Analysis of glycosylphosphatidylinositol membrane anchors by electrospray ionization-mass spectrometry and collision induced dissociation.
The multi-component nature of glycosylphosphatidylinositol membrane anchors makes the analysis of their structure complex. Nuclear magnetic resonance spectroscopy of delipidated glycosylphosphatidylinositol-peptide fractions can supply considerable information but requires relatively large quantities of material. High-sensitivity sequencing techniques are available for the oligosaccharide portions of glycosylphosphatidylinositol anchors, but there is no simple and generally applicable technique to complement this information. In this paper we describe the application of electrospray ionization-mass spectrometry and collision induced dissociation to study intact glycosylphosphatidylinositol-peptides from a Trypanosoma brucei variant surface glycoprotein. Collision of the [M + 4H]4+ pseudomolecular ions of two glycosylphosphatidylinositol-peptide glycoforms produced easily interpretable daughter ion spectra, from which detailed information on the lipid moiety, carbohydrate sequence and site of peptide attachment could be obtained. All of the collision induced dissociation cleavage events occurred in the glycosylphosphatidylinositol portion of the glycosylphosphatidylinositol-peptide. This technique supplies complementary data to the high-sensitivity oligosaccharide sequencing procedures and should greatly assist glycosylphosphatidylinositol anchor structure-function studies, particularly when sample quantities are limiting.
Department of Biochemistry, University of Dundee, Scotland, UK.
C A
CA
Redman
B N
BN
Green
J E
JE
Thomas-Oates
V N
VN
Reinhold
M A
MA
Ferguson
D000595:Amino Acid Sequence; D000818:Animals; D002240:Carbohydrate Sequence; D017261:Glycosylphosphatidylinositols / Q000032:analysis*; D013058:Mass Spectrometry / Q000379:methods; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D014346:Trypanosoma brucei brucei / Q000737:chemistry
D017261:Glycosylphosphatidylinositols; D010446:Peptide Fragments