Giardia duodenalis: direct experimental evidence for the absence of a glycosylphosphatidylinositol anchor in a variant surface protein.
The trophozoites of Giardia duodenalis express variant surface proteins (VSPs) that cover the entire surface of the cell and can be altered by antigenic variation. In the present study, a VSP (VSPH7) expressed by the Giardia GS isolate was purified using Triton-X-114 extraction/phase partitioning and a combination of column chromatography methods. The purified VSP was typed by mass spectrometric fingerprint mapping and peptide sequencing and found to share 58-99.8% peptide identity with the VSPH7 protein sequence previously deduced from the cloned cDNA. Carbohydrate compositional analyses consistently showed the presence of galactose in the VSP preparations but a direct association of carbohydrate with the VSPH7 could not be established. Analysis of the C-terminal part of the purified VSPH7 by off-blot myo-inositol analysis provided for the first time direct experimental evidence that this protein is not modified via a GPI lipid.
Hülsmeier
Andreas J
AJ
Institute of Parasitology, University of Zurich, Winterthurerstrasse 266a, 8057 Zurich, Switzerland.
Köhler
Peter
P
D000595:Amino Acid Sequence; D000818:Animals; D000940:Antigenic Variation; D000953:Antigens, Protozoan / Q000737:chemistry* / Q000276:immunology; D015153:Blotting, Western; D002469:Cell Separation; D002848:Chromatography, DEAE-Cellulose; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D008401:Gas Chromatography-Mass Spectrometry; D016829:Giardia lamblia / Q000737:chemistry* / Q000276:immunology; D017261:Glycosylphosphatidylinositols / Q000737:chemistry*; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D015800:Protozoan Proteins / Q000737:chemistry* / Q000276:immunology; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000953:Antigens, Protozoan; D017261:Glycosylphosphatidylinositols; D015800:Protozoan Proteins; C072464:variant surface protein H7, Giardia lamblia