Enzyme degradation, high performance liquid chromatography and liquid secondary ion mass spectrometry in the analysis of glycoproteins.
Analysis of small amounts of glycoproteins by high performance liquid chromatography (HPLC) and liquid secondary ion mass spectrometry (LSIMS) together with enzyme digestion has been investigated using fetuin as a model. Preliminary data indicates that 71% of the expected peptides were detected by LSIMS analysis of 200 pmol total digest. HPLC profiles of peptides and glycopeptides were obtained from 2 nmol of digest using a reversed phase (C18) column eluted in a solvent system containing TFA, water and acetonitrile. This has provided glycopeptides for subsequent oligosaccharide analysis. Strategies are reviewed for the chromatographic characterization of oligosaccharides following their release from glycopeptides by chemical and enzymatic procedures.
Smith
K D
KD
Section of Glycoconjugate Research, Clinical Research Centre, Harrow, Middlesex, UK.
Harbin
A M
AM
Carruthers
R A
RA
Lawson
A M
AM
Hounsell
E F
EF
D000595:Amino Acid Sequence; D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D006023:Glycoproteins / Q000032:analysis*; D007477:Ions; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D014357:Trypsin; D000509:alpha-Fetoproteins / Q000032:analysis*
D002241:Carbohydrates; D006023:Glycoproteins; D007477:Ions; D000509:alpha-Fetoproteins; D014357:Trypsin