Chemo-enzymatic synthesis of conformationally constrained oligosaccharides.
N-Acetyllactosamine derivative 4, which has a methylene amide tether between C-6 and C-2', was enzymatically glycosylated using rat liver alpha-2,6-sialyltransferase (ST6GalI) or recombinant human fucosyltransferase V (FucT-V) to give conformationally constrained trisaccharides 5 and 6, respectively. The methylene amide linker of 4 was installed by a two-step procedure, which involved acylation of a C-6 amino function of a LacNAc derivative with chloroacetic anhydride followed by macrocyclization by nucleophilic displacement of the chloride by a C-2' hydroxyl. The conformational properties of 4 were determined by a combination of NOE and trans-glycosidic heteronuclear coupling constant measurements and molecular mechanics simulations and these studies established that the glycosidic linkage of 4 is conformationally constrained and resides in only one of the several energy minima accessible to LacNAc. The apparent kinetic parameters of transfer to LacNAc and conformationally constrained saccharides 3 and 4 indicates that fucosyltransferase V recognize LacNAc in its A-conformer whereas alpha-2,6-sialyltransferase recognizes the B-conformer of LacNAc.
Galan
M Carmen
MC
Complex Carbohydrate Research Center, University of Georgia, 220 Riverbend Road, Athens, GA 30602, USA.
Venot
Andre P
AP
Glushka
John
J
Imberty
Anne
A
Boons
Geert-Jan
GJ
D000577:Amides / Q000737:chemistry; D000606:Amino Sugars / Q000737:chemistry; D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D005643:Fucose / Q000737:chemistry; D005647:Fucosyltransferases / Q000737:chemistry; D006153:Guanosine Diphosphate / Q000737:chemistry; D006838:Hydrocarbons; D008099:Liver / Q000201:enzymology; D009682:Magnetic Resonance Spectroscopy; D008697:Methane / Q000031:analogs & derivatives* / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D008962:Models, Theoretical; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis* / Q000737:chemistry*; D011487:Protein Conformation; D051381:Rats; D012799:Sialyltransferases / Q000378:metabolism; D014312:Trisaccharides / Q000737:chemistry; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D000577:Amides; D000606:Amino Sugars; D006838:Hydrocarbons; D009844:Oligosaccharides; D014312:Trisaccharides; D006153:Guanosine Diphosphate; C030011:carbene; D005643:Fucose; C000458:N-acetyllactosamine; D005647:Fucosyltransferases; C435083:fucosyltransferase V; D012799:Sialyltransferases; D008697:Methane; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase