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Analyzing and identifying novel B cell epitopes within Toxoplasma gondii GRA4.

Wang Y, Wang G, Ou J, Yin H, Zhang D - Parasit Vectors (2014)

Bottom Line: The potential B cell epitopes of GRA4 predicted by bioinformatics tools focused on six regions of GRA4, 52-77 aa, 93-112 aa, 127-157 aa, 178-201 aa, 223-252 aa and 314-333 aa.Three of the eleven peptides (amino acids 62-77, 233-252 and 314-333) tested were recognized by all sera.The identified epitopes may be useful for additional studies of epitope-based vaccines and diagnostic reagents.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, 730046, China. wangyh061001@163.com.

ABSTRACT

Background: The identification of specific epitopes targeted by the host antibody response is important for understanding the natural response to infection and for the development of epitope-based marker vaccines and diagnostic tools for toxoplasmosis. In this study, Toxoplasma gondii GRA4 epitopes were identified using software-based prediction and a synthetic peptide technique.

Methods: The complete GRA4 gene sequence was obtained from T. gondii of the Gansu Jingtai strain of tachyzoites. The potential B cell epitopes of GRA4 was predicted using the PROTEAN subroutine in the DNASTAR software package. The peptides with good hydrophilicity, high accessibility, high flexibility and strong antigenicity were chemically synthesized and assessed by ELISA using pig sera from different time points after infection.

Results: The potential B cell epitopes of GRA4 predicted by bioinformatics tools focused on six regions of GRA4, 52-77 aa, 93-112 aa, 127-157 aa, 178-201 aa, 223-252 aa and 314-333 aa. Eleven shorter peptides from the six regions were synthesized and assessed by ELISA using pig sera from different time points after infection. Three of the eleven peptides (amino acids 62-77, 233-252 and 314-333) tested were recognized by all sera.

Conclusions: We precisely located the T. gondii GRA4 epitopes using pig sera collected at different time points after infection. The identified epitopes may be useful for additional studies of epitope-based vaccines and diagnostic reagents.

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Related in: MedlinePlus

ELISA of IgG antibodies against different peptides in the four groups of pig sera. (A), (B), (C) and (D) showing the absorbances targeting P2, P10 and P11, respectively. The cut-off point for the assay is indicated by the horizontal line.
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Fig3: ELISA of IgG antibodies against different peptides in the four groups of pig sera. (A), (B), (C) and (D) showing the absorbances targeting P2, P10 and P11, respectively. The cut-off point for the assay is indicated by the horizontal line.

Mentions: In the past, several experimental techniques were developed for mapping antibody interacting residues on an antigen, including the identification of interacting residues from the structure of antibody-antigen complexes [55,56]. One popular approach is the synthetic peptide technique, which primarily identifies sequential epitopes [56]. Using this technique, we verified the validity of the predicted epitopes in the present study. All eleven of the predicted epitope peptides were evaluated by ELISA using pig sera from various time points after infection. P2, P10 and P11 were recognized by all sera. The other eight peptides were recognized by select sera from various time points after infection (Figure 2; number of positive samples/tested for each peptide as follows: P1:7/51, P3:36/51, P4:23/51, P5:12/51, P6:17/51, P7:27/51, P8:18/51 and P9:21/51). The results of the ELISA for three peptides, P2, P10 and P11, are shown in Figure 3. For each of the three peptides, no significant differences were observed between the mean absorbances of the three groups (G1, G2 and G3) as determined by ANOVA. Furthermore, no significant differences were observed between the mean absorbances of the three peptides, P2, P10 and P11. In our study, we found that the three peptides derived from GRA4 were recognized by pig sera from different time points after infection. The reactivity of these epitopes does not seem to be dependent upon the time of infection, suggesting that the host response is constantly activated by these protein fragments.Figure 2


Analyzing and identifying novel B cell epitopes within Toxoplasma gondii GRA4.

Wang Y, Wang G, Ou J, Yin H, Zhang D - Parasit Vectors (2014)

ELISA of IgG antibodies against different peptides in the four groups of pig sera. (A), (B), (C) and (D) showing the absorbances targeting P2, P10 and P11, respectively. The cut-off point for the assay is indicated by the horizontal line.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4195951&req=5

Fig3: ELISA of IgG antibodies against different peptides in the four groups of pig sera. (A), (B), (C) and (D) showing the absorbances targeting P2, P10 and P11, respectively. The cut-off point for the assay is indicated by the horizontal line.
Mentions: In the past, several experimental techniques were developed for mapping antibody interacting residues on an antigen, including the identification of interacting residues from the structure of antibody-antigen complexes [55,56]. One popular approach is the synthetic peptide technique, which primarily identifies sequential epitopes [56]. Using this technique, we verified the validity of the predicted epitopes in the present study. All eleven of the predicted epitope peptides were evaluated by ELISA using pig sera from various time points after infection. P2, P10 and P11 were recognized by all sera. The other eight peptides were recognized by select sera from various time points after infection (Figure 2; number of positive samples/tested for each peptide as follows: P1:7/51, P3:36/51, P4:23/51, P5:12/51, P6:17/51, P7:27/51, P8:18/51 and P9:21/51). The results of the ELISA for three peptides, P2, P10 and P11, are shown in Figure 3. For each of the three peptides, no significant differences were observed between the mean absorbances of the three groups (G1, G2 and G3) as determined by ANOVA. Furthermore, no significant differences were observed between the mean absorbances of the three peptides, P2, P10 and P11. In our study, we found that the three peptides derived from GRA4 were recognized by pig sera from different time points after infection. The reactivity of these epitopes does not seem to be dependent upon the time of infection, suggesting that the host response is constantly activated by these protein fragments.Figure 2

Bottom Line: The potential B cell epitopes of GRA4 predicted by bioinformatics tools focused on six regions of GRA4, 52-77 aa, 93-112 aa, 127-157 aa, 178-201 aa, 223-252 aa and 314-333 aa.Three of the eleven peptides (amino acids 62-77, 233-252 and 314-333) tested were recognized by all sera.The identified epitopes may be useful for additional studies of epitope-based vaccines and diagnostic reagents.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, 730046, China. wangyh061001@163.com.

ABSTRACT

Background: The identification of specific epitopes targeted by the host antibody response is important for understanding the natural response to infection and for the development of epitope-based marker vaccines and diagnostic tools for toxoplasmosis. In this study, Toxoplasma gondii GRA4 epitopes were identified using software-based prediction and a synthetic peptide technique.

Methods: The complete GRA4 gene sequence was obtained from T. gondii of the Gansu Jingtai strain of tachyzoites. The potential B cell epitopes of GRA4 was predicted using the PROTEAN subroutine in the DNASTAR software package. The peptides with good hydrophilicity, high accessibility, high flexibility and strong antigenicity were chemically synthesized and assessed by ELISA using pig sera from different time points after infection.

Results: The potential B cell epitopes of GRA4 predicted by bioinformatics tools focused on six regions of GRA4, 52-77 aa, 93-112 aa, 127-157 aa, 178-201 aa, 223-252 aa and 314-333 aa. Eleven shorter peptides from the six regions were synthesized and assessed by ELISA using pig sera from different time points after infection. Three of the eleven peptides (amino acids 62-77, 233-252 and 314-333) tested were recognized by all sera.

Conclusions: We precisely located the T. gondii GRA4 epitopes using pig sera collected at different time points after infection. The identified epitopes may be useful for additional studies of epitope-based vaccines and diagnostic reagents.

Show MeSH
Related in: MedlinePlus