Characterization of microsomal and cytosolic alpha-1,2-mannosidases from mung bean hypocotyls.
Microsomal and cytosolic alpha-mannosidase activities, which hydrolyze alpha-1,2-mannosyl-mannose linkages in the Man5GlcNAc2 oligosaccharide, have been isolated from homogenates of mung bean hypocotyls. The alpha-1,2-mannosidase activities were readily distinguished from previously described aryl alpha-mannosidases by several criteria. They were optimally active in the presence of Ca2+ between pH 5.5 and 6, they were inhibited by Zn2+, and they had essentially no activity with p-nitrophenyl-alpha-mannoside. The microsomal and cytosolic alpha-1,2-mannosidases demonstrated specificity for oligosaccharides with terminal nonreducing alpha-1,2-mannosyl linkages, and they were inhibited by mannosyl-mannose disaccharides, with the inhibition decreasing in the order of alpha-1,2-greater than alpha-1,3-greater than alpha-1,6-mannosyl-mannose. The cytosolic alpha-1,2-mannosidase activity, which was present in the 100,000 g supernatant, was separated from the aryl alpha-mannosidase by ammonium sulfate precipitation. The microsomal alpha-1,2-mannosidase, which was tightly associated with the particulate fraction, was solubilized with Triton X-100 and 0.2 M KCl. The two alpha-1,2-mannosidase activities were readily differentiated by gel-filtration chromatography. The solubilized microsomal enzyme chromatographed in approximately the same position as a Mr 460,000 globular protein whereas the cytosolic enzyme was eluted in a retarded position, indicating a much smaller protein.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002850:Chromatography, Gel; D003600:Cytosol / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D008361:Mannosidases / Q000378:metabolism*; D008861:Microsomes / Q000201:enzymology; D008970:Molecular Weight; D010944:Plants / Q000201:enzymology*; D011817:Rabbits; D013379:Substrate Specificity; D015032:Zinc / Q000378:metabolism; D043323:alpha-Mannosidase
D008361:Mannosidases; D043323:alpha-Mannosidase; D015032:Zinc; D002118:Calcium