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Identification of an Immunogenic Mimic of a Conserved Epitope on the Plasmodium falciparum Blood Stage Antigen AMA1 Using Virus-Like Particle (VLP) Peptide Display.

Crossey E, Frietze K, Narum DL, Peabody DS, Chackerian B - PLoS ONE (2015)

Bottom Line: Most of the selected VLPs failed to reliably elicit AMA1 specific antibodies.However, one VLP consistently induced antibodies that cross-reacted with AMA1.Taken together, these data demonstrate that VLP-peptide display can identify immunogenic mimics of a complex conformational epitope and illustrate the promise and challenges of this approach.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology, University of New Mexico, Albuquerque, NM 87131, United States of America.

ABSTRACT
We have developed a peptide display platform based on VLPs of the RNA bacteriophage MS2 that combines the high immunogenicity of VLP display with affinity selection capabilities. Random peptides can be displayed on the VLP surface by genetically inserting sequences into a surface-exposed loop of the viral coat protein. VLP-displayed peptides can then be isolated by selection using antibodies, and the VLP selectants can then be used directly as immunogens. Here, we investigated the ability of this platform to identify mimotopes of a highly conserved conformational epitope present on the Plasmodium falciparum blood-stage protein AMA1. Using 4G2, a monoclonal antibody that binds to this epitope and is a potent inhibitor of erythrocyte invasion, we screened three different VLP-peptide libraries and identified specific VLPs that bound strongly to the selecting mAb. We then tested the ability of a handful of selected VLPs to elicit anti-AMA1 antibody responses in mice. Most of the selected VLPs failed to reliably elicit AMA1 specific antibodies. However, one VLP consistently induced antibodies that cross-reacted with AMA1. Surprisingly, this VLP bound to 4G2 more weakly than the other selectants we identified. Taken together, these data demonstrate that VLP-peptide display can identify immunogenic mimics of a complex conformational epitope and illustrate the promise and challenges of this approach.

No MeSH data available.


Related in: MedlinePlus

Comparison of the structure of the 4G2 epitope with the putative structure of the 4G2 mimotope on MS2 coat protein.(A) Predicted structure of the MS2-VTHDAWRPD coat protein dimer as determined by the One-to-One Threading function in the Web-based Phyre2 Server by modeling on MS2 coat protein (PDB: 1MSC). The VTHDAWRPD peptide, highlighted in panels B-D, is indicated by a dotted circle. Charge of proteins indicated by red (negative), blue (positive), and white (neutral). (B-D) A magnified view of the predicted structure of the VTHDAWRPD in the context of the MS2 AB-loop. (E-G) The structure of domains I and II of PfAMA1 (PDB: 1Z40_A). In panels (A,D) the charge of proteins indicated by red (negative), blue (positive), and white (neutral). Panel C highlights the VTHDAWRPD peptide insert and panel F shows a subset of the residues identified as necessary for 4G2 binding; both are indicated in magenta. In yellow space-fill, panel G shows selected residues on AMA1 involved in 4G2 binding (aa 351–353) and panel D shows putative residues that may be involved in 4G2 binding to VTHDAWRPD.
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pone.0132560.g008: Comparison of the structure of the 4G2 epitope with the putative structure of the 4G2 mimotope on MS2 coat protein.(A) Predicted structure of the MS2-VTHDAWRPD coat protein dimer as determined by the One-to-One Threading function in the Web-based Phyre2 Server by modeling on MS2 coat protein (PDB: 1MSC). The VTHDAWRPD peptide, highlighted in panels B-D, is indicated by a dotted circle. Charge of proteins indicated by red (negative), blue (positive), and white (neutral). (B-D) A magnified view of the predicted structure of the VTHDAWRPD in the context of the MS2 AB-loop. (E-G) The structure of domains I and II of PfAMA1 (PDB: 1Z40_A). In panels (A,D) the charge of proteins indicated by red (negative), blue (positive), and white (neutral). Panel C highlights the VTHDAWRPD peptide insert and panel F shows a subset of the residues identified as necessary for 4G2 binding; both are indicated in magenta. In yellow space-fill, panel G shows selected residues on AMA1 involved in 4G2 binding (aa 351–353) and panel D shows putative residues that may be involved in 4G2 binding to VTHDAWRPD.

Mentions: In order to investigate the structural basis of 4G2 mimics further, we generated predicted structural models of the selectant VLPs by using the Phyre2 Server. This web-based application can model protein structures based on sequence homology and predict structural features of user-entered protein sequences. For our purpose, we modeled the MS2 coat protein amino acid sequence with the foreign peptides against the structure for wild-type MS2 coat protein by using the One-to-One Threading function. This function allows modeling of the short constrained peptides in the AB-loop on the known structure of the MS2 coat protein dimer. Interestingly, the best immunologic mimic (VTHDAWRPD) had a striking similarity to a prominent feature of the AMA1 4G2 binding site (Fig 8), including a negatively charged “knuckle” (shown in red) and a positively charged “thumb” (shown in blue). Selectants that showed weak or no immunologic mimicry of the 4G2 epitope did not exhibit these features, although in some cases either a “knuckle” or a “thumb” feature was present (S1 Fig). These data suggest a possible explanation for the immunologic mimicry demonstrated by a subset of selectant VLPs.


Identification of an Immunogenic Mimic of a Conserved Epitope on the Plasmodium falciparum Blood Stage Antigen AMA1 Using Virus-Like Particle (VLP) Peptide Display.

Crossey E, Frietze K, Narum DL, Peabody DS, Chackerian B - PLoS ONE (2015)

Comparison of the structure of the 4G2 epitope with the putative structure of the 4G2 mimotope on MS2 coat protein.(A) Predicted structure of the MS2-VTHDAWRPD coat protein dimer as determined by the One-to-One Threading function in the Web-based Phyre2 Server by modeling on MS2 coat protein (PDB: 1MSC). The VTHDAWRPD peptide, highlighted in panels B-D, is indicated by a dotted circle. Charge of proteins indicated by red (negative), blue (positive), and white (neutral). (B-D) A magnified view of the predicted structure of the VTHDAWRPD in the context of the MS2 AB-loop. (E-G) The structure of domains I and II of PfAMA1 (PDB: 1Z40_A). In panels (A,D) the charge of proteins indicated by red (negative), blue (positive), and white (neutral). Panel C highlights the VTHDAWRPD peptide insert and panel F shows a subset of the residues identified as necessary for 4G2 binding; both are indicated in magenta. In yellow space-fill, panel G shows selected residues on AMA1 involved in 4G2 binding (aa 351–353) and panel D shows putative residues that may be involved in 4G2 binding to VTHDAWRPD.
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pone.0132560.g008: Comparison of the structure of the 4G2 epitope with the putative structure of the 4G2 mimotope on MS2 coat protein.(A) Predicted structure of the MS2-VTHDAWRPD coat protein dimer as determined by the One-to-One Threading function in the Web-based Phyre2 Server by modeling on MS2 coat protein (PDB: 1MSC). The VTHDAWRPD peptide, highlighted in panels B-D, is indicated by a dotted circle. Charge of proteins indicated by red (negative), blue (positive), and white (neutral). (B-D) A magnified view of the predicted structure of the VTHDAWRPD in the context of the MS2 AB-loop. (E-G) The structure of domains I and II of PfAMA1 (PDB: 1Z40_A). In panels (A,D) the charge of proteins indicated by red (negative), blue (positive), and white (neutral). Panel C highlights the VTHDAWRPD peptide insert and panel F shows a subset of the residues identified as necessary for 4G2 binding; both are indicated in magenta. In yellow space-fill, panel G shows selected residues on AMA1 involved in 4G2 binding (aa 351–353) and panel D shows putative residues that may be involved in 4G2 binding to VTHDAWRPD.
Mentions: In order to investigate the structural basis of 4G2 mimics further, we generated predicted structural models of the selectant VLPs by using the Phyre2 Server. This web-based application can model protein structures based on sequence homology and predict structural features of user-entered protein sequences. For our purpose, we modeled the MS2 coat protein amino acid sequence with the foreign peptides against the structure for wild-type MS2 coat protein by using the One-to-One Threading function. This function allows modeling of the short constrained peptides in the AB-loop on the known structure of the MS2 coat protein dimer. Interestingly, the best immunologic mimic (VTHDAWRPD) had a striking similarity to a prominent feature of the AMA1 4G2 binding site (Fig 8), including a negatively charged “knuckle” (shown in red) and a positively charged “thumb” (shown in blue). Selectants that showed weak or no immunologic mimicry of the 4G2 epitope did not exhibit these features, although in some cases either a “knuckle” or a “thumb” feature was present (S1 Fig). These data suggest a possible explanation for the immunologic mimicry demonstrated by a subset of selectant VLPs.

Bottom Line: Most of the selected VLPs failed to reliably elicit AMA1 specific antibodies.However, one VLP consistently induced antibodies that cross-reacted with AMA1.Taken together, these data demonstrate that VLP-peptide display can identify immunogenic mimics of a complex conformational epitope and illustrate the promise and challenges of this approach.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology, University of New Mexico, Albuquerque, NM 87131, United States of America.

ABSTRACT
We have developed a peptide display platform based on VLPs of the RNA bacteriophage MS2 that combines the high immunogenicity of VLP display with affinity selection capabilities. Random peptides can be displayed on the VLP surface by genetically inserting sequences into a surface-exposed loop of the viral coat protein. VLP-displayed peptides can then be isolated by selection using antibodies, and the VLP selectants can then be used directly as immunogens. Here, we investigated the ability of this platform to identify mimotopes of a highly conserved conformational epitope present on the Plasmodium falciparum blood-stage protein AMA1. Using 4G2, a monoclonal antibody that binds to this epitope and is a potent inhibitor of erythrocyte invasion, we screened three different VLP-peptide libraries and identified specific VLPs that bound strongly to the selecting mAb. We then tested the ability of a handful of selected VLPs to elicit anti-AMA1 antibody responses in mice. Most of the selected VLPs failed to reliably elicit AMA1 specific antibodies. However, one VLP consistently induced antibodies that cross-reacted with AMA1. Surprisingly, this VLP bound to 4G2 more weakly than the other selectants we identified. Taken together, these data demonstrate that VLP-peptide display can identify immunogenic mimics of a complex conformational epitope and illustrate the promise and challenges of this approach.

No MeSH data available.


Related in: MedlinePlus