IN THE BIOSYNTHESIS OF N-GLYCANS IN CONNECTIVE-TISSUE OF THE SNAIL LYMNAEA-STAGNALIS OF INCORPORATION GLCNAC BY BETA-2GLCNAC-TRANSFERASE-I IS AN ESSENTIAL PREREQUISITE FOR THE ACTION OF BETA-2GLCNAC-TRANSFERASE-II AND BETA-2XYL-TRANSFERASE
H. Mulder et al., IN THE BIOSYNTHESIS OF N-GLYCANS IN CONNECTIVE-TISSUE OF THE SNAIL LYMNAEA-STAGNALIS OF INCORPORATION GLCNAC BY BETA-2GLCNAC-TRANSFERASE-I IS AN ESSENTIAL PREREQUISITE FOR THE ACTION OF BETA-2GLCNAC-TRANSFERASE-II AND BETA-2XYL-TRANSFERASE, European journal of biochemistry, 232(1), 1995, pp. 272-283
Using a series of relevant substrates, connective tissue of the snail
Lymnaea stagnalis was shown to contain beta 1-2 xylosyltransferase (be
ta 2Xyl-T), beta 1-2 N-acetylglucosaminyltransferase I (beta 2GlcNAc-T
I), and beta 1-2 N-acetylglucosaminyltransferase II (beta 2GlcNAc-T I
l) activities. These enzymes are probably involved in the biosynthesis
of the N-linked carbohydrate chains, like those present in hemocyanin
. The products formed by incubation of GlcNAc beta 1-2Man alpha 1-6(Gl
cNAc beta 1-2Man alpha 1-3)Man beta 1-R [where R = -4GlcNAc beta 1-4Gl
cNAc or O-(CH2)(7)CH3] with UDP-Xyl and connective tissue microsomes h
ave been purified and characterized by H-1-NMR spectroscopy in conjunc
tion with methylation analysis to be GlcNAc beta 1-2Man alpha 1-6(GlcN
Ac beta 1-2Man alpha 1-3)(Xyl beta 1-2)Man beta 1-R. Substrate specifi
city studies focused on connective tissue beta 2Xyl-T show that the mi
nimal structure requirements are fulfilled in GlcNAc beta 1-2Man alpha
1-3Man beta 1-O-(CH2)(7)CH3. The enzyme activity can therefore be cha
racterized as UDP-Xyl:GlcNAc beta 1-2Man alpha 1-3Man beta-R (Xyl to M
an beta) beta 1-2 xylosyltransferase. In substrate-specificity studies
directed to connective tissue beta 2GlcNAc-T I, it could be demonstra
ted that the enzyme is active towards accepters having at the minimum
a Man alpha 1-3Man beta-R sequence, and that introduction of a beta Xy
l residue at C2 of beta Man totally abolishes the enzyme activity. Xyl
ose-containing oligosaccharides are not accepters for beta 2GlcNAc-T I
. In combination with the substrate specificity of beta 2Xyl-T, this s
hows that in snail connective tissue beta 2GlcNAc-T I must act before
beta 2Xyl-T. The connective tissue beta 2GlcNAc-T II activity follows
the earlier established biosynthetic routes. Based on the substrate sp
ecificities of the various connective tissue glycosyltransferases know
n so far, and the structures isolated from L. stagnalis hemocyanin, a
partial biosynthetic scheme for N-glycosylation in snail connective ti
ssue is proposed.