Giorgi M Eugenia, Lopez Rosana, Agusti Rosalia, Marino Carla, de Lederkremer Rosa M
Universidad de Buenos Aires, Consejo Nacional de Investigaciones Científicas y Técnicas, Centro de Investigaciones en Hidratos de Carbono (CIHIDECAR), Facultad de Ciencias Exactas y Naturales, Departamento de Química Orgánica, Buenos Aires, Argentina.
Universidad de Buenos Aires, Consejo Nacional de Investigaciones Científicas y Técnicas, Centro de Investigaciones en Hidratos de Carbono (CIHIDECAR), Facultad de Ciencias Exactas y Naturales, Departamento de Química Orgánica, Buenos Aires, Argentina.
Carbohydr Res. 2017 Oct 10;450:30-37. doi: 10.1016/j.carres.2017.08.007. Epub 2017 Aug 18.
Trypanosoma cruzi, the etiologic agent of Chagas disease, is covered by a dense glycocalix mainly composed by glycoproteins called mucins which are also the acceptors of sialic acid in a reaction catalyzed by a trans-sialidase (TcTS). Sialylation of trypomastigote mucins protects the parasite from lysis by the anti α-Galp antibodies from serum. The TcTS is essential for the infection process since T. cruzi is unable to biosynthesize sialic acid. The enzyme specifically transfers it from a terminal β-d-Galp unit in the host glycoconjugate to terminal β-d-Galp units in the parasite mucins to construct the d-NeuNAc(α2→3)β-d-Galp motif. On the other hand, although galactose is the most abundant sugar in mucins of both, the infective trypomastigotes and the insect stage epimastigotes, α-d-Galp is only present in the infective stage whereas β-d-Galf is characteristic of the epimastigote stage of the less virulent strains. Neither α-d-Galp nor d-Galf is acceptor of sialic acid. In the mucins, some of the oligosaccharides are branched with terminal β-d-Galp units to be able to accept sialic acid in the TcTS reaction. Based on previous reports showing that anti α-Galp antibodies only partially colocalize with sialic acid, we have undertaken the synthesis of the trisaccharide α-d-Galp(1→3)-[β-d-Galp(1→6)]-d-Galp, the smallest structure containing both, the antigenic d-Galp(α1→3)-d-Galp unit and the sialic acid-acceptor β-d-Galp unit. The trisaccharide was obtained as the 6-aminohexyl glycoside to facilitate further conjugation for biochemical studies. The synthetic approach involved the α-galactosylation at O-4 of a suitable precursor of the reducing end, followed by β-galactosylation at O-6 of the same precursor and introduction of the 6-aminohexyl aglycone. The fully deprotected trisaccharide was successfully sialylated by TcTS using either 3'-sialyllactose or fetuin as donors. The product, 6-aminohexyl α-d-NeuNAc(2→3)-β-d-Galp(1→6)-[α-d-Galp(1→3)]-β-d-Galp, was purified and characterized.
克氏锥虫是恰加斯病的病原体，其表面覆盖着一层致密的糖萼，主要由称为粘蛋白的糖蛋白组成，这些粘蛋白也是转唾液酸酶（TcTS）催化反应中唾液酸的受体。锥鞭毛体粘蛋白的唾液酸化可保护寄生虫免受血清中抗α-Galp抗体的裂解作用。由于克氏锥虫无法生物合成唾液酸，因此TcTS对感染过程至关重要。该酶将唾液酸从宿主糖缀合物中的末端β-d-Galp单元特异性转移至寄生虫粘蛋白中的末端β-d-Galp单元，以构建d-NeuNAc(α2→3)β-d-Galp基序。另一方面，尽管半乳糖是感染性锥鞭毛体和昆虫阶段上鞭毛体粘蛋白中最丰富的糖类，但α-d-Galp仅存在于感染阶段，而β-d-Galf是毒性较低菌株上鞭毛体阶段的特征性糖类。α-d-Galp和d-Galf都不是唾液酸的受体。在粘蛋白中，一些寡糖带有末端β-d-Galp单元分支，以便能够在TcTS反应中接受唾液酸。基于先前的报道显示抗α-Galp抗体仅与唾液酸部分共定位，我们合成了三糖α-d-Galp(1→3)-[β-d-Galp(1→6)]-d-Galp，这是包含抗原性d-Galp(α1→3)-d-Galp单元和唾液酸受体β-d-Galp单元的最小结构。该三糖以6-氨基己基糖苷形式获得，便于进一步偶联用于生化研究。合成方法包括在还原端的合适前体的O-4位进行α-半乳糖基化，然后在同一前体的O-6位进行β-半乳糖基化，并引入6-氨基己基糖苷配基。使用3'-唾液酸乳糖或胎球蛋白作为供体，完全脱保护的三糖成功地被TcTS唾液酸化。产物6-氨基己基α-d-NeuNAc(2→3)-β-d-Galp(1→6)-[α-d-Galp(1→3)]-β-d-Galp被纯化并进行了表征。
