Synthetic study on glycophosphatidyl inositol (GPI) anchor of trypanosoma brucei: Glycoheptaosyl core

1990 ◽  
Vol 31 (17) ◽  
pp. 2439-2442 ◽  
Author(s):  
Chikara Murakata ◽  
Tomoya Ogawa
2021 ◽  
pp. 101153
Author(s):  
Samuel M. Duncan ◽  
Rupa Nagar ◽  
Manuela Damerow ◽  
Dmitry V. Yashunsky ◽  
Benedetta Buzzi ◽  
...  

2014 ◽  
Vol 63 (3) ◽  
pp. 513-518 ◽  
Author(s):  
Masayuki Nakanishi ◽  
Moe Karasudani ◽  
Takahiro Shiraishi ◽  
Kazunori Hashida ◽  
Mami Hino ◽  
...  

1991 ◽  
Vol 32 (5) ◽  
pp. 671-674 ◽  
Author(s):  
Chikara Murakata ◽  
Tomoya Ogawa

2021 ◽  
Author(s):  
Samuel Martin Duncan ◽  
Rupa Nagar ◽  
Manuela Damerow ◽  
Dmitry V. Yashunsky ◽  
Benedetta Buzzi ◽  
...  

Trypanosoma brucei has large carbohydrate extensions on its N-linked glycans and glycosylphosphatidylinositol (GPI) anchors in its bloodstream form (BSF) and procyclic form (PCF), respectively. The parasites glycoconjugate repertoire suggests at least 38 glycosyltransferase (GT) activities, 16 of which are unknown. Here, we probe the function(s) of a putative β3GT gene, TbGT10. The BSF null mutant is viable in vitro and in vivo and can differentiate into PCF, demonstrating non-essentiality. However, the absence of TbGT10 led to impaired elaboration of N-glycans and GPI anchor sidechains in BSF and PCF parasites, respectively. Glycosylation defects include reduced BSF glycoprotein binding to ricin and to monoclonal antibodies mAb139 and mAbCB1. The latter bind a carbohydrate epitope of lysosomal glycoprotein p67 that we show here, using synthetic glycans, consists of (-6Gal1-4GlcNAc1-)≥4 poly-N-acetyllactosamine repeats. Methylation linkage analysis of Pronase glycopeptides isolated from BSF wild-type and TbGT10 null parasites show a reduction in 6-O-substituted- and 3,6-di-O-substituted-Gal residues. Together, these data suggest that TbGT10 encodes a UDP-GlcNAc : βGal β1-6 GlcNAc-transferase active in both BSF and PCF life-cycle stages elaborating complex N-glycans and GPI sidechains, respectively. The β1-6 specificity of this β3GT gene product and its dual roles in N-glycan and GPI glycan elaboration are notable.


2020 ◽  
Author(s):  
Zhe Ji ◽  
Michele Tinti ◽  
Michael A.J. Ferguson

AbstractThe first step of glycosylphosphatidylinositol (GPI) anchor biosynthesis in all eukaryotes is the addition of N-acetylglucosamine (GlcNAc) to phosphatidylinositol (PI) which is catalysed by a UDP-GlcNAc : PI α1-6 GlcNAc-transferase. This enzyme has been shown to be a complex of at least seven subunits in mammalian cells and a similar complex of homologous subunits has been postulated in yeast. Homologs of most of these mammalian and yeast subunits were identified in the Trypanosoma brucei predicted protein database. The putative catalytic subunit of the T. brucei complex, TbGPI3, was epitope tagged with three consecutive c-Myc sequences at its C-terminus. Immunoprecipitation of TbGPI3-3Myc followed by native polyacrylamide gel electrophoresis and anti-Myc Western blot showed that it is present in a ~240 kDa complex. Label-free quantitative proteomics were performed to compare anti-Myc pull-downs from lysates of TbGPI-3Myc expressing and wild type cell lines. TbGPI3-3Myc was the most highly enriched protein in the TbGPI3-3Myc lysate pull-down and partner proteins TbGPI15, TbGPI9, TbGPI2, TbGPI1 and TbERI1 were also identified with significant enrichment. Our proteomics data also suggest that an Arv1-like protein (TbArv1) is a subunit of the T. brucei complex. Yeast and mammalian Arv1 have been previously implicated in GPI biosynthesis, but here we present the first experimental evidence for physical association of Arv1 with GPI biosynthetic machinery. A putative E2-ligase has also been tentatively identified as part of the T. brucei UDP-GlcNAc : PI α1-6 GlcNAc-transferase complex.Graphical abstractFirst step of GPI anchor biosynthesis pathway in T.brucei BSF is catalysed by TbGPI3 complex.


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