Expression of disulphide-bridge-dependent conformational epitopes and immunogenicity of the carboxy-terminal 19 kDa domain of Plasmodium yoelii merozoite surface protein-1 in live attenuated Salmonella vaccine strains

Abstract

The 19 kDa carboxy-terminal domain of Plasmodium yoelii merozoite surface protein-I (MSPI,,) was expressed in Salmonella vaccine strains as a carboxyterminal fusion to fragment C of tetanus toxin (TetC). This study demonstrates that antibodies that recognize disulphide-dependent conformational epitopes in native MSPI react with the TetC-MSPI,, fusion protein expressed in Salmonella. The proper folding of MSPI,, polypeptide is dependent on both the Salmonella host strain and the protein to which the MSPI,, polypeptide is fused. Serum from mice immunized with Salmonella fyphimurium C5aroD expressing TetC-MSPI ,, recognized native MSPI as shown by immunofluorescence with P. yoelii-infected erythrocytes. Antibody levels to MSPI,, were highest in out-bred mice immunized with S-typhimurium C5aroD carrying pTECH2-MSPI 19 and antibody was mostly directed against reductionsensitive conformational epitopes. However, antibody levels were lower than in BALBlc mice immunized with a glutathione S-transferase (GST)-MSPI ,, fusion protein in Freund's adjuvant, and which were protected against P. yoelii challenge infection. In challenge experiments with P. yoelii the Salmonellaimmunized mice were not protected, probably reflecting the magnitude of the antibody response. The results of this study have important implications in the design of live multivalent bacterial vaccines against eukaryotic pathogens.

Acquired Immune Responses to Plasmodium falciparum Merozoite Surface Protein-1 in the Human Fetus

“…IgG Abs in mothers and their newborns recognized the Q-KNG as well as the E-KNG, Q-TSR, and E-TSR alleles, although Ab to the E-TSR variant was less than that observed to the other variants. This is consistent with the notion that Abs to MSP-1 19 bind to conformational epitopes conserved in the two epidermal growth factor motifs, i.e., binding is not constrained by changes in primary amino acid sequence that do not affect folding through disulfide bonds in cysteine residues (33,39,(62)(63)(64)(65). Similarly, cytokine responses in mothers and their newborns were stimulated equally well by each of the four constructs.…”

A chimeric protein-based malaria vaccine candidate induces robust T cell responses against Plasmodium vivax MSP119

“…To remove antibodies elicited against linear epitopes the samples tested in ELISA were first absorbed with reduced PvRMC-MSP1 at 2 μg/ml for 2 hours. For absorption under reducing conditions, PvRMC-MSP1 was initially treated with 0.05 M dithiothreitol (DTT) at 37 °C for 1 h. The reduced PvRMC-MSP1 was then diluted in 0.1 M carbonate buffer, containing 0.05 M DTT as described 93 and Immulon 2HB plates coated overnight. After 2 hours absorption, the samples were then tested for recognition of reduced or non-reduced PvRMC-MSP1 at 1 μg/ml.…”

Production and Preclinical Evaluation of Plasmodium falciparum MSP-1 19 and MSP-3 11 Chimeric Protein, PfMSP-Fu 24

“…linkages, which provide it with conformational stability (47,48). Maintaining proper conformation of PfMSP-1 19 is critical for the generation of protective antibodies.…”