Biosynthesis of glycoproteins in Candida albicans: biochemical characterization of a soluble alpha-mannosidase.
Most alpha-mannosidase activity (80%) in C. albicans was found in a soluble form. Addition of protease inhibitors to explore proteolytic release from a particulate cell component during enzyme preparation did not change this distribution. Molecular mass, calculated from gel filtration chromatography, was 417 kDa. Optimum pH was 6.0 with 50 mM Mes-Tris when p-nitrophenyl-alpha-D-mannopyranoside was used as substrate. Optimum temperature was 42 degrees C with either 10 mM phosphate buffer (pH 6.8) or 50 mM Mes-Tris buffer (pH 6.0) and with 4-methylumbelliferyl-alpha-D-mannopyranoside as substrate. Apparent Km values for p-nitrophenyl-alpha-D-mannopyranoside and 4-methylumbelliferyl-alpha-D-mannopyranoside were 3.3 mM and 0.1 mM, respectively. 1 mM 1-deoxymannojirimycin and 0.3 mM swainsonine inhibited the hydrolysis of 4-methylumbelliferyl-alpha-D-mannopyranoside by 67% and 83%, respectively, whereas that of p-nitrophenyl-alpha-D-mannopyranoside was only slightly diminished (10-15%).
Vázquez-Reyna
A B
AB
Instituto de Investigación en Biología Experimental, Facultad de Química, Universidad de Guanajuato, Mexico.
Balcázar-Orozco
R
R
Flores-Carreón
A
A
D001426:Bacterial Proteins / Q000096:biosynthesis; D002176:Candida albicans / Q000378:metabolism*; D002240:Carbohydrate Sequence; D003300:Copper / Q000494:pharmacology; D006023:Glycoproteins / Q000096:biosynthesis*; D008361:Mannosidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry; D012995:Solubility; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity; D043323:alpha-Mannosidase
D001426:Bacterial Proteins; D006023:Glycoproteins; D009844:Oligosaccharides; D003300:Copper; D008361:Mannosidases; D043323:alpha-Mannosidase