Partial purification and properties of porcine thymus lactosylceramide beta-galactosidase.
Porcine thymus lactosylceramide beta-galactosidase was purified by a simple procedure. In the final step of isoelectric focusing the enzyme was separated into two peaks of pI 6.3 (peak I) and 7.0 (peak II), which showed 3,600- and 4,000-fold enhancement of lactosylceramide-hydrolysing activity, respectively. The two peaks had identical mobility on polyacrylamide gel electrophoresis. The apparent molecular weight was 34,000. Neither monosialoganglioside (GM1) nor galactosylceramide was hydrolysed by the purified enzyme fractions. The optimal pH was at 4.6, and sodium taurocholate was essential for the reaction. The apparent Km was 2.3 x 10-5 M. The reaction was stimulated by sodium chloride and linoleic acid, while it was strongly inhibited by Triton X-100 and bovine serum albumin. Galactosylceramide, p-nitrophenyl beta-galactoside, and p-nitrophenol were weak inhibitors. No effects of GM1 and galactose were observed on the hydrolysis of lactosylceramide.
D000818:Animals; D002554:Cerebrosides / Q000494:pharmacology; D005696:Galactosidases / Q000302:isolation & purification / Q000378:metabolism; D005697:Galactosides / Q000494:pharmacology; D005732:Gangliosides / Q000494:pharmacology; D008970:Molecular Weight; D009596:Nitrophenols / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000494:pharmacology; D013552:Swine; D013656:Taurocholic Acid / Q000494:pharmacology; D013950:Thymus Gland / Q000201:enzymology*
D002554:Cerebrosides; D005697:Galactosides; D005732:Gangliosides; D009596:Nitrophenols; D011092:Polyethylene Glycols; D013656:Taurocholic Acid; D005696:Galactosidases