scholarly journals A new method for the rapid purification of both the membrane-bound and released forms of the variant surface glycoprotein from Trypanosoma brucei

1985 ◽  
Vol 230 (1) ◽  
pp. 195-202 ◽  
Author(s):  
D G Jackson ◽  
M J Owen ◽  
H P Voorheis
Keyword(s):  
Amino Acid ◽  
Trypanosoma Brucei ◽  
Aqueous Salt Solution ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
High Yield ◽  
Dependent Manner ◽  
Wide Range ◽  
Membrane Bound ◽  
Rapid Purification

A simple new technique was developed for the rapid purification of either the membrane-bound or the released forms of the variant surface glycoprotein of Trypanosoma brucei in high yield. Whole cells were used as the source of the membrane-bound form, and the supernatant of benzyl alcohol-treated cells was used as the source of the released form. The technique was based on extraction of the acid-treated protein into chloroform/methanol, followed by selective re-partition into aqueous salt solution. The yield of purified protein was found to be dependent critically on a low pH during the extraction/re-partition stages. This finding and the ability to cycle the protein repeatedly through organic and aqueous phases in a strictly pH-dependent manner suggested that the protein could undergo fully reversible denaturation/renaturation only while in an extensively protonated form. The yield was independent of the polarity of the organic phase and the protein concentration over a wide range. After purification, both forms retain their ability to react with specific antibody raised against the authentic native protein purified by conventional means. The amino acid composition and the identity of the N-terminal amino acid was the same for both forms of the protein. In addition, both forms had blocked C-terminal residues. There were determined to be 1.13 × 10(7) copies of the variant surface glycoprotein per cell.

scholarly journals Carbohydrate is linked through ethanolamine to the C-terminal amino acid of Trypanosoma brucei variant surface glycoprotein

1983 ◽  
Vol 209 (1) ◽  
pp. 261-262 ◽  
Author(s):  
A A Holder
Keyword(s):  
Amino Acid ◽  
Aspartic Acid ◽  
Trypanosoma Brucei ◽  
Carboxy Group ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Side Chain ◽  
Serine Residue ◽  
Terminal Amino Acid ◽  
Terminal Amino

The C-terminal amino acid of the variant surface glycoprotein from Trypanosoma brucei is glycosylated. For two variant proteins that terminate in an aspartic acid and a serine residue respectively, it was shown that the sugar side chain is linked through ethanolamine to the alpha-carboxy group of the amino acid.

scholarly journals Locations of the six disulphide bonds in a variant surface glycoprotein (VSG 117) from Trypanosoma brucei

1983 ◽  
Vol 209 (2) ◽  
pp. 481-487 ◽  
Author(s):  
G Allen ◽  
L P Gurnett
Keyword(s):  
Amino Acid ◽  
Trypanosoma Brucei ◽  
Thiol Group ◽  
Cysteine Residue ◽  
Terminal Region ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
British Library ◽  
African Trypanosome ◽  
Disulphide Bonds

The locations of the six disulphide bonds and the single free cysteine residue in a variant surface glycoprotein, VSG 117, from the African trypanosome Trypanosoma brucei have been determined to be Cys-14-Cys-140, Cys-121-Cys-182, Cys-389-Cys-404, Cys-398-417, Cys-447-Cys-461 and Cys-455-Cys-468. Cys-244 bears the single thiol group, which is unreactive towards 2-nitro-5-thiocyanobenzoate in the native molecule and is probably buried. Biosynthetically incorporated [35S]cysteine aided the location of the disulphide bonds. Two proteinase-resistant glycosylated domains, each containing two disulphide bonds, were identified in the C-terminal region of the glycoprotein. Details of purification of [35S]cysteine-containing peptides, and Tables of amino acid analyses, are presented in Supplementary Publication SUP 50119 (32 pages), which has been deposited with the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1981) 193,5.

scholarly journals Expression of a variant surface glycoprotein of Trypanosoma gambiense in procyclic forms of Trypanosoma brucei shows that the cell type dictates the nature of the glycosylphosphatidylinositol membrane anchor attached to the glycoprotein

1997 ◽  
Vol 324 (3) ◽  
pp. 885-895 ◽  
Author(s):  
Françoise PATURIAUX-HANOCQ ◽  
Nicole ZITZMANN ◽  
Jacqueline HANOCQ-QUERTIER ◽  
Luc VANHAMME ◽  
Sylvie ROLIN ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Degradation Products ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Mrna Levels ◽  
Membrane Anchor ◽  
Gpi Anchor ◽  
Intact Membrane ◽  
Membrane Bound ◽  
Trypanosoma Gambiense

Procyclic forms of Trypanosoma brucei have been genetically modified to express the major metacyclic variant surface glycoprotein (VSG variant AnTat 11.17) of Trypanosoma gambiense. The VSG is expressed in an intact membrane-bound form that can be detected over the entire plasma membrane, together with procyclin, and as a series of lower-molecular-mass fragments that are mostly soluble degradation products. The presence of degraded VSG in the cells and the culture medium suggests that VSG is not efficiently processed and/or efficiently folded when expressed in procyclic cells. The level of procyclin expressed on the surface of these cells is slightly reduced, although there is no difference in procyclin mRNA levels. The intact membrane-bound form of the VSG is N-glycosylated with oligomannose structures and contains a glycosylphosphatidylinositol (GPI) membrane anchor that can be biosynthetically labelled with [3H]ethanolamine. The anchor is sensitive to mammalian GPI-specific phospholipase D but, like the anchor of procyclin, it is resistant to the action of bacterial phosphatidylinositol-specific phospholipase C. This pattern of phospholipase sensitivity suggests that the GPI anchor acquired by VSG when expressed in procyclics is acylated on the inositol ring and therefore resembles a procyclic procyclin-type anchor rather than a trypomastigote VSG-type anchor with respect to the lipid structure. The VSG expressed in procyclics was sensitive to the action of a mixture of sialidase, β-galactosidase and β-hexosaminidase, suggesting that the VSG GPI anchor also contains a sialylated polylactosamine side-chain modification similar to that described for procyclin. These results indicate that the nature of the protein expressed has little influence on the post-translational modifications performed in the secretory pathway of procyclic trypanosomes.

scholarly journals Trypanosoma brucei brucei variant surface glycoprotein contains non-N-acetylated glucosamine

1986 ◽  
Vol 234 (2) ◽  
pp. 481-484 ◽  
Author(s):  
A M Strang ◽  
J M Williams ◽  
M A J Ferguson ◽  
A A Holder ◽  
A K Allen
Keyword(s):  
Amino Acid ◽  
Acid Hydrolysis ◽  
Trypanosoma Brucei ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Amino Group ◽  
Terminal Amino Acid ◽  
Trypanosoma Brucei Brucei ◽  
Parasitic Protozoan ◽  
Terminal Amino

The C-terminal amino acid of the variant surface glycoprotein of the parasitic protozoan Trypanosoma brucei brucei is glycosylated and the oligosaccharide has been shown to contain glucosamine. By acid hydrolysis, HNO2 deamination and 1H-n.m.r. studies we have demonstrated that the amino group of this glucosamine is not N-acetylated and is most probably unmodified.

scholarly journals Synchronous expression of individual metacyclic variant surface glycoprotein genes in Trypanosoma brucei

2015 ◽  
Vol 200 (1-2) ◽  
pp. 1-4 ◽  
Author(s):  
Kiantra Ramey-Butler ◽  
Elisabetta Ullu ◽  
Nikolay G. Kolev ◽  
Christian Tschudi
Keyword(s):  
Trypanosoma Brucei ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein
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