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Recovery and Purification of Oligosaccharides from Copra Meal by Recombinant Endo-β-mannanase and Deciphering Molecular Mechanism Involved and Its Role as Potent Therapeutic Agent

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Abstract

Production of manno-oligosaccharides (MOSs) from pretreated and defatted copra meal (dFCO) hydrolysis was achieved by endo-mannanase. Structural characterization of dFCO by FT-IR and NMR exhibited resemblance with galactomannan. The time-dependent hydrolysis of dFCO by recombinant endo-β-(1 → 4)-mannanase of Clostridium thermocellum by TLC and HPAEC displayed the release of mannose and MOSs mannobiose and mannotriose. Purified MOSs yielded 40 % mannobiose and 18 % mannotriose confirmed by mass spectroscopy which showed mannobiose (m/z = 365) and mannotriose (m/z = 527). The homology based structural analysis of catalytic endo-mannanase (CtManT) showed the catalytic core composed of Glu181 and Glu300 acting as acid/base and Glu288 as a nucleophile during galactomannan hydrolysis. Sub-site map** of CtManT exhibited two aglycone and four glycone sites at cleavage sites existing on either side of β-(1 → 4)-linkage of galactomannan. Isolated MOSs displayed potential prebiotic characteristics and supported higher growth of probiotic Lactobacillus acidophilus and Bifidobacterium infantis than with standard inulin. Moreover, MOSs displayed over 97 % tolerance to simulated gastric juice, intestinal fluid, and α-amylase proving its potential as a stable prebiotic over inulin. In vitro cytotoxicity assay of MOSs (500 µg/mL) on human epithelial colorectal adenocarcinoma cell line (HT-29) demonstrated 60 % decreased viability of cells after 48 h displaying anti-tumorigenic property.

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Acknowledgments

Arabinda Ghosh is supported by a scholarship from University Grants Commission (UGC), New Delhi, India. The mass spectrometric analysis performed in the Department of Chemistry, IIT Guwahati is gratefully acknowledged. The funder has no role in designing the experiments, preparation of this manuscript and bearing any further publication charges. Moreover, authors declare no conflict of interest.

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  1. Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, 781 039, Assam, India

    Arabinda Ghosh, Anil Kumar Verma, Jaganmohan Rao Tingirikari, Rishikesh Shukla & Arun Goyal

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  1. Arabinda Ghosh
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Correspondence to Arun Goyal.

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Arabinda Ghosh and Anil Kumar Verma contributed equally.

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Ghosh, A., Verma, A.K., Tingirikari, J.R. et al. Recovery and Purification of Oligosaccharides from Copra Meal by Recombinant Endo-β-mannanase and Deciphering Molecular Mechanism Involved and Its Role as Potent Therapeutic Agent. Mol Biotechnol 57, 111–127 (2015). https://doi.org/10.1007/s12033-014-9807-4

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