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Glycoconj J. 2014 Oct;31(6-7):485-96. doi: 10.1007/s10719-014-9540-z.

Purification and characterization of a class II α-Mannosidase from Moringa oleifera seed kernels.

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Department of Biochemistry, Central University of Rajasthan, Kishangarh, Ajmer, 305817, Rajasthan, India,


α-Mannosidase (EC. belonging to class II glycosyl hydrolase family 38 was purified from Moringa oleifera seeds to apparent homogeneity by conventional protein purification methods followed by affinity chromatography on Con A Sepharose and size exclusion chromatography. The purified enzyme is a glycoprotein with 9.3 % carbohydrate and exhibited a native molecular mass of 240 kDa, comprising two heterogeneous subunits with molecular masses of 66 kDa (α-larger subunit) and 55 kDa (β-smaller subunit). Among both the subunits only larger subunit stained for carbohydrate with periodic acid Schiff's staining. The optimum temperature and pH for purified enzyme was 50 °C and pH 5.0, respectively. The enzyme was stable within the pH range of 3.0-7.0. The enzyme was inhibited by EDTA, Hg(2+), Ag(2+), and Cu(2+). The activity lost by EDTA was completely regained by addition of Zn(2+). The purified enzyme was characterized in terms of the kinetic parameters K m (1.6 mM) and V max (2.2 U/mg) using para-nitrophenyl-α-D-mannopyranoside as substrate. The enzyme was very strongly inhibited by swainsonine (SW) at 1 μM concentration a class II α-Mannosidase inhibitor, but not by deoxymannojirimycin (DMNJ). Chemical modification studies revealed involvement of tryptophan at active site. The inhibition by SW and requirement of the Zn(2+) as a metal ion suggested that the enzyme belongs to class II α-Mannosidase.

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