Purification and properties of the two major isozymes of alpha-galactosidase from human placenta.

This is the authors' abstract. We don't add key points for this paper.

Two cu-galactosidase (a-o-galactoside galactohydrolase, EC 3.2.1.22) isozymes have been highly purified from human placental tissue. Both isozymes appeared to be homogeneous based upon disc gel electrophoresis. Both were bound by concanavalin A-Sepharose and stained positively with the periodic acid-Schiff stain indicating that each contained carbohydrate. The A isozyme had a molecular weight of 103,000 and the B isozyme 117,000 monitored by gel filtration and sodium dodecyl sulfate electrophoresis. Based upon sodium dodecyl sulfate electrophoresis under denaturing conditions, the A isozyme had a minimum subunit molecular weight of 57,700 and the B, 47,700, indicating that each isozyme is a dimer of equal molecular weight subunits. (YGalactosidase A had a K, of 1.55 mM for the artificial substrate 4-methylumbelliferyl-a-o-galactopyranoside while cu-galactosidase B had a K, of 13.1 mM. Purified A isozyme was 300 times more active with the natural substrate, ceramide trihexoside (CTH, Gal-Gal-Glu-ceramide) than purified B isozyme. Each isozyme exhibited a different pattern of inhibition by various carbohydrates. a-Galactosidase A was heat-labile and susceptible to neuraminidase treatment. cu-Galactosidase B was heat-stable, and not susceptible to neuraminidase treatment. Both isozymes had a pH optimum of 4.4 with the artificial substrate I-methylumbelliferyl-cu-o-galactopyranoside and the A isozyme had a pH optimum of 4.1 with ceramide trihexoside. cu-Galactosidase A had a p1 of 4.7 and the B isozyme had a p1 of 4.4. Only artificial cu-galactosidase and ceramide trihexosidase activities co-purified with the A isozyme while artificial agalactosidase, ceramide trihexosidase, and p-nitrophenol-2acetamido-cu-o-galactopyranosidase activities co-purified with the B isozyme. Albumin enhanced the artificial and natural activity of the A isozyme and the artificial activity of the B isozyme. These results support the theory that the two major isozymes of cx-galactosidase from human placenta are not structurally related and do not share a precursor-product relationship.