Modification of triantennary glycopeptide into probes for the asialoglycoprotein receptor of hepatocytes.
Triantennary glycopeptide was oxidized with galactose oxidase to convert the -CH2OH group on terminal galactose residues to the aldehyde group (oxo-form). Kinetic profiling by reverse phase high performance liquid chromatography allowed termination of the reaction when intermediate mono-oxo- and di-oxo-triantennary glycopeptides had been produced. The mixture of the oxo-glycopeptides was derivatized with 2,4-dinitrophenylhydrazine for efficient separation, and each isomeric triantennary hydrazone was separated by reverse phase high performance liquid chromatography. The purified hydrazones were reverted to three original isomeric mono-oxo- and di-oxo-glycopeptides, and a single tri-oxo-glycopeptide. Each of these isomers was characterized by proton NMR by a downfield shift in the anomeric signals of 6-oxo-Gal residue(s). The functionalized glycopeptides were successively modified with dansyl and naphthyl groups through the 6-oxo-Gal residue and the amino terminus of the peptide to prepare three isomeric glycopeptide probes suitable for conformation studies by fluorescence energy transfer measurements. Alternatively, glycopeptides were derivatized by attaching t-butyloxycarbonyl-L-tyrosine to the amino terminus of the peptide, and reductive amination of the 6-oxo-Gal residue, provided three isomeric triantennary photoaffinity probes which allow photolyzable groups to be attached to the newly introduced 6-amino-Gal residue.
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218.
K G
KG
Rice
Y C
YC
Lee
D000345:Affinity Labels; D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins / Q000378:metabolism*; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D005456:Fluorescent Dyes; D005692:Galactose Oxidase / Q000494:pharmacology; D006020:Glycopeptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011971:Receptors, Immunologic / Q000378:metabolism*
D000345:Affinity Labels; D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D005456:Fluorescent Dyes; D006020:Glycopeptides; D011971:Receptors, Immunologic; D005692:Galactose Oxidase