Multiple Deficiency of Mucopolysaccharide Sulfatases in Mucosulfatidosis

Abstract

Summaryiduronide 2-sulfate sulfatase, sulfamidase, and arylsulfatase B (Nacetylgalactosamine 4-sulfate sulfatase) were assayed in four muFibroblasts of four patients affected with mucosulfatidosis (mul-cosulfatidosis fibroblast lines using natural substrates.tiple sulfatase deficiency, Austin variant of metachromatic leukodystrophy) were assayed for activities of the five sulfatases known to degrade mucopolysaccharides. These were iduronide 2-sulfate sulfatase, sulfamidase, N-acetyl-galactosamine 6-sulfate sulfatase, (specific activity 27 mCi/mmole) from hen oviduct (lo), the monosulfated tri-saccharide N-acetylgalactosamine 6-sulfate Speculation (GlcNAC (6s)) D-glucuronic acid (G1cu~)-[1-3H]anhydromannit01 (specific activity 68.8 mCi/mmole) prepared from heparin Mucosulfatidosis is a disease characterized by the deficiency of sulfate (9), and the disulfated trisaccharide GalNAc(6S)-GlcUAmultiple sulfatases in cultured fibroblasts in contrast to the defi-~-acet~l-[l-~H]galactosaminitol(6S) (specific activity 24.9 mCi/ ciencies of singe sulfatases that are known for the five mucopoly-mmole) prepared from chondroitin 6-sulfate (12) were obtained saccharide degrading sulfatases and arylsulfatase A. The genes as described. responsible for the expression of the sulfatases are located both on autosomes and the X-chromosomes. Mucosulfatidosis fibroblasts should provide an experimental model for the study of a PATIENTS hitherto unknown common mechanism responsible for the exPresSkin biopsies or fibroblast cultures from patients with mucosulsion of sulfatase activities. fatidosis were kindly provided by Drs. J. Couchot (Reims), cell line 1, U. Wiesmann (Bern), cell line 2, and M. Niermeijer Mucosulfatidosis is a lysosomal storage disease characterized by (Rotterdam), line 3. line (GM 2407)9 was from the storage of sulfatides, glycosaminoglycans, and steroid sulfates. the Human Genetic Mutant Camden* N.J.The storage is caused by the deficiency of multiple sulfatases (for review, see refs. I and 2). In fibroblasts and tissues so far the METHODS deficiency of arylsulfatases A, B, and C, cholesteryl sulfatase and dehydroepiandrosterone sulfatase are known (3,4). It is, however, Cell Culture. Human skin fibroblasts from normal individuals still unclear whether activities of arylsulfatase C and the steroid and from patients of various genotypes were cultured in 25 cm2 sulfatases reside in the same enzyme or whether different enzymes Falcon plastic flasks with Eagle's Minimal Essential Medium (18) catalyze these reactions (2). Furthermore, mucosulfatidosis fibro-containing 10% fetal calf serum (LS-Labor Service, Miinchen), blasts do not correct the deranged mucopolysaccharide catabolism nonessential amino acids, and antibiotics as described previously in fibroblasts derived from patients affected with Hunter or Sari-(14). Cultures grown to confluency were harvested by trypsinizafilippo A syndrome which implies that mucosulfatidosis fibre-tion, the cells were washed twice with 0.15 M NaCI, and cell blasts are deficient in iduron...

Molecular Characterization of the Human Cα-formylglycine-generating Enzyme

“…All peptides served as substrates for FGE, albeit the sulfamidase-and the galactose 6-sulfatase-derived peptides showed a rather low turnover. Both sulfamidase and galactose 6-sulfatase are known as substrates of FGE in vivo, as MSD-causing mutations of FGE lead to inactivity of the two sulfatases (31). These data show that FGE is capable of generating FGly residues in all sulfatases.…”

Human liver N-acetylglucosamine-6-sulphate sulphatase. Purification and characterization

“…This is an extremely simple purification protocol utilizing a rapid threecolumn procedure that includes a combined concanavalin A-Sepharose/Blue A-agarose recycling step [9]. Partially purified rat skin 6SS had a molecular mass of 70 kDa and, although most activity did not bind to DEAEcellulose at pH 7.2, a small amount did bind and was eluted by a salt gradient, but was not further characterized [10].…”

“…A single form of 6SS with activity towards GlcNAc6S at the non-reducing terminal of a heparan sulphatederived trisaccharide has been purified 136-fold from human urine with a recovery of 1 % enzyme activity and shown to have a molecular mass of 97 kDa [9]. Habuchi et al [10] have reported 6SS activity in rat skin towards GlcNAc6S residues at the non-reducing end of heparan sulphate-and keratan sulphate-derived substrates with a molecular mass of 70 kDa.…”

“…Evidence of a sulphatase activity towards non-reducing terminal 2-sulphaminoglucosamine 6-sulphate residues in heparan sulphate distinct from activity towards GlcNAc6S residues has been reported to be present in bovine kidney extracts [11]. Resolution of questions raised in these studies [5][6][7][8][9][10][11] about the physical properties and substrate-specificity of 6SS requires the purification of this low-abundance protein to homogeneity in high yields. We now report the purification to homogeneity of two major forms of 6SS from human liver, both with native molecular masses of Vol.…”

Impaired degradation of keratan sulphate by Morquio A fibroblasts

“…However, this proposal could not be verified due to the lack of appropriate substrates of galactose 6-sulphate sulphatase (Gl6ssl et al, 1979;DiFerrante, 1980). We observed recently that N-acetylglucosamine 6-sulphate residues of keratan sulphate-derived oligosaccharides may be removed in vitro as sulphated monosaccharides by fl-N-acetylhexosaminidase A , thus bypassing the action of an N-acetylglucosamine 6-sulphate sulphatase (Basner et al, 1979 . O (Kresse et al, 1980) and other materials (Ludolph et al, 1981) were obtained as described.…”