Stimulation of bovine milk galactosyltransferase activity by bovine colostrum N-acetylglucosaminyltransferase I.
Purified bovine milk galactosyltransferase was stimulated by purified bovine colostrum N-acetylglucosaminyltransferase I by more than 10-fold. Only slight stimulation of the N-acetylglucosaminyltransferase I by galactosyltransferase was observed. Heat inactivation destroyed the ability of the N-acetylglucosaminyltransferase I to stimulate the galactosyltransferase. The stimulation of galactosyltransferase was accompanied by a decrease in Km of this enzyme from 9.7 to 3.3. mM and an increase in Vmax from 1.87 to 3.71 nmol galactose transferred/min per mg galactosyltransferase when GlcNAc was the substrate. When the Km for UDPgalactose was determined, it increased from 0.19 to 0.42 mM in the presence of N-acetylglucosaminyltransferase I and the Vmax increased from 0.66 to 2.76 nmol galactose transferred/min per mg galactosyltransferase. In phosphatidylcholine vesicles, no effect on Km values with GlcNAc as substrate was noted, while an increase in the Km of UDPgalactose was observed. The Vmax values were generally higher in the lipid vesicles. Complex formation between galactosyltransferase and N-acetylglucosaminyltransferase I was demonstrated both by glycerol density gradient centrifugation and Bio-Gel P-100 column chromatography. An approximate molecular weight for the complex was obtained on a calibrated Sephadex G-200 column and found to be about 75 000, consistent with a 1:1 complex. The stimulation of galactosyltransferase involved the N-acetyllactosamine synthetase activity of this enzyme and not the lactose synthetase activity, since the latter activity was only slightly affected. Since N-acetylglucosaminyltransferase I is not involved in the lactose synthetase reaction, the stimulation is consistent with the known biosynthetic role of N-acetylglucosaminyltransferase I in the biosynthesis of asparagine-linked oligosaccharides.
Moscarello
M A
MA
Mitranic
M M
MM
Vella
G
G
D000818:Animals; D002417:Cattle; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D003126:Colostrum / Q000201:enzymology*; D004789:Enzyme Activation; D005700:Galactosyltransferases / Q000378:metabolism*; D005964:Glucosyltransferases / Q000378:metabolism*; D007700:Kinetics; D007788:Lactose Synthase / Q000378:metabolism; D008892:Milk / Q000201:enzymology*; D017351:N-Acetylglucosaminyltransferases; D010713:Phosphatidylcholines; D011485:Protein Binding
D010713:Phosphatidylcholines; D005700:Galactosyltransferases; D005964:Glucosyltransferases; D017351:N-Acetylglucosaminyltransferases; C024850:alpha-1,3-mannosyl-glycoprotein beta-1,2-N-acetylglucosaminyltransferase I; D007788:Lactose Synthase