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Endo-N-acetyl-β-D-glucosaminidase (EC 3.2.1.96), which may be involved in fruit ripening by control the free N-glycans, were partially purified and characterized from ´Golden Delicious´ apple fruit. The enzyme was extracted from acetone powder prepared with apple cortical tissues in 50 mM Na-acetate (pH 5.5) containing 2% PVP, 0.3 M KCl, 1% NaHSO₃, 0.5 mM DTT, and 0.1 mM EDTA and purified through ammonium sulfate 30 to 70% saturation, QMA-accell plus FPLC chromatography, Con A-Sepharose 4B, and Superose 12 gel filtration column chromatography in series. The final products resulted in above 700 fold in purity with 0.07% recovery. The enzyme activity of endo-N-acetyl-β-D-glucosaminidase was detected in the range of pH 4 to 9, but the optimum pH was 6.0. The molecular weight of endo-Nacetyl-β-D-glucosaminidase was estimated to be 52.9 kD, when determined through the superose 12 column with standard proteins.
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