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Additional stabilization of stem bromelain coupled to a thermosensitive polymer by uniform orientation and using polyclonal antibodies

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Abstract

Stem bromelain was covalently coupled to a thermosensitive polymer of N-isopropylacrylamide (p(NIPAm)) either through the amino groups of the enzyme (randomly coupled) or via the lone oligosaccharide chain (uniformly coupled). The enzyme coupled via the oligosaccharide chain exhibited better access to the substrate casein as compared to the preparation in which the amino groups formed the point of contact between the enzyme and the polymer. Native bromelain exhibited a pH optimum of 8.0 and a broad pH-activity profile. The polymer-coupled preparations exhibited broader pH-activity profiles and shifting of pH optimum to 10.0 at 35°C. At 25°C, the shifting of pH optimum was observed for the randomly coupled enzyme only. The temperature-activity profiles of bromelain coupled to p(NIPAm) also showed appreciable broadening and the preparations retained greater fraction of maximum activity above the temperature optimum. The optimum temperature of the uniformly oriented preparation also rose to 70°C. Inactivation rates of the polymer-coupled bromelain were remarkably low at 60°C as compared to the native protease, and binding of antibromelain antibodies improved the resistance to inactivation of the polymer-coupled preparations. The cleavage patterns of hemoglobin and IgG by the native bromelain and the polymer-coupled preparations were comparable.

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Abbreviations

LCST:

low critical solution temperature

p(NIPAm)):

poly(N-isopropylacrylamide)

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Authors and Affiliations

  1. Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, 202002, India

    Rubab Mahmood & M. Saleemuddin

  2. Department of Biochemistry, Aligarh Muslim University, Aligarh, 202002, India

    M. Saleemuddin

Authors
  1. Rubab Mahmood
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  2. M. Saleemuddin
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Correspondence to M. Saleemuddin.

Additional information

Published in Russian in Biokhimiya, 2007, Vol. 72, No. 3, pp. 375–382.

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Mahmood, R., Saleemuddin, M. Additional stabilization of stem bromelain coupled to a thermosensitive polymer by uniform orientation and using polyclonal antibodies. Biochemistry Moscow 72, 307–312 (2007). https://doi.org/10.1134/S000629790703008X

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  • DOI: https://doi.org/10.1134/S000629790703008X

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