Structures of oligomannoside chains of alpha-mannosidase from porcine kidney.
Lysosomal acid alpha-mannosidase from porcine kidney was found to contain mannose (4.8%), galactose (0.9%), fucose (0.5%), N-acetylglucosamine (3.1%), and mannose 6-phosphate (0.1%). Approximately 50% of the total hexose of the oligosaccharide chains could be released by endo-beta-N-acetylglucosaminidase-H (endo-H). They were predominantly neutral, oligomannoside-type oligosaccharides containing 5, 6, and 9 mannose residues, respectively, in the centesimal ratio of 36:25:34. 500-MHz 1H-NMR spectroscopy in conjunction with sequential exoglycosidase digestion of the reduced compounds revealed that each of the three fractions consisted of a single isomer only; the Man9 compound has the following structure: (Formula: see tex). The Man6-compound lacks Man residues D1, D2, and D3, while the Man5-compound lacks Man-C as well. In addition to the neutral ones, some (5%) phosphorylated oligomannoside-type oligosaccharides were obtained. The endo-H resistant glycopeptides were subjected to hydrazinolysis. Approximately 60% of the oligosaccharides released by hydrazine were found to be of rather small size; their composition can be represented asMan2-3GlcNAc[Fuc]0-1GlcNAcol. The remaining 40% consist of larger-size galactose-containing, N-acetyllactosamine-type oligosaccharides. Studies involving sequential exoglycosidase digestion and 500-MHz 1H-NMR spectroscopy performed on the highly purified small-sized compounds revealed the following four structures for the endo-H-resistant oligosaccharides: (Formula: see text).
Y
Y
Kozutsumi
Y
Y
Nakao
T
T
Teramura
T
T
Kawasaki
I
I
Yamashina
J H
JH
Mutsaers
H
H
van Halbeek
J F
JF
Vliegenthart
D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D002850:Chromatography, Gel / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D006834:Hydrazines / Q000494:pharmacology; D006859:Hydrogen; D007668:Kidney / Q000201:enzymology*; D009682:Magnetic Resonance Spectroscopy; D008361:Mannosidases / Q000302:isolation & purification*; D009844:Oligosaccharides / Q000032:analysis* / Q000302:isolation & purification; D013057:Spectrum Analysis / Q000379:methods; D013552:Swine; D043323:alpha-Mannosidase
D002241:Carbohydrates; D006834:Hydrazines; D009844:Oligosaccharides; C029424:hydrazine; D006859:Hydrogen; D008361:Mannosidases; D043323:alpha-Mannosidase