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Crystal structure of Plasmodium knowlesi apical membrane antigen 1 and its complex with an invasion-inhibitory monoclonal antibody.

Vulliez-Le Normand B, Faber BW, Saul FA, van der Eijk M, Thomas AW, Singh B, Kocken CH, Bentley GA - PLoS ONE (2015)

Bottom Line: R31C2 inhibits binding of the Rhoptry Neck Protein 2 (RON2) receptor by steric blocking of the hydrophobic groove and by preventing the displacement of the D2 loop which is essential for exposing the complete binding site on AMA1.PkAMA1 is much less polymorphic than the P. falciparum and P. vivax orthologues.Unlike these two latter species, there are no polymorphic sites close to the RON2-binding site of PkAMA1, suggesting that P. knowlesi has not developed a mechanism of immune escape from the host's humoral response to AMA1.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur, Unité d'Immunologie Structurale, Département de Biologie Structurale et Chimie, Paris, France; CNRS URA 2185, Paris, France.

ABSTRACT
The malaria parasite Plasmodium knowlesi, previously associated only with infection of macaques, is now known to infect humans as well and has become a significant public health problem in Southeast Asia. This species should therefore be targeted in vaccine and therapeutic strategies against human malaria. Apical Membrane Antigen 1 (AMA1), which plays a role in Plasmodium merozoite invasion of the erythrocyte, is currently being pursued in human vaccine trials against P. falciparum. Recent vaccine trials in macaques using the P. knowlesi orthologue PkAMA1 have shown that it protects against infection by this parasite species and thus should be developed for human vaccination as well. Here, we present the crystal structure of Domains 1 and 2 of the PkAMA1 ectodomain, and of its complex with the invasion-inhibitory monoclonal antibody R31C2. The Domain 2 (D2) loop, which is displaced upon binding the Rhoptry Neck Protein 2 (RON2) receptor, makes significant contacts with the antibody. R31C2 inhibits binding of the Rhoptry Neck Protein 2 (RON2) receptor by steric blocking of the hydrophobic groove and by preventing the displacement of the D2 loop which is essential for exposing the complete binding site on AMA1. R31C2 recognizes a non-polymorphic epitope and should thus be cross-strain reactive. PkAMA1 is much less polymorphic than the P. falciparum and P. vivax orthologues. Unlike these two latter species, there are no polymorphic sites close to the RON2-binding site of PkAMA1, suggesting that P. knowlesi has not developed a mechanism of immune escape from the host's humoral response to AMA1.

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The PkAMA1 epitope recognized by R31C2.PkAMA1 is shown in surface representation with the epitope residues on Domain 1 in green and the epitope residues on the D2 loop in red. In addition, residues of both PkAMA1 and R31C2 that are involved in polar interactions are drawn in stick representation with hydrogen bonds indicated by dotted lines. The stick models of the PkAMA1 residues and their labels are green for Domain 1 and red for the D2 loop. Stick models for R31C2 are blue for VH residues and yellow for VL residues with residue labels in black. (The complete list of PkAMA1 and R31C2 residues in contact is given in Table 3, Table 4).
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pone.0123567.g004: The PkAMA1 epitope recognized by R31C2.PkAMA1 is shown in surface representation with the epitope residues on Domain 1 in green and the epitope residues on the D2 loop in red. In addition, residues of both PkAMA1 and R31C2 that are involved in polar interactions are drawn in stick representation with hydrogen bonds indicated by dotted lines. The stick models of the PkAMA1 residues and their labels are green for Domain 1 and red for the D2 loop. Stick models for R31C2 are blue for VH residues and yellow for VL residues with residue labels in black. (The complete list of PkAMA1 and R31C2 residues in contact is given in Table 3, Table 4).

Mentions: The PkAMA1-Fab R31C2 complex, which crystallized in the space group C2 with one molecule of complex in the asymmetric unit, was refined at 3.1 Å resolution (Table 1, Table 2). The main chain of PkAMA1 was traced from Met51 to Phe386, with gaps Glu301-Met302 and Phe330-Asn331 which occur in the D2 loop (Fig 2B). The heavy and light chains of the Fab fragment were both traced in their entirety to the C-terminal cysteine residues that form the interchain disulfide bridge. (Fig 3) The Fab fragment binds to the hydrophobic groove of PkAMA1 and makes significant contact with the D2 loop. A total of 2465 Å2 of solvent-accessible surface is buried at the antibody-antigen interface (1180 Å2 from the antibody; 1285 Å2 from the antigen). The heavy-chain variable domain (VH) contributes 838 Å2 to the buried surface while the light-chain variable domain (VL) contributes 342 Å2. All three Complementarity-Determining Regions (CDR) of both VH and VL make direct contacts with PkAMA1. The shape complementarity factor (shape correlation statistic, Sc) is 0.74, which is significantly above values typical of antibody-antigen complexes [47]. The more significant participation of VH is also reflected in the number and nature of the contacts across the interface; for VH there are 116 interatomic distances less than 3.8 Å, of which 22 are polar (including three salt bridges) while for VL there are only 25 contacts, of which two are polar (Table 3, Table 4, Fig 4). The D2 loop is contacted by CDR-H1, CDR-H2 and CDR-H3 (first, second and third CDR of VH, respectively), with 46 interatomic contacts < 3.8 Å.


Crystal structure of Plasmodium knowlesi apical membrane antigen 1 and its complex with an invasion-inhibitory monoclonal antibody.

Vulliez-Le Normand B, Faber BW, Saul FA, van der Eijk M, Thomas AW, Singh B, Kocken CH, Bentley GA - PLoS ONE (2015)

The PkAMA1 epitope recognized by R31C2.PkAMA1 is shown in surface representation with the epitope residues on Domain 1 in green and the epitope residues on the D2 loop in red. In addition, residues of both PkAMA1 and R31C2 that are involved in polar interactions are drawn in stick representation with hydrogen bonds indicated by dotted lines. The stick models of the PkAMA1 residues and their labels are green for Domain 1 and red for the D2 loop. Stick models for R31C2 are blue for VH residues and yellow for VL residues with residue labels in black. (The complete list of PkAMA1 and R31C2 residues in contact is given in Table 3, Table 4).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4401722&req=5

pone.0123567.g004: The PkAMA1 epitope recognized by R31C2.PkAMA1 is shown in surface representation with the epitope residues on Domain 1 in green and the epitope residues on the D2 loop in red. In addition, residues of both PkAMA1 and R31C2 that are involved in polar interactions are drawn in stick representation with hydrogen bonds indicated by dotted lines. The stick models of the PkAMA1 residues and their labels are green for Domain 1 and red for the D2 loop. Stick models for R31C2 are blue for VH residues and yellow for VL residues with residue labels in black. (The complete list of PkAMA1 and R31C2 residues in contact is given in Table 3, Table 4).
Mentions: The PkAMA1-Fab R31C2 complex, which crystallized in the space group C2 with one molecule of complex in the asymmetric unit, was refined at 3.1 Å resolution (Table 1, Table 2). The main chain of PkAMA1 was traced from Met51 to Phe386, with gaps Glu301-Met302 and Phe330-Asn331 which occur in the D2 loop (Fig 2B). The heavy and light chains of the Fab fragment were both traced in their entirety to the C-terminal cysteine residues that form the interchain disulfide bridge. (Fig 3) The Fab fragment binds to the hydrophobic groove of PkAMA1 and makes significant contact with the D2 loop. A total of 2465 Å2 of solvent-accessible surface is buried at the antibody-antigen interface (1180 Å2 from the antibody; 1285 Å2 from the antigen). The heavy-chain variable domain (VH) contributes 838 Å2 to the buried surface while the light-chain variable domain (VL) contributes 342 Å2. All three Complementarity-Determining Regions (CDR) of both VH and VL make direct contacts with PkAMA1. The shape complementarity factor (shape correlation statistic, Sc) is 0.74, which is significantly above values typical of antibody-antigen complexes [47]. The more significant participation of VH is also reflected in the number and nature of the contacts across the interface; for VH there are 116 interatomic distances less than 3.8 Å, of which 22 are polar (including three salt bridges) while for VL there are only 25 contacts, of which two are polar (Table 3, Table 4, Fig 4). The D2 loop is contacted by CDR-H1, CDR-H2 and CDR-H3 (first, second and third CDR of VH, respectively), with 46 interatomic contacts < 3.8 Å.

Bottom Line: R31C2 inhibits binding of the Rhoptry Neck Protein 2 (RON2) receptor by steric blocking of the hydrophobic groove and by preventing the displacement of the D2 loop which is essential for exposing the complete binding site on AMA1.PkAMA1 is much less polymorphic than the P. falciparum and P. vivax orthologues.Unlike these two latter species, there are no polymorphic sites close to the RON2-binding site of PkAMA1, suggesting that P. knowlesi has not developed a mechanism of immune escape from the host's humoral response to AMA1.

View Article: PubMed Central - PubMed

Affiliation: Institut Pasteur, Unité d'Immunologie Structurale, Département de Biologie Structurale et Chimie, Paris, France; CNRS URA 2185, Paris, France.

ABSTRACT
The malaria parasite Plasmodium knowlesi, previously associated only with infection of macaques, is now known to infect humans as well and has become a significant public health problem in Southeast Asia. This species should therefore be targeted in vaccine and therapeutic strategies against human malaria. Apical Membrane Antigen 1 (AMA1), which plays a role in Plasmodium merozoite invasion of the erythrocyte, is currently being pursued in human vaccine trials against P. falciparum. Recent vaccine trials in macaques using the P. knowlesi orthologue PkAMA1 have shown that it protects against infection by this parasite species and thus should be developed for human vaccination as well. Here, we present the crystal structure of Domains 1 and 2 of the PkAMA1 ectodomain, and of its complex with the invasion-inhibitory monoclonal antibody R31C2. The Domain 2 (D2) loop, which is displaced upon binding the Rhoptry Neck Protein 2 (RON2) receptor, makes significant contacts with the antibody. R31C2 inhibits binding of the Rhoptry Neck Protein 2 (RON2) receptor by steric blocking of the hydrophobic groove and by preventing the displacement of the D2 loop which is essential for exposing the complete binding site on AMA1. R31C2 recognizes a non-polymorphic epitope and should thus be cross-strain reactive. PkAMA1 is much less polymorphic than the P. falciparum and P. vivax orthologues. Unlike these two latter species, there are no polymorphic sites close to the RON2-binding site of PkAMA1, suggesting that P. knowlesi has not developed a mechanism of immune escape from the host's humoral response to AMA1.

Show MeSH
Related in: MedlinePlus