ABSTRACT
Tetanus toxoid (TTd) plays an important role in the pharmaceutical world, especially in vaccines. The toxoid is obtained after formaldehyde treatment of the tetanus toxin. In parallel, current emphasis in the drug discovery field is put on producing well-defined and safer drugs, explaining the interest in finding new alternative proteins. The tetanus toxin fragment C (TTFC) has been extensively studied both as a neuroprotective agent for central nervous system disorders owing to its neuronal properties and as a carrier protein in vaccines. Indeed, it is derived from a part of the tetanus toxin and, as such, retains its immunogenic properties without being toxic. Moreover, this fragment has been well characterized, and its entire structure is known. Here, we propose a systematic review of TTFC by providing information about its structural features, its properties and its methods of production. We also describe the large uses of TTFC in the field of drug discovery. TTFC can therefore be considered as an attractive alternative to TTd and remarkably offers a wide range of uses, including as a carrier, delivery vector, conjugate, booster, inducer, and neuroprotector.
ABSTRACT
Aldehyde detoxification is a process used to convert toxin into toxoid for vaccine applications. In the case of tetanus toxin (TT), formaldehyde is used to obtain the tetanus toxoid (TTd), which is used either for the tetanus vaccine or as carrier protein in conjugate vaccines. Several studies have already been conducted to better understand the exact mechanism of this detoxification. Those studies led to the identification of a number of formaldehyde-induced modifications on lab scale TTd samples. To obtain greater insights of the changes induced by formaldehyde, we used three industrial TTd batches to identify repeatable modifications in the detoxification process. Our strategy was to combine seven analytical tools to map these changes. Mass spectrometry (MS), colorimetric test and amino acid analysis (AAA) were used to study modifications on amino acids. SDS-PAGE, asymmetric flow field flow fractionation (AF4), fluorescence spectroscopy and circular dichroism (CD) were used to study formaldehyde modifications on the whole protein structure. We identified 41 formaldehyde-induced modifications across the 1315 amino acid primary sequence of TT. Of these, five modifications on lysine residues were repeatable across TTd batches. Changes in protein conformation were also observed using SDS-PAGE, AF4 and CD techniques. Each analytical tool brought a piece of information regarding formaldehyde induced-modifications, and all together, these methods provided a comprehensive overview of the structural changes that occurred with detoxification. These results could be the first step leading to site-directed TT mutagenesis studies that may enable the production of a non-toxic equivalent protein without using formaldehyde.
