Purification and characterization of UDP-GlcNAc: GlcNAcbeta 1-6(GlcNAcbeta 1-2)Manalpha 1-R [GlcNAc to Man]-beta 1, 4-N-acetylglucosaminyltransferase VI from hen oviduct.
A new beta1,4-N-acetylglucosaminyltransferase (GnT) responsible for the formation of branched N-linked complex-type sugar chains has been purified 64,000-fold in 16% yield from a homogenate of hen oviduct by column chromatography procedures using Q-Sepharose FF, Ni(2+)-chelating Sepharose FF, and UDP-hexanolamine-agarose. This enzyme catalyzes the transfer of GlcNAc from UDP-GlcNAc to tetraantennary oligosaccharide and produces pentaantennary oligosaccharide with the beta1-4-linked GlcNAc residue on the Manalpha1-6 arm. It requires a divalent cation such as Mn(2+) and has an apparent molecular weight of 72,000 under nonreducing conditions. The enzyme does not act on biantennary oligosaccharide (GnT I and II product), and beta1,6-N-acetylglucosaminylation of the Manalpha1-6 arm (GnT V product) is essential for its activity. This clearly distinguishes it from GnT IV, which is known to generate a beta1-4-linked GlcNAc residue only on the Manalpha1-3 arm. Based on these findings, we conclude that this enzyme is UDP-GlcNAc:GlcNAcbeta1-6(GlcNAcbeta1-2)Manalpha1-R [GlcNAc to Man]-beta1,4-N-acetylglucosaminyltransferase VI. This is the only known enzyme that has not been previously purified among GnTs responsible for antenna formation on the cores of N-linked complex-type sugar chains.
Taguchi
T
T
RIKEN (Institute of Physical and Chemical Research), Wako, Saitama 351-0198, Japan.
Ogawa
T
T
Inoue
S
S
Inoue
Y
Y
Sakamoto
Y
Y
Korekane
H
H
Taniguchi
N
N
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002458:Cell Fractionation; D002645:Chickens; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D005260:Female; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017351:N-Acetylglucosaminyltransferases / Q000302:isolation & purification* / Q000378:metabolism*; D009844:Oligosaccharides / Q000737:chemistry* / Q000378:metabolism*; D010057:Oviducts / Q000201:enzymology*
D009844:Oligosaccharides; D017351:N-Acetylglucosaminyltransferases; C038111:alpha-1,3-mannosylglycoprotein beta-1,4-N-acetylglucosaminyltransferase