Concanavalin A bound to Sepharose has been used for the purification of brain β-galactosidase, α-L-fucosidase, α-D-mannosidase, arylsulphatase and β-glucuronidase.0 Several factorsviz pH, temperature and concentration of α-methyl glucoside influenced the binding and elution of these enzymes. A lysosomal acid α-mannosidase and a cytosolic neutral mannosidase were separable by concanavalin A-Sepharose chromatography. Similarly lysosomal and microsomal β-glucuronidases were separable using gradient elution with α-methyl glucoside. The results indicate the usefulness of this lectin for the isolation of wide variety of enzymes under specified experimental conditions.

The anionic form of arylsulphatase B (arylsulphatase Bm) was purified to apparent homogeneity from monkey brain through steps involving chromatography on diethylaminoethyl-cellulose, Blue-Sepharose, Biogel HTP and finally Biogel P-300 gel filtration. The molecular weight of the purified enzyme as deduced by gel filtration on Biogel P-300 and by sodium dodecylsulphate gel electrophoresis was ∼ 30,000.Escherichia coli alkaline phosphatase treatment of arylsulphatase Bm resulted in the conversion of upto 84% of the enzyme into a less charged form of enzyme, that could not bind to diethylaminoethyl cellulose. Potassium phosphate an inhibitor of alkaline phosphatase prevented this conversion. Upon acid hydrolysis the purified enzyme yielded approximately 7.0 mol of inorganic phosphate per mol of protein.Vibrio cholerae neuraminidase treatment did not alter the charge on arylsulphatase Bm.

The lysosomal enzyme binding protein (receptor protein) isolated from monkey brain was immobilised on Sepharose 4B and used to study the binding of brain lysosomal enzymes. The immobilised protein could bind \-D-glucosaminidase, α-D-mannosidase, α-L-fucosidase and²-D-glucuronidase. The bound enzymes could be eluted either at an acid pH of 4.5 or by mannose 6-phosphate but not by a number of other sugars tested. Binding could be abolished by prior treatment of the lysosomal enzymes with sodium periodate. Alkaline phosphatase treatment of the enzymes did not prevent the binding of the lysosomal enzymes to the column but decreased their affinity, as seen by a shift in their elution profile, when a gradient elution with mannose 6-phosphate was employed. These results suggested that an ‘uncovered’ phosphate on the carbohydrate moiety of the enzymes was not essential for binding but can enhance the binding affinity.

Sulfoconjugates occur ubiquitously as sulfopolysaccharides, sulfolipids and sulfoproteins. A variety of sulfotransferases catalyze the sulfation process with 3’-phosphoadenosine 5’-phosphosulfate as the sulfate donor. Sulfatases that catalyze the desulfation of different sulfoconjugates are known to be deficient in a number of genetic storage disorders.