| First Author | Nomura T | Year | 1998 |
| Journal | J Biol Chem | Volume | 273 |
| Issue | 22 | Pages | 13570-7 |
| PubMed ID | 9593693 | Mgi Jnum | J:47946 |
| Mgi Id | MGI:1206519 | Doi | 10.1074/jbc.273.22.13570 |
| Citation | Nomura T, et al. (1998) Purification, cDNA cloning, and expression of UDP-Gal: glucosylceramide beta-1,4-galactosyltransferase from rat brain. J Biol Chem 273(22):13570-7 |
| abstractText | Lactosylceramide synthase is an enzyme that catalyzes the transfer of galactose from UDP-Gal to glucosylceramide, and thus participates in the biosynthesis of most glycosphingolipids in mammals. We purified this enzyme over 61,000-fold to near homogeneity with a 29. 7% yield from rat brain membrane fractions. The isolation procedure included solubilization with Triton X-100, affinity chromatography on wheat germ agglutinin-agarose and UDP-hexanolamine-agarose, and hydroxylapatite column chromatography, followed by ion exchange chromatography. The final preparation migrated as a broad band with an apparent molecular mass of 61 kDa on SDS-polyacrylamide gel electrophoresis. This apparent molecular mass was reduced to 51 kDa by N-glycanase digestion, suggesting that the enzyme has a glycoprotein nature. The enzyme required Mn2+ for its activity, and glucosylceramide was its preferred substrate. The cDNA for the enzyme was cloned from a rat brain cDNA library. The cDNA insert encoded a polypeptide of 382 amino acid residues, with a molecular weight of 44,776. The polypeptide contained eight putative glycosylation sites and a 20-amino acid residue transmembrane domain at its N terminus. Amino acid sequence homology analysis revealed that this enzyme shared 39% homology with mouse beta- 1, 4-galactosyltransferase (EC 2.4.1.38), which catalyzes the transfer of Gal to beta-1,4-GlcNAc in glycoproteins. |
