Isolation and characterization of UDP-galactose:N-acetylglucosamine 4 beta-galactosyltransferase activity induced in rat parotid glands treated with isoproterenol.
Chronic injection of the beta-adrenergic receptor agonist isoproterenol causes a 6- to 10-fold increase in the specific activity of the Golgi membrane marker enzyme UDP-galactose:D-glucose 4-beta-D-galactosyltransferase (EC 2.4.1.22) in rat parotid glands. Using a combination of size exclusion chromatography and affinity chromatography on alpha-lactalbumin-agarose, the Triton X-100-solubilized enzyme was purified to homogeneity from treated and control animals. The enzyme purified from both sources had a single Mr = 60,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme isolated from control and isoproterenol-treated animals had identical amino acid compositions and isoelectric points (pI = 5.9). Neutral sugar content as determined by the phenol-sulfuric acid assay was 10%. UDP-galactose:N-acetylglucosamine 4 beta-galactosyltransferase activity purified by alpha-lactalbumin column chromatography was inhibited by the presence of 200 mM N-acetylglucosamine and by the inclusion of alpha-lactalbumin in assays using 10 mM N-acetylglucosamine as acceptor. Increased enzyme activity was immune precipitated from Triton X-100-solubilized membrane prepared from isoproterenol-treated animals versus control animals. This was also true of protein synthesized by in vitro translation of poly(A+) RNA prepared from parotid glands, indicating an increase in mRNA levels for the enzyme. The molecular mass of the immune-precipitated, in vitro translation product was determined to be 45,000 Da.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002417:Cattle; D004790:Enzyme Induction; D005260:Female; D007545:Isoproterenol / Q000494:pharmacology*; D007788:Lactose Synthase / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D008297:Male; D008892:Milk / Q000201:enzymology; D008970:Molecular Weight; D010306:Parotid Gland / Q000187:drug effects / Q000201:enzymology*; D011061:Poly A / Q000235:genetics; D014176:Protein Biosynthesis; D012313:RNA / Q000235:genetics; D012333:RNA, Messenger; D051381:Rats; D011919:Rats, Inbred Strains; D014018:Tissue Distribution
D000596:Amino Acids; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D007788:Lactose Synthase; D007545:Isoproterenol