Synthesis of poly-N-acetyllactosamine in core 2 branched O-glycans. The requirement of novel beta-1,4-galactosyltransferase IV and beta-1,3-n-acetylglucosaminyltransferase.
Poly-N-acetyllactosamine is a unique carbohydrate composed of N-acetyllactosamine repeats and provides the backbone structure for additional modifications such as sialyl Lex. Poly-N-acetyllactosamines in mucin-type O-glycans can be formed in core 2 branched oligosaccharides, which are synthesized by core 2 beta-1,6-N-acetylglucosaminyltransferase. Using a beta-1, 4-galactosyltransferase (beta4Gal-TI) present in milk and the recently cloned beta-1,3-N-acetylglucosaminyltransferase, the formation of poly-N-acetyllactosamine was found to be extremely inefficient starting from a core 2 branched oligosaccharide, GlcNAcbeta1-->6(Galbeta1-->3)GalNAcalpha-->R. Since the majority of synthesized oligosaccharides contained N-acetylglucosamine at the nonreducing ends, galactosylation was judged to be inefficient, prompting us to test novel members of the beta4Gal-T gene family for this synthesis. Using various synthetic acceptors and recombinant beta4Gal-Ts, beta4Gal-TIV was found to be most efficient in the addition of a single galactose residue to GlcNAcbeta1-->6(Galbeta1-->3)GalNAcalpha-->R. Moreover, beta4Gal-TIV, together with beta-1,3-N-acetylglucosaminyltransferase, was capable of synthesizing poly-N-acetyllactosamine in core 2 branched oligosaccharides. On the other hand, beta4Gal-TI was found to be most efficient for poly-N-acetyllactosamine synthesis in N-glycans. In contrast to beta4Gal-TI, the efficiency of beta4Gal-TIV decreased dramatically as the acceptors contained more N-acetyllactosamine repeats, consistent with the fact that core 2 branched O-glycans contain fewer and shorter poly-N-acetyllactosamines than N-glycans in many cells. These results, as a whole, indicate that beta4Gal-TIV is responsible for poly-N-acetyllactosamine synthesis in core 2 branched O-glycans.
Ujita
M
M
Glycobiology Program, Cancer Research Center, The Burnham Institute, La Jolla, California 92037, USA. minoru@burnham-inst.org
McAuliffe
J
J
Schwientek
T
T
Almeida
R
R
Hindsgaul
O
O
Clausen
H
H
Fukuda
M
M
D000818:Animals; D002240:Carbohydrate Sequence; D002417:Cattle; D005690:Galactose / Q000378:metabolism; D005700:Galactosyltransferases / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D008892:Milk / Q000201:enzymology; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D011134:Polysaccharides / Q000096:biosynthesis*; D013379:Substrate Specificity
D011134:Polysaccharides; C037199:poly-N-acetyllactosamine; D005700:Galactosyltransferases; D017351:N-Acetylglucosaminyltransferases; C111678:beta-1,4-galactosyltransferase IV; D005690:Galactose