Dataset Information

Synthesis of Gal?(1,3)Gal?(1,4)GlcNAc?-, Gal?(1,4)GlcNAc?- and GlcNAc-containing neoglycoproteins and their immunological evaluation in the context of Chagas disease.

ABSTRACT: The protozoan parasite, Trypanosoma cruzi, the etiologic agent of Chagas disease (ChD), has a cell surface covered by immunogenic glycoconjugates. One of the immunodominant glycotopes, the trisaccharide Gal?(1,3)Gal?(1,4)GlcNAc?, is expressed on glycosylphosphatidylinositol-anchored mucins of the infective trypomastigote stage of T. cruzi and triggers high levels of protective anti-?-Gal antibodies (Abs) in infected individuals. Here, we have efficiently synthesized the mercaptopropyl glycoside of that glycotope and conjugated it to maleimide-derivatized bovine serum albumin (BSA). Chemiluminescent-enzyme-linked immunosorbent assay revealed that Gal?(1,3)Gal?(1,4)GlcNAc?-BSA is recognized by purified anti-?-Gal Abs from chronic ChD patients ?230-fold more strongly than by anti-?-Gal Abs from sera of healthy individuals (NHS anti-?-Gal). Similarly, the pooled sera of chronic Chagas disease patients (ChHSP) recognized Gal?(1,3)Gal?(1,4)GlcNAc? ?20-fold more strongly than pooled NHS. In contrast, the underlying disaccharide Gal?(1,4)GlcNAc? and the monosaccharide GlcNAc? or GlcNAc? conjugated to BSA are poorly or not recognized by purified anti-?-Gal Abs or sera from Chagasic patients or healthy individuals. Our results highlight the importance of the terminal Gal? moiety for recognition by Ch anti-?-Gal Abs and the lack of Abs against nonself Gal?(1,4)GlcNAc? and GlcNAc? glycotopes. The substantial difference in binding of Ch vs. NHS anti-?-Gal Abs to Gal?(1,3)Gal?(1,4)GlcNAc?-BSA suggests that this neoglycoprotein (NGP) might be suitable for experimental vaccination. To this end, the Gal?(1,3)Gal?(1,4)GlcNAc?-BSA NGP was then used to immunize ?1,3-galactosyltransferase-knockout mice, which produced antibody titers 40-fold higher as compared with pre-immunization titers. Taken together, our results indicate that the synthetic Gal?(1,3)Gal?(1,4)GlcNAc? glycotope coupled to a carrier protein could be a potential diagnostic and vaccine candidate for ChD.

SUBMITTER: Schocker NS

PROVIDER: S-EPMC4672149 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Synthesis of Galα(1,3)Galβ(1,4)GlcNAcα-, Galβ(1,4)GlcNAcα- and GlcNAc-containing neoglycoproteins and their immunological evaluation in the context of Chagas disease.

Schocker Nathaniel S NS Portillo Susana S Brito Carlos R N CR Marques Alexandre F AF Almeida Igor C IC Michael Katja K

Glycobiology 20150918 1

The protozoan parasite, Trypanosoma cruzi, the etiologic agent of Chagas disease (ChD), has a cell surface covered by immunogenic glycoconjugates. One of the immunodominant glycotopes, the trisaccharide Galα(1,3)Galβ(1,4)GlcNAcα, is expressed on glycosylphosphatidylinositol-anchored mucins of the infective trypomastigote stage of T. cruzi and triggers high levels of protective anti-α-Gal antibodies (Abs) in infected individuals. Here, we have efficiently synthesized the mercaptopropyl glycoside ...[more]

PMID: 26384953

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Project description:The protozoan parasite Leishmania (Viannia) braziliensis is among Latin America's most widespread Leishmania species and is responsible for tegumentary leishmaniasis (TL). This disease has multiple clinical presentations, with cutaneous leishmaniasis (CL) being the most frequent. It manifests as one or a few localized skin ulcers, which can spread to other body areas. Hence, early diagnosis and treatment, typically with pentavalent antimonials, is critical. Traditional diagnostic methods, like parasite culture, microscopy, or the polymerase chain reaction (PCR) for detection of the parasite DNA, have limitations due to the uneven distribution of parasites in biopsy samples. Nonetheless, studies have revealed high levels of parasite-specific anti-α-Gal antibodies in L. (V.) braziliensis-infected patients. Previously, we demonstrated that the neoglycoprotein NGP28b, consisting of the L. (Leishmania) major type-2 glycoinositolphospholipid (GIPL)-3-derived trisaccharide Galpα1,6Galpα1,3Galfβ conjugated to bovine serum albumin (BSA) via a linker, acts as a reliable serological biomarker (BMK) for L. (V.) braziliensis infection in Brazil. This indicates the presence of GIPL-3 or a similar structure in this parasite, and its terminal trisaccharide either functions as or is part of an immunodominant glycotope. Here, we explored whether extending the trisaccharide with a mannose unit would enhance its efficacy as a biomarker for the serological detection of L. (V.) braziliensis. We synthesized the tetrasaccharide Galpα1,6Galpα1,3Galfβ1,3Manpα(CH2)3SH (G31SH) and conjugated it to maleimide-functionalized BSA to afford NGP31b. When we assessed the efficacy of NGP28b and NGP31b by chemiluminescent enzyme-linked immunosorbent assay on a cohort of CL patients with L. (V.) braziliensis infection from Bolivia and Argentina against a healthy control group, both NGPs exhibited similar or identical sensitivity, specificity, and accuracy. This finding implies that the mannose moiety at the reducing end is not part of the glycotope recognized by the parasite-specific anti-α-Gal antibodies in patients' sera, nor does it exert a relevant influence on the terminal trisaccharide's conformation. Moreover, the mannose does not seem to inhibit glycan-antibody interactions. Therefore, NGP31b is a viable and dependable BMK for the serodiagnosis of CL caused by L. (V.) braziliensis.

Project description:The ?-Gal epitope (Gal?1,3Gal?1,4GlcNAc?R) is ubiquitously presented in non-primate mammals, marsupials and New World Monkeys, but it is absent in humans, apes and Old World monkeys. However, the anti-Gal antibody (~1% of immunoglobulins) is naturally generated in human, and is found as the immunoglobulin G (IgG), IgM and IgA isotypes. Owing to the specific binding of the anti?Gal antibody with the ??Gal epitope, humans have a distinct anti???gal reactivity, which is responsible for hyperacute rejection of organs transplanted from ??gal donors. In addition, the ?1,3 galactosyltransferases (?1,3GT) can catalyze the synthesis of the ??Gal epitope. Therefore, the ?1,3GT gene, which encodes the ?1,3GT, is developed profoundly. The distributions of the ??Gal epitope and anti?Gal antibody, and the activation of ?1,3GT, reveal that the enzyme of ?1,3GT in ancestral primates is ineffective. Comparison of the nucleotide sequence of the human ?1,3?GT pseudogene to the corresponding different species sequence, and according to the evolutionary tree of different species, the results of evolutionary inactivation of the ?1,3GT gene in ancestral primates attribute to the mutations under a stronger selective pressure. However, on the basis of the structure, the mechanism and the specificity of the ??Gal epitope and anti?Gal antibody, they can be applied to clinical exploitation. Knocking out the ?1,3GT gene will eliminate the xenoantigen, Gal(?1,3)Gal, so that the transplantation of ?1,3GT gene knockout pig organ into human becomes a potential clinically acceptable treatment for solving the problem of organ shortage. By contrast, the ??Gal epitope expressed through the application of chemical, biochemical and genetic engineering can be exploited for the clinical use. Targeting anti?Gal?mediated autologous tumor vaccines, which express ??Gal epitope to antigen?presenting cells, would increase their immunogenicity and elicit an immune response, which will be potent enough to eradicate the residual tumor cells. For tumor vaccines, the way of increasing immunogenicity of certain viral vaccines, including flu vaccines and human immunodeficiency virus vaccines, can also be used in the elderly. Recently, ??Gal epitope nanoparticles have been applied to accelerate wound healing and further directions on regeneration of internally injured tissues.

Dataset Information

Synthesis of Gal?(1,3)Gal?(1,4)GlcNAc?-, Gal?(1,4)GlcNAc?- and GlcNAc-containing neoglycoproteins and their immunological evaluation in the context of Chagas disease.

Publications

Synthesis of Galα(1,3)Galβ(1,4)GlcNAcα-, Galβ(1,4)GlcNAcα- and GlcNAc-containing neoglycoproteins and their immunological evaluation in the context of Chagas disease.

Similar Datasets

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