Transglutaminase-catalysed glycosidation of trypsin with aminated polysaccharides

Dextran (MW=7.2x10(4)), carboxymethylcellulose (MW=2.5x10(4), substitution degree=0.7) and Ficoll (MW=6.9x10(4)) 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 degrees C over 10 min incubation. The prepared conjugates were also more stable against thermal incubation at different temperatures ranging from 50 degrees C to 60 degrees 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.