Synthesis of the O-linked hexasaccharide containing β-d-Galp-(1→2)-d-Galf in Trypanosoma cruzi mucins. Differences on sialylation by trans-sialidase of the two constituent hexasaccharides

Abstract

The hexasaccharide β-D-Galp-(1→2)-[β-D-Galp-(1→3)]-β-D-Galp-(1→6)-[β-D-Galp(1→2)-β-D-Galf(1→4)]-D-GlcNAc (10) and its β-D-Galf-(1→2)-β-D-Galf containing isomer (7) are the largest carbohydrates in mucins of some strains of Trypanosoma cruzi. The terminal β-D-Galp units are sites of sialylation by the parasite trans-sialidase. Hexasaccharide 10 was chemically synthesized for the first time by a [3+3] nitrilium based convergent approach, using the trichloroacetimidate method of glycosylation. The (1)H NMR spectrum of its alditol was identical to the spectrum of the product released by β-elimination from the parasite mucin. The trans-sialylation reaction studied on the benzyl glycoside of 10 showed two monosialylated products whose relative abundance changed with time. On the other hand, only one product was produced by sialylation of the benzyl glycoside of 7. A preparative synthesis of the latter and spectroscopic analysis of the product unequivocally established the sialylation site at the less hindered (1→3)-linked galactopyranose.

Mucins and Pathogenic Mucin-Like Molecules Are Immunomodulators During Infection and Targets for Diagnostics and Vaccines

“…Initial synthesis includes the preparation of the O-linked saccharides 1 – 5 (Figure 4) present in T. cruzi Colombiana and Tulahuen strains (de Lederkremer and Agusti, 2009). The first synthetic target was disaccharide 1 (Gallo-Rodriguez et al, 1996), which is the basis of synthesizing other molecules including trisaccharides 2 (Gallo-Rodriguez et al, 1998), 3 and 4 (Mendoza et al, 2010), tetrasaccharide 5 (Gallo-Rodriguez et al, 2003), pentasaccharide 6 (Mendoza et al, 2006), and hexasaccharide 7 (Agusti et al, 2015). Further reports include the synthesis of glycan 8 from the T. cru zi Y strain (Figure 4) (van Well et al, 2008).…”

trans-Sialylation: a strategy used to incorporate sialic acid into oligosaccharides

“…In the case of sialylation of the benzyl glycoside of the other hexasaccharide, with three terminal Galp units, two monosialylated compounds and a minor amount of a disialylated product were formed (Fig. 7) (132). A study on the comparative rates of sialylation of the synthetic oligosaccharides, showed that the presence of Galf did not impair the reaction.…”

“…Colostrum, secreted by the mammary gland a few days before and after parturition, is a good source of oligosaccharides (128). Sialyllactose may be obtained from bovine colostrum in about 500 mg/L (129) and has been used as donor for the synthesis of complex sialylated oligosaccharides (130)(131)(132)(133)(134)(135)(136). Since sialyl galactooligosaccharides (SiaGOS) are much less abundant in bovine milk there are several reports describing their enzymatic synthesis with the aim to enrich baby food.…”

“…Since both constituents, βGalf and βGalp, may coexist in the same molecule, it was interesting to study their behavior in the TS reaction. The Galf-containing oligosaccharides have been chemically synthesized (132,133,(147)(148)(149)(150)(151). with TcTS showed selective trans-sialylation from the donor 3′-sialyllactose to the less hindered (1-3)-linked βGalp.…”

“…Sialylation of the more flexible alditol 2 was not selective and a mixture of compounds 3 and 4 was obtained (Scheme 6), suggesting that the open zig-zag conformation adopted by the alditol turned both galactoses almost equally accessible for TcTS recognition (130). Accordingly, the benzyl glycosides of the pentasaccharide and one of the hexasaccharides of the mucins, compounds 5 and 7 respectively, were also selectively sialylated in the same residue to give 6 and 8, respectively (Scheme 7) (132,133). All the structures were confirmed by NMR spectroscopy.…”