scholarly journals Site of palmitoylation of a phospholipase C-resistant glycosylphosphatidylinositol membrane anchor

1992 ◽  
Vol 284 (2) ◽  
pp. 297-300 ◽  
Author(s):  
M A J Ferguson
Keyword(s):  
Phospholipase C ◽  
Trypanosoma Brucei ◽  
Periodate Oxidation ◽  
Membrane Anchor ◽  
Glycosyl Phosphatidylinositol ◽  
Oxidation Analysis ◽  
Inositol Ring ◽  
Repetitive Protein

The site of palmitoylation of the phosphatidylinositol moiety of the glycosyl-phosphatidylinositol membrane anchor of Trypanosoma brucei procyclic acidic repetitive protein was studied by using periodate oxidation. Analysis of the products by g.c.-m.s. allowed the assignment of 40 and 60% of the palmitate to the 2-position and the 3-position respectively of the myo-inositol ring.

Intracellular localization of the glycosyl-phosphatidylinositol-specific phospholipase C of Trypanosoma brucei

1989 ◽  
Vol 93 (2) ◽  
pp. 233-240
Author(s):  
R. Bolow ◽  
G. Griffiths ◽  
P. Webster ◽  
Y.D. Stierhof ◽  
F.R. Opperdoes ◽  
...  
Keyword(s):  
Phospholipase C ◽  
Trypanosoma Brucei ◽  
Cellular Localization ◽  
Intracellular Localization ◽  
Surface Glycoprotein ◽  
Glycosyl Phosphatidylinositol ◽  
Membrane Bound ◽  
Cyclic Phosphate ◽  
Inositol Ring ◽  
Cytoplasmic Side

Glycosyl-phosphatidylinositol-specific phospholipase C (GPI-PLC) is a membrane-bound enzyme of bloodstream forms of Trypanosoma brucei, which cleaves the GPI-membrane anchor of the variant surface glycoprotein forming diacylglycerol and 1,2-cyclic phosphate on the inositol ring. The cellular localization of the enzyme was studied by fractionation of sub-cellular organelles and immunofluorescence microscopy and was found to be primarily cytoplasmic. This was confirmed by immuno-electron microscopy using cryo-sections, which showed that the labelling was predominantly on the cytoplasmic side of intracellular membranes but was absent from the plasma membrane including the region lining the flagellar pocket. The significance of these results for the possible function of the phospholipase is discussed.

scholarly journals A simple purification of procyclic acidic repetitive protein and demonstration of a sialylated glycosyl-phosphatidylinositol membrane anchor

1993 ◽  
Vol 291 (1) ◽  
pp. 51-55 ◽  
Author(s):  
M A Ferguson ◽  
P Murray ◽  
H Rutherford ◽  
M J McConville
Keyword(s):  
Hydrophobic Interaction Chromatography ◽  
Glycosylation Site ◽  
Surface Glycoprotein ◽  
Membrane Anchor ◽  
Cell Surface Glycoprotein ◽  
African Trypanosomes ◽  
Glycosyl Phosphatidylinositol ◽  
Repeat Domain ◽  
Rapid Purification ◽  
Repetitive Protein

The procyclic acidic repetitive protein is the major cell-surface glycoprotein of the insect-dwelling procyclic forms of the Trypanosoma brucei species of African trypanosomes. The glycoprotein contains an acidic Glu-Pro repeat domain, a glycosyl-phosphatidylinositol membrane anchor and a putative asparagine glycosylation site. In this paper we describe a rapid purification scheme for this glycoprotein, using solvent extraction and hydrophobic interaction chromatography, and a partial characterization of the glycosylphosphatidylinositol membrane anchor. The carbohydrate composition of the anchor is extremely unusual; it contains on average nine GlcNAc, nine Gal, and five sialic acid residues. This is the first description of such a heavily substituted and negatively charged anchor. A comparison between the trypanosome procyclic surface and the Leishmania promastigote surface is also presented.

Phospholipase Resistance of the Glycosyl-Phosphatidylinositol Membrane Anchor on Human Alkaline Phosphatase

1992 ◽  
Vol 38 (12) ◽  
pp. 2517-2525 ◽  
Author(s):  
Y W Wong ◽  
M G Low
Keyword(s):  
Alkaline Phosphatase ◽  
Cell Surface ◽  
Human Cell Lines ◽  
Membrane Anchor ◽  
Gpi Anchor ◽  
Glycosyl Phosphatidylinositol ◽  
In Vivo Release ◽  
Mammalian Tissues ◽  
Inositol Ring

Abstract Alkaline phosphatase (ALP) is attached to the cell surface in mammalian tissues via a glycosyl-phosphatidylinositol (GPI) anchor and can be released from the membrane by GPI-specific phospholipases. In a range of cultured human cell lines, however, the sensitivity of ALP to phospholipases was observed to be variable in magnitude (approximately 20-90%). The mechanism of phospholipase resistance was explored with phospholipases of different bond specificities. The results suggest that phospholipase resistance is the result of acylation of the inositol ring in the GPI anchor. The occurrence of phospholipase-resistant forms of ALP may have important implications for the in vivo release and disposition of plasma ALP.

scholarly journals Ectoenzymes of the kidney microvillar membrane. Differential solubilization by detergents can predict a glycosyl-phosphatidylinositol membrane anchor

1988 ◽  
Vol 250 (3) ◽  
pp. 865-869 ◽  
Author(s):  
N M Hooper ◽  
A J Turner
Keyword(s):  
Membrane Proteins ◽  
Phospholipase C ◽  
Transmembrane Proteins ◽  
Sodium Deoxycholate ◽  
Membrane Anchor ◽  
Glycosyl Phosphatidylinositol ◽  
Octyl Glucoside

The pattern of solubilization of nine kidney microvillar ectoenzymes by a range of detergents distinguished two classes of membrane proteins: those released from the membrane by bacterial phosphatidylinositol-specific phospholipase C and those not so released. The latter group of transmembrane proteins were solubilized efficiently (greater than 80%) by all the detergents examined. In contrast, proteins released by phosphatidylinositol-specific phospholipase C were solubilized effectively only by octyl glucoside, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulphonate and sodium deoxycholate. Octyl glucoside solubilized the amphipathic forms of the ectoenzymes examined, suggesting that this may be a useful detergent in the purification of glycosyl-phosphatidylinositol-anchored ectoenzymes.

scholarly journals The hydrophobic mannoside Manα1-6Manα1-S-(CH2)7-CH3 acts as an acceptor for the UDP-Gal:glycosylphosphatidylinositol anchor α1,3-galactosyltransferase of Trypanosoma brucei

1995 ◽  
Vol 309 (3) ◽  
pp. 877-882 ◽  
Author(s):  
S Pingel ◽  
R A Field ◽  
M L S Güther ◽  
M Duszenko ◽  
M A J Ferguson
Keyword(s):  
Trypanosoma Brucei ◽  
Electrospray Mass Spectrometry ◽  
Periodate Oxidation ◽  
Major Product ◽  
Side Chain ◽  
Membrane Anchor ◽  
Gpi Anchor ◽  
The Core ◽  
Variant Surface Glycoproteins

The variant surface glycoproteins (VSGs) of Trypanosoma brucei are attached to the plasma membrane via a glycosylphosphatidylinositol (GPI) membrane anchor. This anchor contains the core sequence ethanolamine-PO4-6Man alpha 1-2Man alpha 1-6Man alpha 1-4GlcN alpha 1-6myo-inositol, which is conserved in all GPI anchors, and a unique alpha Gal side chain attached to the 3-position of the alpha Man residue adjacent to the alpha GlcN residue. Here we report that trypanosome membranes can catalyse the transfer of Gal from UDP-Gal to the hydrophobic thioglycoside Man alpha 1-6Man alpha 1-S-(CH2)7-CH3. Characterization of the galactosylated products by electrospray mass spectrometry, exoglycosidase digestion and periodate-oxidation studies revealed that the major product was Man alpha 1-6(Gal alpha 1-3)Man alpha 1-S-(CH2)7-CH3. The similarity of this product to part of the mature VSG GPI anchor suggests that the thioglycoside is able to act as an acceptor for the trypanosome-specific UDP-Gal-GPI anchor alpha 1,3-galactosyltransferase.

Sign in / Sign up

Export Citation Format

Share Document