah, R., Haq, I.U., Kaleem, A. and Iqtedar, M. (2019). Isolation and identification of multi stress tolerant polygalacturonase producing fungi from various fruits. Journal of Animal and Plant Sciences, 29(3), 825-832.
Obafemi, Y.D., Ajayi, A.A., Taiwo, O.S., Olorunsola, S.J. and Isibor, P.O. (2019). Isolation of Polygalacturonase-Producing Bacterial Strain from Tomatoes (Lycopersicon esculentum Mill.). International Journal of Microbiology 7, l-9. (https://doi.org/10.1155/2019/7505606)
Okafor, U.A., Okochi, V.I., Chinedu, S.N., Ebuehi, O.A.T. and Okerenta, B.M.O. (2010). Pectinolytic activity of wild-type filamentous fungi fermented on agro-wastes. African Journal of Microbiology Research 4, 2729-2734.
Oumer, O.J. (2017). Pectinase: substrate, production and their biotechnological applications. International Journal of Environment, Agriculture and Biotechnology 2, 10071014. (http://dx.doi.org/10.22161/ijeab/2.3.1)
Sandhya, R. and Kurup, G. (2013). Screening and isolation of pectinase from fruit and vegetable wastes and the use of orange waste as a substrate for pectinase production. International Research Journal of Biological Sciences 2, 34-39.
Saxena, R.K., Sheoran, A., Giri, B. and Davidson, W.S. (2003). Purification strategies for microbial lipases. Journal of Microbiology Methods 52, 1-18. (https://doi.org/10.1016/S0167-7012(02)00161-6)
Sethi, B.K., Nanda, P.K. and Sahoo, S. (2016). Enhanced production of pectinase by Aspergillus terreus NCFT 4269.10 using banana peels as substrate. 3 Biotech 6, 36. (https://doi.org/10.1007/s13205-015-0353-y).
Siddiqui, M., Pande, V. and Arif, M. (2012). Production, purification, and characterization of polygalacturonase from Rhizomucor pusillus isolated from decomposting orange peels. Enzyme Research 2012, 1-8. (https://doi.org/10.1155/2012/138634).
Suneetha, V. and Prathyusha, K. (2011). Bacterial pectinases and their potent biotechnological application in fruit processing/juice production industry: a review. The Journal of Phytological Research 3, 11-15.
Thakur, A., Pahwa, R., Singh, S. and Gupta, R. (2010). Production, purification, and characterization of polygalacturonase from Mucor circinelloides ITCC 6025. Enzyme Research 2010, 21-27. (https://doi.org/10.4061/2010/170549)
Trindade, L.V., Desagiacomo, C., Polizeli, M.D.L.T.D., Damasio, A.R.D.L., Lima, A.M. F., Gomes, E. and Bonilla-Rodriguez, G.O. (2016). Biochemical characterization, thermal stability, and partial sequence of a novel exo-polygalacturonase from the thermophilic fungus Rhizomucor pusillus A13. 36 obtained by submerged cultivation. BioMed Research International 2016, 110. (https://doi.org/10.1155/2016/8653583)
Udenwobele, D.I., Nsude, C.A., Ezugwu, A.L., Eze, S.O., Anyawu, C., Uzoegwu, P.Z. and Chilaka, F.C. (2014). Extraction, partial purification and characterization of pectinases isolated from Aspergillus species cultured on mango (Mangifera indica) peels. African Journal of Biotechnology 13, 2445-2454. (DOI: 10.5897/AJB2014.13771).
Copyright (c) 2020 Revista Mexicana de Ingeniería Química
This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
By publishing your paper in our journal you are also granting it the copyright of the information that it contains.