[Fluorescent derivatives of carbohydrates in the analysis of the structure of glycoconjugates. N-(4-methylcoumarin-7-yl) glycamines in the study of asparagine-bound carbohydrate chains of glycoproteins].
4-(N-Methylcoumarin-7-yl) glycamines were employed in studying asparagine-linked carbohydrate chains of acid desialylated fetuin. The procedure was optimised for the reductive amination of oligosaccharides with 7-amino-4-methylcoumarin in the presence of Na(CN)BH3 to lead to oligosaccharide glycamines (AMC-OS). AMC-OS were obtained from dextran oligosaccharides and from oligosaccharides released by hydrazinolysis of asparagine-linked sugar chains of asialofetuin. Reverse-phase HPLC and exclusion HPLC with fluorimetric quantitation of AMC-OS is described. TSK Gel 2000 SW column was calibrated using dextran AMC-OS to give linear relationship ln Ni = k(ti/tr)+b, where ti/tr is retention time of the AMC-OS relatively to the reference AMC-trisaccharide, and Ni is calibration unit value, characterizing molecular size of AMC-OS. Three AMC-OS, Gal3GlcNAc3Man3GlcNAc2-AMC (I) and (II), and Gal2GlcNAc3Man3GlcNAc2AMC (III), were obtained from asialofetuin in a molar ration of 1:1.8:0.1. Acid treatment of AMK-OS (II) in desialylation conditions also gave AMC-OS (III), thus suggesting a partial degalactosylation of the glycoprotein sugar chains during the desialylation. Consequent digestion of AMC-OS (II) and (III) with Jack bean beta-galactosidase and beta-N-acetylhexosaminidase led to the same AMC-OS, Man3GlcNAc2AMC. The final digestion product of AMC-OS (I) was GalGlcNAcMan3GlcNAc2AMC, suggesting a structural difference in one of the antennas of the minor sugar chain of asialofetuin. The monosaccharide quantitation and exoglycosidase sequencing were carried out at a 100 pmole level.
A Ia
AIa
Khorlin
S D
SD
Shiian
V V
VV
Nasonov
V A
VA
Markin
D001216:Asparagine / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D003374:Coumarins; D005456:Fluorescent Dyes; D006023:Glycoproteins / Q000032:analysis*; D006868:Hydrolysis; D009844:Oligosaccharides / Q000032:analysis
D003374:Coumarins; D005456:Fluorescent Dyes; D006023:Glycoproteins; D009844:Oligosaccharides; D001216:Asparagine; C028743:7-amino-4-methylcoumarin
Fluorestsentnye proizvodnye uglevodov v strukturnom analize glikokon''iugatov. N-(4-metilkumarin-7-il)glikaminy v izuchenii asparaginsviazannykh uglevodnykh tsepeĭ glikoproteinov.