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Soluble and Cross-Linked Aggregated Forms of α-Galactosidase from Vigna mungo Immobilized on Magnetic Nanocomposites: Improved Stability and Reusability

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Abstract

α-Galactosidases hold immense potential due to their biotechnological applications in various industrial and functional food sectors. In the present study, soluble and covalently cross-linked aggregated forms of a low molecular weight, thermo-labile α-galactosidase from Vigna mungo (VM-αGal) seeds were immobilized onto chitosan-coated magnetic nanoparticles for improved stability and repeated usage by magnetic separation. Parameters like precipitants (type, amount, and ratio), glutaraldehyde concentration, and enzyme load were optimized for the preparation of chitosan-coated magnetic nanocomposites of cross-linked VM-αGal (VM-αGal-MC) and VM-αGal (VM-αGal-M) resulted in 100% immobilization efficiency. Size and morphology of VM-αGal-M were studied through dynamic light scattering (DLS) and scanning electron microscopy (SEM), while Fourier transform infrared spectroscopy (FTIR) was used to study the chemical composition of VM-αGal-MC and VM-αGal-M. VM-αGal-MC and VM-αGal-M were found more active in a broad range of pH (3–8) and displayed optimal temperatures up to 25 °C higher than VM-αGal. Addition of non-ionic detergents (except Tween-40) improved VM-αGal-MC activity by up to 44% but negatively affected VM-αGal-M activity. Both VM-αGal-MC (15% residual activity after 21 min at 85 °C, Ed 92.42 kcal/mol) and VM-αGal-M (69.0% residual activity after 10 min at 75 °C, Ed 39.87 kcal/mol) showed remarkable thermal stability and repeatedly hydrolyzed the substrate for 10 cycles.

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Funding

We thank Director, CSIR-CFTRI, for constant encouragement and support. The work was carried out under the CSIR networking project entitled Nano-materials: Applications and Impact on Safety, Health and Environment (NanoSHE, Project No. BSC0112) under the 12th five year plan, Govt of India, New Delhi. The grant of Junior Research Fellowship (JRF) to DP and PRM from CSIR and UGC, Govt. of India, New Delhi, respectively is gratefully acknowledged. JEJ thank CSIR, Ministry of Science and Technology, Govt. of India, for providing the project fellowship under NanoSHE, Project No. BSC0112.

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  1. Juby Elsa Joseph and Priyanka Rose Mary contributed equally to this work.

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  1. Department of Protein Chemistry and Technology, CSIR-Central Food Technological Research Institute, Mysuru, 570 020, India

    Juby Elsa Joseph, Priyanka Rose Mary, K. V. Haritha, Deepesh Panwar & Mukesh Kapoor

  2. Academy of Scientific and Innovative Research (AcSIR), CSIR-Human Resource Development Centre (CSIR-HRDC) Campus, Ghaziabad, UP, 201 002, India

    Priyanka Rose Mary, Deepesh Panwar & Mukesh Kapoor

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Highlights

• Synthesis of chitosan magnetic nanocomposites of soluble and cross-linked VM-αGal.

• VM-αGal-MC and VM-αGal-M were more active and stable than VM-αGal between pH 3 and 8.

• VM-αGal-MC and VM-αGal-M showed up to 25 °C higher temperature optima than VM-αGal.

• VM-αGal-MC and VM-αGal-M showed thermal stability up to 85 °C.

• VM-αGal-MC and VM-αGal-M repeatedly hydrolyzed the substrate for 10 cycles.

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Joseph, J.E., Mary, P.R., Haritha, K.V. et al. Soluble and Cross-Linked Aggregated Forms of α-Galactosidase from Vigna mungo Immobilized on Magnetic Nanocomposites: Improved Stability and Reusability. Appl Biochem Biotechnol 193, 238–256 (2021). https://doi.org/10.1007/s12010-020-03408-5

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