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Biochemical properties and application of a novel pectinase from a mutant strain of Bacillus subtilis

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Abstract

Pectinases have broad applications in food industries attributed to their role as a biocatalyst in degrading pectic compounds. The present work reports the purification and characterization of a pectinase from a mutant strain of Bacillus subtilis. A three-step enzyme purification involving ammonium sulfate precipitation, followed by ion-exchange chromatography (IEC) and size-exclusion chromatography (SEC), was evaluated to recover the enzyme effectively. The pectinase was purified up to 7.98-fold with 65.63% total recovery, resulting in maximum specific activity of 362.04 U/mg after three purification steps. The eluted fractions of IEC and SEC showed pectinase activities of 9520 U and 8288 U, respectively. The molecular weight of this extracellular pectinase was found to be 30 kDa after sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The optimal conditions of temperature and pH for maximum pectinase activity were 37 °C and 4–5, respectively. The enzyme also exhibited high stability when incubated at optimal conditions for 60 min. The addition of calcium and magnesium ions elicited pectinase activity at 4 mM concentrations, while copper and zinc ions inhibited the pectinase activity. The Vmax and Km values were 362 U and 0.62 mg/ml, respectively, for the pectin substrate obtained from citrus peel. Further analysis on the efficiency of clarification in sweet lime, pineapple, and lime juice showed 79.25, 72.76, and 61.24% reductions in turbidity within 1-h incubation. Therefore, characteristics of pectinase suggest that it can be potentially used for industrial applications as a biocatalyst.

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Acknowledgements

Ram Balak Mahto and Mukesh Yadav express gratitude to HoD, Department of Biotechnology, Maharishi Markandeshwar (Deemed to be University), Mullana 133207, Ambala, India, for their encouragement and support. Dr. Biswanath Bhunia and Muthusivaramapandian Muthuraj express gratitude to the Director, National Institute of Technology Agartala for the constant inspiration and funding.

Funding

Dr. Biswanath Bhunia would like to express their gratitude to the Department of Science and Technology (DST), Government of India, for providing financial assistance to Fast Track Young Scientist (SERC/LS-167/2012).

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Authors and Affiliations

  1. Department of Biotechnology, Maharishi Markandeshwar (Deemed To Be University), Mullana 133207, Ambala, India

    Ram Balak Mahto, Mukesh Yadav & Anil K. Sharma

  2. Department of Bio Engineering, National Institute of Technology Agartala, Agartala, 799046, India

    Muthusivaramapandian Muthuraj & Biswanath Bhunia

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  1. Ram Balak Mahto
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Mahto, R.B., Yadav, M., Muthuraj, M. et al. Biochemical properties and application of a novel pectinase from a mutant strain of Bacillus subtilis. Biomass Conv. Bioref. 13, 10463–10474 (2023). https://doi.org/10.1007/s13399-021-02225-y

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