Purification and partial characterization of alpha-N-acetylglucosaminidase from human liver.
A deficiency in alpha-N-acetylglucosaminidase is known as mucopolysaccharidosis IIIB or Sanfilippo B syndrome. We purified this enzyme almost 39,000-fold from liver to homogeneity with 3% recovery. Use of concanavalin A (Con A)-Sepharose and heparin-Sepharose resulted in 13.4-fold and 11.6-fold purifications of the enzymatic activity, respectively. The molecular mass was estimated to be 300 kDa by gel filtration and 80 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The isoelectric point was 5.1, optimal pH was 4.5, and the Km for p-nitrophenyl alpha-N-acetylglucosamine was 0.13-0.20 mM. The purified enzyme was stable at 50 degrees C for 1 h and within the pH range of 6.5-8.5. Anti-serum against the purified enzyme raised in BALB/c mice inhibited the activities of alpha-N-acetylglucosaminidase.
Sasaki
T
T
Department of Pediatrics, Gifu University School of Medicine.
Sukegawa
K
K
Masue
M
M
Fukuda
S
S
Tomatsu
S
S
Orii
T
T
D000118:Acetylglucosaminidase / Q000737:chemistry / Q000302:isolation & purification*; D000818:Animals; D004795:Enzyme Stability; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000201:enzymology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight
C033079:alpha-N-acetyl-D-glucosaminidase; D000118:Acetylglucosaminidase