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Architecture Effects on the Binding of Cholera Toxin by Helical Glycopolypeptides.

ABSTRACT: A variety of binding events in biological systems are mediated by multivalent interactions between oligosaccharides and saccharide receptors present on pathogens and cell surfaces. In particular, given the important role of multivalent interaction between proteins and carbohydrates in the initial step of pathogen recognition, many glycosylated molecules and polymers have been synthesized in order to mimic the carbohydrate ligands and to inhibit the binding of the pathogen to its target. In this work, we extend our evaluation of the impact of the architecture of well-defined glycopolypeptides on the inhibition of binding of the cholera toxin B pentamer (CT B(5)) subunit. Here we report the production of two families of ?-helical glycopolypeptides which were synthesized via a combination of protein engineering and chemical methods. The presentation of pendant saccharides on the polypeptide backbones, as well as their valencies, can be well controlled via these methods. Control of the backbone conformation, introduced in this report, is also possible via these strategies. The polypeptides and glycopolypeptides were characterized via SDS-PAGE analysis, (1)H NMR, and MALDI-TOF mass spectrometry. Their conformation and hydrodynamic volume were characterized by circular dichroic (CD) spectroscopy and gel permeation chromatography (GPC), respectively. The binding of CT B(5) by these glycopolypeptides was evaluated via direct enzyme-linked immunosorbent assay (DELA). The effects of spacing and conformation were elucidated by comparison of the binding exhibited by helical glycopolypeptides with that of random-coil glycopolypeptides.

SUBMITTER: Liu S

PROVIDER: S-EPMC2639716 | biostudies-literature | 2008 Feb

REPOSITORIES: biostudies-literature

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Architecture Effects on the Binding of Cholera Toxin by Helical Glycopolypeptides.

Liu Shuang S Kiick Kristi L KL

Macromolecules 20080201 3

A variety of binding events in biological systems are mediated by multivalent interactions between oligosaccharides and saccharide receptors present on pathogens and cell surfaces. In particular, given the important role of multivalent interaction between proteins and carbohydrates in the initial step of pathogen recognition, many glycosylated molecules and polymers have been synthesized in order to mimic the carbohydrate ligands and to inhibit the binding of the pathogen to its target. In this ...[more]

PMID: 19214239

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Project description:The calcium-binding protein S100A4 has been described to promote pathological inflammation in experimental autoimmune and inflammatory disorders and in allergy and to contribute to antigen presentation and antibody response after parenteral immunization with an alum-adjuvanted antigen. In this study, we extend these findings by demonstrating that mice lacking S100A4 have a defective humoral and cellular immune response to mucosal (sublingual) immunization with a model protein antigen [ovalbumin (OVA)] given together with the strong mucosal adjuvant cholera toxin (CT), and that this impairment is due to defective adjuvant-stimulated antigen presentation by antigen-presenting cells. In comparison to wild-type (WT) mice, mice genetically lacking S100A4 had reduced humoral and cellular immune responses after immunization with OVA plus CT, including a complete lack of detectable germinal center reaction. Further, when stimulated in vitro with OVA plus CT, S100A4-/- dendritic cells (DCs) showed impaired responses in several CT-stimulated immune regulatory molecules including the co-stimulatory molecule CD86, inflammasome-associated caspase-1 and IL-1β. Coculture of OVA-specific OT-II T cells with S100A4-/- DCs that had been pulse incubated with OVA plus CT resulted in impaired OT-II T cell proliferation and reduced production of Th1, Th2, and Th17 cytokines compared to similar cocultures with WT DCs. In accordance with these findings, transfection of WT DCs with S100A4-targeting small interfering RNA (siRNA) but not mock-siRNA resulted in significant reductions in the expression of caspase-1 and IL-1β as well as CD86 in response to CT. Importantly, also engraftment of WT DCs into S100A4-/- mice effectively restored the immune response to immunization in the recipients. In conclusion, our results demonstrate that deficiency in S100A4 has a strong impact on the development of both humoral and cellular immunity after mucosal immunization using CT as adjuvant.

Dataset Information

Architecture Effects on the Binding of Cholera Toxin by Helical Glycopolypeptides.

Publications

Architecture Effects on the Binding of Cholera Toxin by Helical Glycopolypeptides.

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