Multimeric bivalent immunogens from recombinant tetanus toxin HC fragment, synthetic hexasaccharides, and a glycopeptide adjuvant

Abstract

Using recombinant tetanus toxin H(C) fragment (rTT-H(C)) as carrier, we prepared multimeric bivalent immunogens featuring the synthetic hexasaccharide fragment of O-PS of Vibrio cholerae O:1, serotype Ogawa, in combination with either the synthetic hexasaccharide fragment of O-PS of Vibrio cholerae O:1, serotype Inaba, or a synthetic disaccharide tetrapeptide peptidoglycan fragment as adjuvant. The conjugation reaction was effected by squaric acid chemistry and monitored in virtually real time by SELDI-TOF MS. In this way, we could prepare well-defined immunogens with predictable carbohydrate-carrier ratio, whose molecular mass and the amount of each saccharide attached could be independently determined. The ability to prepare such neoglycoconjugates opens unprecedented possibilities for preparation of conjugate vaccines for bacterial diseases from synthetic carbohydrates.

Mapping the glycation sites in the neoglycoconjugate from hexasaccharide antigen of Vibrio cholerae, serotype Ogawa and the recombinant tetanus toxin C‐fragment carrier

“…[13] Briefly, rTT–Hc (26.8 mg, 1 eq) was dissolved in pH 9 borate buffer (775 µ1) to form a clear solution. Ogawa hexasaccharide squarate [4] (5.5 mg, 6 eq) was added, and the clear solution was stirred at room temperature for 24 h. The mixture was dialyzed (eight times) against 10 mM ammonium carbonate using Millipore Amicon Ultra centrifugal filter device (30 kDa cut-off, 4°C).…”

“…[4,8,9] Using this method, they synthesized different carbohydrate-protein neoglycoconjugates by attachment of fragments of the O-specific polysaccharide of Vibrio cholerae O1 serotype Ogawa [9,10] and lnaba [11,12] to bovine serum albumin (BSA), and also the Ogawa hexasaccharide to the medically acceptable carrier, rTT–Hc. [13] These hapten–protein glycoconjugates are not only immunogenic but also offer protective capacity. [14–16] …”

“…[17,18] The toxoid form of this protein has also been used as a carrier protein for antigens in experimental carbohydrate-protein conjugate vaccines. [13] However, because the structure of TT is not fully characterized, the reproducibility of the synthesis of a well-defined polysaccharide–TT neoglycoconjugate vaccine was found to be poor. Consequently, the use of a fully characterized Hc fragment for the preparation of such vaccines is generally preferred.…”

The Future of Synthetic Carbohydrate Vaccines: Immunological Studies on Streptococcus pneumoniae Type 14

“…The immunogenicity of the synthetic oligosaccharide fragment of the O-specific polysaccharide (O-PS) of Vibrio cholera O1, serotype Ogawa, conjugated to bovine serum albumin has been investigated in a mouse model [62,63]. A multimeric bivalent synthetic hexasaccharide fragment of the O-specific polysaccharide of Vibrio cholera O 1 , s e r o t y p e O g a w a , i n combination with Inaba:1 or a synthetic disaccharide tetrapeptide peptidoglycan fragment as adjuvant were prepared and conjugated to recombinant tetanus toxin H(C) fragment as protein carrier [64]. The immunogenicity of synthetic oligosaccharides mimicking the Oantigen of the Shigella flexneri 2a lipopolysaccharide (LPS) was also investigated in mice [65,66].…”

Binding activity and immunogenic characterization of recombinant C-terminal quarter and half of the heavy chain of botulinum neurotoxin serotype A

“…6,14,19,20 Especially, the Hc-C subdomains of BoNTs might be used as a carrier or component of a chimeric vaccine as fragments of TeNT. [21][22][23] Therefore, recombinant AHc-C as antigen or vectors for chimeric vaccines has been also explored in our laboratory. In the current study, the AHc-C was co-expressed in E. coli with Trx and its binding activity and immunogenic characterization were explored and compared with the AHc.…”