Isolation, characterization and the role of rabbit testicular arysulphatase A in fertilization.
Arysulphatase A was purified from rabbit testis. The purification was accomplished by a four-step procedure involving (NH4)2SO4 fractionation, chromatography on DEAE-cellulose, SP(sulphopropyl)-Sephadex and affinity chromatography on concanavalin A-Sepharose. The specific activity of purified preparation was 135 mumol/min per mg of protein, which represented an increase of 900-fold above that of the crude homogenate. The purified enzyme (20-50 micrograms) was found to move electrophoretically as a single band on polyacrylamide gel at pH 7.2 and 8.4. The homogeneous enzyme was shown to be a glycoprotein with 0.8% (w/w) of N-acetylneuraminic acid and 20% neutral sugar. The treatment of purified enzyme with bacterial neuraminidase had no effect on enzyme activity or kinetic properties, but it changed the elution prolife of rabbit testis arylsulphatase A through DEAE-Sephadex. The purified enzyme was strongly inhibited by Cu2+, Fe3+ and Ag+. It hydrolysed several sulphate esters including cerebroside 3-sulphate, ascorbic acid 2-sulphate and steroid sulphates. Pure arysulphatase was effective in dispersing the cumulus cells of rabbit ova.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D002553:Cerebroside-Sulfatase / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology; D002847:Chromatography, Agarose; D005306:Fertilization; D008297:Male; D009439:Neuraminidase; D011817:Rabbits; D013379:Substrate Specificity; D013429:Sulfatases / Q000378:metabolism*; D013737:Testis / Q000201:enzymology*
D000596:Amino Acids; D002241:Carbohydrates; D013429:Sulfatases; D002553:Cerebroside-Sulfatase; D009439:Neuraminidase