Insect cells encode a class II alpha-mannosidase with unique properties.
Previously, we cloned and characterized an insect (Sf9) cell cDNA encoding a class II alpha-mannosidase with amino acid sequence and biochemical similarities to mammalian Golgi alpha-mannosidase II. Since then, it has been demonstrated that other mammalian class II alpha-mannosidases can participate in N-glycan processing. Thus, the present study was performed to evaluate the catalytic properties of the Sf9 class II alpha-mannosidase and to more clearly determine its relationship to mammalian Golgi alpha-mannosidase II. The results showed that the Sf9 enzyme is cobalt-dependent and can hydrolyze Man(5)GlcNAc(2) to Man(3)GlcNAc(2), but it cannot hydrolyze GlcNAcMan(5)GlcNAc(2). These data establish that the Sf9 enzyme is distinct from Golgi alpha-mannosidase II. This enzyme is not a lysosomal alpha-mannosidase because it is not active at acidic pH and it is localized in the Golgi apparatus. In fact, its sensitivity to swainsonine distinguishes the Sf9 enzyme from all other known mammalian class II alpha-mannosidases that can hydrolyze Man(5)GlcNAc(2). Based on these properties, we designated this enzyme Sf9 alpha-mannosidase III and concluded that it probably provides an alternate N-glycan processing pathway in Sf9 cells.
Department of Molecular Biology, University of Wyoming, Laramie, Wyoming 82071-3944, USA.
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D000818:Animals; D002413:Cations, Divalent / Q000494:pharmacology; D002460:Cell Line; D002846:Chromatography, Affinity; D005260:Female; D017930:Genes, Reporter; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006056:Golgi Apparatus / Q000201:enzymology; D049452:Green Fluorescent Proteins; D007700:Kinetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008322:Mammals; D008361:Mannosidases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D010053:Ovary; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D018411:Spodoptera / Q000201:enzymology*; D013379:Substrate Specificity
D002413:Cations, Divalent; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D005982:Glutathione Transferase; D008361:Mannosidases; C038829:mannosyl-oligosaccharide 1,3 - 1,6-alpha-mannosidase