Enzymatic Midchain Branching of Polylactosamine Backbones Is Restricted in a Site-Specific Manner in 1,3-Fucosylated Chains
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- 作者：Anne Leppä ; nen ; Ritva Niemelä ; and Ossi Renkonen
- 刊名：Biochemistry
- 出版年：1997
- 出版时间：November 4, 1997
- 年：1997
- 卷：36
- 期：44
- 页码：13729 - 13735
- 全文大小：252K
- 年卷期：v.36,no.44(November 4, 1997)
- ISSN：1520-4995
文摘
Branched polylactosamines on animal cell surfaces are believed tocontribute to multivalentinteractions in cell adhesion and cell signalling. Theirbiosynthesis proceeds via linear precursorsthatbecome branched by 
1,6-GlcNAc transferases (IGnT6, GlcNAc to Gal).Previous work has identifiedthe tetrasaccharide Gal1-4GlcNAc1-3Gal1-4GlcNAc(1) and the hexasaccharideGal1-4GlcNAc1-3Gal1-4GlcNAc1-3Gal1-4GlcNAc (4) as acceptorsfor a rat serum enzyme activity (cIGnT6), whichtransfers GlcNAc1-6 units to the midchain galactose residues.Thereby, 1 is converted to the branchedpentasaccharideGal1-4GlcNAc1-3(GlcNAc1-6)Gal1-4GlcNAc and4 to the doubly branchedoctasaccharideGal1-4GlcNAc1-3(GlcNAc1-6)Gal1-4GlcNAc1-3(GlcNAc1-6)Gal1-4GlcNAc [Leppänen, A., Salminen, H., Zhu, Y., Maaheimo, H.,Helin, J., Costello, C. E., & Renkonen,O. (1997) Biochemistry 36, 7026-7036]. Here wereport that neither the 
1,3-fucose-containingderivativesof 1[Gal1-4GlcNAc1-3Gal1-4(Fuc1-3)GlcNAc andGal1-4(Fuc1-3)GlcNAc1-3Gal1-4GlcNAc] nor a similar derivative of 4[Gal1-4GlcNAc1-3Gal1-4(Fuc1-3)GlcNAc1-3Gal1-4GlcNAc] were acceptors for the rat serum cIGnT6 activity. Hence,the enzyme's branch-forming actionwas completely prevented at sites in the immediate neighborhood of thefucosylated loci of thepolylactosamines. InGal1-4GlcNAc1-3Gal1-4GlcNAc1-3Gal1-4(Fuc1-3)GlcNAc,theinhibition of the branch-forming reaction was restricted to thefucose-carrying LacNAc unit; at the middleLacNAc, the branching proceeded normally. However, in the isomericGal1-4(Fuc1-3)GlcNAc1-3Gal1-4GlcNAc1-3Gal1-4GlcNAc, the fucose residueprevented branching completely at the middleLacNAc and almost completely at the reducing end LacNAc. Insummary, 1,3-fucose residues inpolylactosamine chains inhibited the cIGnT6 reaction in a site-specificmanner, at the fucosylated LacNAcunit itself and also at sites one and two LacNAc units upstream, butnot at the LacNAc units downstreamfrom the fucosylated locus. These data imply thatsite-directed branching in polylactosamines is possiblein vitro with the aid of specifically positioned1,3-fucosyl units, that can be removed afterwardwithoutharming the branched backbones.
1,6-GlcNAc transferases (IGnT6, GlcNAc to Gal).Previous work has identifiedthe tetrasaccharide Gal1-4GlcNAc1-3Gal1-4GlcNAc(1) and the hexasaccharideGal1-4GlcNAc1-3Gal1-4GlcNAc1-3Gal1-4GlcNAc (4) as acceptorsfor a rat serum enzyme activity (cIGnT6), whichtransfers GlcNAc1-6 units to the midchain galactose residues.Thereby, 1 is converted to the branchedpentasaccharideGal1-4GlcNAc1-3(GlcNAc1-6)Gal1-4GlcNAc and4 to the doubly branchedoctasaccharideGal1-4GlcNAc1-3(GlcNAc1-6)Gal1-4GlcNAc1-3(GlcNAc1-6)Gal1-4GlcNAc [Leppänen, A., Salminen, H., Zhu, Y., Maaheimo, H.,Helin, J., Costello, C. E., & Renkonen,O. (1997) Biochemistry 36, 7026-7036]. Here wereport that neither the 1,3-fucose-containingderivativesof 1[Gal1-4GlcNAc1-3Gal1-4(Fuc1-3)GlcNAc andGal1-4(Fuc1-3)GlcNAc1-3Gal1-4GlcNAc] nor a similar derivative of 4[Gal1-4GlcNAc1-3Gal1-4(Fuc1-3)GlcNAc1-3Gal1-4GlcNAc] were acceptors for the rat serum cIGnT6 activity. Hence,the enzyme's branch-forming actionwas completely prevented at sites in the immediate neighborhood of thefucosylated loci of thepolylactosamines. InGal1-4GlcNAc1-3Gal1-4GlcNAc1-3Gal1-4(Fuc1-3)GlcNAc,theinhibition of the branch-forming reaction was restricted to thefucose-carrying LacNAc unit; at the middleLacNAc, the branching proceeded normally. However, in the isomericGal1-4(Fuc1-3)GlcNAc1-3Gal1-4GlcNAc1-3Gal1-4GlcNAc, the fucose residueprevented branching completely at the middleLacNAc and almost completely at the reducing end LacNAc. Insummary, 1,3-fucose residues inpolylactosamine chains inhibited the cIGnT6 reaction in a site-specificmanner, at the fucosylated LacNAcunit itself and also at sites one and two LacNAc units upstream, butnot at the LacNAc units downstreamfrom the fucosylated locus. These data imply thatsite-directed branching in polylactosamines is possiblein vitro with the aid of specifically positioned1,3-fucosyl units, that can be removed afterwardwithoutharming the branched backbones.