title: Isolation of a human hepatic 60 kDa aspartylglucosaminidase consisting of three non-identical polypeptides.
abstract: We have characterized the properties of human aspartylglucosaminidase (EC 3.5.1.26), the lysosomal enzyme which is deficient in the human inherited disease aspartylglucosaminuria. The purification procedure from human liver included affinity chromatography, gel filtration, strong-anion- and strong-cation-exchange h.p.l.c., chromatofocusing and reverse-phase h.p.l.c. In a denaturing SDS/polyacrylamide-gel electrophoresis, the 6600-fold purified enzyme was shown to be composed of three non-identical inactive polypeptide chains of molecular masses 24, 18 and 17 kDa. In a native polyacrylamide-gel electrophoresis, these polypeptide chains ran as one active enzyme complex. As judged from the elution position of the native enzyme in a Biogel P-100 gel filtration, the approximate molecular mass of this complex was 60 kDa. The enzyme had a pI of 5.7, a pH optimum at 6, of 0.48 mM and a specific activity of 200,000 nkat for the substrate 2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-D-glucose. The enzyme showed a 57% loss of activity at 60 degrees C after 45 h but was practically inactive after incubation at 72 degrees C for a few minutes. The molecular structure, Km and specific activity as well as the thermostability of the enzyme described here are different from those reported previously for human aspartylglucosaminidase.
authors.affiliation: Department of Medical Chemistry, University of Helsinki, Finland.
authors.forename: M
authors.initials: M
authors.lastname: Baumann
authors.forename: L
authors.initials: L
authors.lastname: Peltonen
authors.forename: P
authors.initials: P
authors.lastname: Aula
authors.forename: N
authors.initials: N
authors.lastname: Kalkkinen
mesh_terms: D000581:Amidohydrolases / Q000302:isolation & purification*; D001227:Aspartylglucosylaminase / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007231:Infant, Newborn; D008099:Liver / Q000201:enzymology*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis
chemical_list: D010446:Peptide Fragments; D000581:Amidohydrolases; D001227:Aspartylglucosylaminase