N-acetylgalactosamine-6-sulfate sulfatase in human placenta: purification and characteristics.
N-Acetylgalactosamine-6-sulfate sulfatase from human placenta was purified 33,600-fold using beta-N-acetyl-D-galactosamine 6-sulfate-(1----4)-beta-D-glucuronic acid-(1----3)-N-acetyl-D-[3H]galactosaminitol 6-sulfate as the substrate. This enzyme is an oligomer with a molecular mass of 120 kDa and consists of polypeptides of 40 and 15 kDa. The 15 kDa polypeptide is a glycoprotein. This purified protein has activities of N-acetylgalactosamine-6-sulfate sulfatase and galactose-6-sulfate sulfatase. Rabbit antiserum was raised against the purified protein. The antibody titrated N-acetylgalactosamine-6-sulfate sulfatase and galactose-6-sulfate sulfatase. The size of the precursor of the enzyme is 60 kDa, as determined by cell-free translation. The optimal pH values of the N-acetylgalactosamine-6-sulfate sulfatase and galactose-6-sulfate sulfatase activities are pH 3.8-4.0, and the Kms are 8 and 13 microM, respectively. Sulfate and phosphate ions are potent competitive inhibitors for the enzyme and their inhibition constants are 35 and 200 microM, respectively. Cross-reactive materials of 40 and 15 kDa were detected by immunoblot analysis, in the placenta, liver, and normal fibroblasts, but not in fibroblasts from a patient with Morquio disease.
Masue
M
M
Department of Pediatrics, Gifu University School of Medicine.
Sukegawa
K
K
Orii
T
T
Hashimoto
T
T
D002811:Chondroitinsulfatases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D011487:Protein Conformation; D013379:Substrate Specificity
D002811:Chondroitinsulfatases