Distribution, purification and characterization of rat intestinal UDPgalactose: N-acetylglucosaminyl(beta 1----4)galactosyltransferase.
Rat intestinal UDPgalactose: N-acetylglucosaminyl(beta 1----4)galactosyltransferase activity was studied as to its intestinal and villus-to-crypt distribution, and then purified and characterized. Rapid UDPgalactose hydrolysis was noted in the duodenum and jejunum; little to no breakdown was detected in the distal ileum, cecum and proximal colon. Product analysis suggested that UDPgalactose hydrolysis was due to nucleotide-sugar pyrophosphatase and galactose-1-phosphate phosphatase activities; ileum appeared to have little of the first activity and none of the latter. An aboral gradient of galactosyltransferase activity was noted, activity being 3-4-fold higher in the ileum, cecum and proximal colon. Total homogenate exogenous acceptor galactosyltransferase activities showed no villus-crypt differences but activity measured with intact isolated cells demonstrated higher activity with crypt cells; this was particularly evident in the ileum. Galactosyltransferase activity was purified from ileal-colonic mucosa. An over 4000-fold purification with 75 percent yield was achieved. Only one band of approx. 70-75 kDa was noted on sodium dodecyl sulfate polyacrylamide electrophoresis. As with other eukaryotic galactosyltransferase activities, there was an absolute requirement for Mn2+; the concentration required for half maximal activity was only 2.5 microM and higher concentrations did not inhibit. The Km for UDPgalactose was 30 microM.
D000818:Animals; D007413:Intestinal Mucosa / Q000201:enzymology; D007422:Intestines / Q000033:anatomy & histology / Q000201:enzymology*; D007788:Lactose Synthase / Q000378:metabolism*; D008297:Male; D008970:Molecular Weight; D009237:N-Acetyllactosamine Synthase / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D013499:Surface Properties; D014018:Tissue Distribution
D007788:Lactose Synthase; D009237:N-Acetyllactosamine Synthase