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Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides

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Summary

Dextran (MW=7.2×104), carboxymethylcellulose (MW=2.5×104, substitution degree=0.7) and Ficoll (MW=6.9×104) were derivatized with 1,4-diaminobutane and covalently attached to bovine pancreatic trypsin through a transglutaminase-catalysed reaction. The conjugates contained an average of 0.7–1.8 mol of polymers per mol of protein, and retained about 61–82% of the original esterolytic activity of trypsin. The optimum pH for trypsin was shifted to alkaline values after enzymatic glycosidation. The thermostability of the polymer–enzyme complexes was increased in about 13.7–50.0 °C over 10 min incubation. The prepared conjugates were also more stable against thermal incubation at different temperatures ranging from 50 °C to 60 °C. In comparison with native trypsin, the enzyme-polymer complexes were about 22- to 48-fold more resistant to autolytic degradation at pH 9.0. Transglutaminase-catalysed glycosidation also protected trypsin against denaturation in surfactant media, with 9- to 68–fold increased half-life times in the presence of 0.3% (w/v) sodium dodecylsulfate.

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Acknowledgments

This research was supported by the International Foundation for Science, Stockholm, Sweden, and the Organization for the Prohibition of Chemical Weapons, The Hague, The Netherlands, through a grant to R. Villalonga (Grant F/3004-1). Financial support from The Third World Academy of Sciences, through a grant to R. Villalonga (Grant 01-279 RG/CHE/LA), is also gratefully acknowledged.

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Correspondence to Reynaldo Villalonga.

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Villalonga, M.L., Villalonga, R., Mariniello, L. et al. Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides. World J Microbiol Biotechnol 22, 595–602 (2006). https://doi.org/10.1007/s11274-005-9076-2

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Keywords

  • Enzyme thermostability
  • modified enzyme
  • polysaccharide
  • transglutaminase
  • trypsin