Immunological Evidence for Functional Rather than Structural Mimicry by aShigella flexneriY Polysaccharide-Mimetic Peptide

Abstract

Shigellosis, caused by Shigella species (gram negative), is a prominent, and the most infectious, diarrheal disease. Shigella flexneri, the species responsible for the highest mortality rate, is endemic in most developing countries (23). The 13 serotypes of S. flexneri, with the exception of serotype 6, result from structural modifications of the O-antigen polysaccharide (PS), the outer portion of the lipopolysaccharide (LPS) on the bacterial surface, which is both an essential virulence factor and a protective antigen (10). The basic O-antigen PS repeat, common to the 12 serotypes, is referred to as serotype Y. A murine immunoglobulin G3 (IgG3) monoclonal antibody, SYA/J6, specific for the PS O antigen of the S. flexneri Y LPS, was developed by Bundle (7) and Carlin and coworkers (8). The O-antigen Y PS is a linear heteropolymer with a tetrasaccha- -(13] and is recognized by the monoclonal antibody SYA/J6 (7, 9). The oligosaccharides of S. flexneri serotype Y have been well studied by nuclear magnetic resonance techniques, which have provided a three-dimensional model of the determinant in solution (4,30,31,32) and the identity of the biological repeating unit (9).Glycoconjugate vaccines to prevent shigellosis, focused on oligosaccharide analogues related to Shigella flexneri 2a and other serotypes, have been synthesized and evaluated (3,10,29,38). An interesting approach to vaccine design is the use of molecules that mimic the immunogenic element of interest (11,20,28). Carbohydrate-mimetic peptides have potential as surrogate ligands for traditional carbohydrate vaccines, providing more discriminating immune responses (19,20,28). However, there are few examples of immunological responses with peptide-based PS mimics (18,20,28). Therefore, the requirements for cross-reactivity are not fully understood and are certainly not predictable, because of the limited data set available.In order to further exploit this principle, a carbohydratemimetic peptide of the S. flexneri Y O-antigen PS, MDWNM HAA, cross-reactive with the anti-S. flexneri Y O-PS monoclonal antibody, SYA/J6, was identified by phage library screening (15). The structures of complexes of the antibody SYA/J6 Fab fragment with synthetic deoxytrisaccharide and pentasaccharide ligands, related to the S. flexneri Y O antigen, and with the carbohydrate-mimetic peptide have been determined by X-ray crystallography (17,33,34,35). The structure of the Fab complex with MDWNMHAA revealed differences, and few similarities, with respect to the oligosaccharide complexes (34), providing the first evidence that the modes of binding of the pentasaccharide and octapeptide differ considerably and that few aspects of structural mimicry exist (34).

Design, synthesis, and immunochemical characterization of a chimeric glycopeptide corresponding to the Shigella flexneri Y O-polysaccharide and its peptide mimic MDWNMHAA

“…23 In order to test these hypotheses, we synthesized protein conjugates of the octapeptide 24 and recently evaluated their immunogenicities. 16 Indeed, we found that, although the kinetics of the immune response in mice were slow, cross-reactivity was observed, and an effective prime-boost strategy was developed. 16 We now report the design of two chimeric glycopeptides ( Fig.…”

“…[14][15][16][17] In addition to their ability to elicit carbohydrate-binding antibody responses, these peptide mimics should focus the immune responses on particular epitopes since greater discrimination of the peptides for the corresponding antibody-combining sites has been observed, 18 thus minimizing autoimmune reactions with self structures where the original carbohydrate antigens would pose a threat as immunogens. [17][18][19] Shigella flexneri Y is a virulent bacterium that causes bacillary dysentery by invading the colonic mucosa.…”

Investigation of the Binding of a Carbohydrate‐Mimetic Peptide to its Complementary Anticarbohydrate Antibody by STD‐NMR Spectroscopy and Molecular‐Dynamics Simulations

“…It is noteworthy that despite the differences in peptide versus oligosaccharide binding, a protein conjugate of the peptide ligand is immunogenic and elicits an immune response that is cross-reactive with the bacterial polysaccharide. [6] Thus, the peptide is an antigenic mimic of the polysaccharide.…”

“…[3] Traditionally, carbohydrate-based vaccines are weakly immunogenic, as determined by the thymus dependent immune response; therefore, studies were undertaken to identify carbohydrate-mimetic peptides as surrogate ligands. [4,5] Using phase display, a weakly immunogenic [6] carbohydrate-mimetic peptide of the Shigella flexneri Y O-linked polysaccharide, with the amino acid sequence MDWNMHAA 1 (Figure 2), was identified to be cross-reacAbstract: Saturation transfer difference (STD)-NMR spectroscopy was used to probe experimentally the bioactive solution conformation of the carbohydrate mimic MDWNMHAA 1 of the O-polysaccharide of Shigella flexneri Y when bound to its complementary antibody, mAb SYA/J6. Molecular dynamics simulations using the ZymeCAD Molecular Dynamics platform were also undertaken to give a more accurate picture of the conformational flexibility and the possibilities for bound ligand conformations.…”

Developing strategies to enhance and focus humoral immune responses using filamentous phage as a model antigen

“…29 In support of this, others have found synthetic MD10 peptide conjugated to other carrier proteins to be very poorly immunogenic. 30 Moreover, while recombinant peptides displayed on pVIII can be resolved by NMR, 31 it is likely the SPDP amide and disulfide linkages introduce a degree of conformational flexibility to a conjugated peptide. The 4E10 L peptide, on the other hand, is extremely hydrophobic, and in recombinant form may embed itself in hydrophobic "grooves" between pVIII monomers, 32 making it poorly immunogenic as a recombinant fusion.…”