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Diversity of T cell epitopes in Plasmodium falciparum circumsporozoite protein likely due to protein-protein interactions.

Aragam NR, Thayer KM, Nge N, Hoffman I, Martinson F, Kamwendo D, Lin FC, Sutherland C, Bailey JA, Juliano JJ - PLoS ONE (2013)

Bottom Line: Using the information from these parasite populations, structural analysis reveals that polymorphic amino acids within TH2 and TH3 colocalize to one side of the protein, surround, but do not involve, the hydrophobic pocket in CS, and predominately involve charge switches.In addition, free energy analysis shows polymorphic residues tend to be populated by energetically unfavorable amino acids.In combination, these findings suggest the diversity of T cell epitopes in CS may be primarily an evolutionary response to intermolecular interactions at the surface of the protein potentially counteracting antibody-mediated immune recognition or evolving host receptor diversity.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, University of North Carolina School of Medicine, Chapel Hill, North Carolina, United States of America.

ABSTRACT
Circumsporozoite protein (CS) is a leading vaccine antigen for falciparum malaria, but is highly polymorphic in natural parasite populations. The factors driving this diversity are unclear, but non-random assortment of the T cell epitopes TH2 and TH3 has been observed in a Kenyan parasite population. The recent publication of the crystal structure of the variable C terminal region of the protein allows the assessment of the impact of diversity on protein structure and T cell epitope assortment. Using data from the Gambia (55 isolates) and Malawi (235 isolates), we evaluated the patterns of diversity within and between epitopes in these two distantly-separated populations. Only non-synonymous mutations were observed with the vast majority in both populations at similar frequencies suggesting strong selection on this region. A non-random pattern of T cell epitope assortment was seen in Malawi and in the Gambia, but structural analysis indicates no intramolecular spatial interactions. Using the information from these parasite populations, structural analysis reveals that polymorphic amino acids within TH2 and TH3 colocalize to one side of the protein, surround, but do not involve, the hydrophobic pocket in CS, and predominately involve charge switches. In addition, free energy analysis suggests residues forming and behind the novel pocket within CS are tightly constrained and well conserved in all alleles. In addition, free energy analysis shows polymorphic residues tend to be populated by energetically unfavorable amino acids. In combination, these findings suggest the diversity of T cell epitopes in CS may be primarily an evolutionary response to intermolecular interactions at the surface of the protein potentially counteracting antibody-mediated immune recognition or evolving host receptor diversity.

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Related in: MedlinePlus

Significantly energetically-constrained amino acid positions identified by MUMBO Analysis.The 5 amino acids identified by MUMBO as having constrained ΔΔG mutational profiles relative to other identical amino acids within the crystal structure are color coded: ASN340 (red), Gly341 (orange) Ile342 (yellow) Glu343 (green), and Ser332 (cyan), are shown with respect to the TH2 and TH3 domains’ surface area (colored as in Figure 4). These residues cluster behind the conserved hydrophobic pocket and were identified because there mutational profile differed on average by 2 standard deviations from all other identical residues within the crystal structure.
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pone-0062427-g007: Significantly energetically-constrained amino acid positions identified by MUMBO Analysis.The 5 amino acids identified by MUMBO as having constrained ΔΔG mutational profiles relative to other identical amino acids within the crystal structure are color coded: ASN340 (red), Gly341 (orange) Ile342 (yellow) Glu343 (green), and Ser332 (cyan), are shown with respect to the TH2 and TH3 domains’ surface area (colored as in Figure 4). These residues cluster behind the conserved hydrophobic pocket and were identified because there mutational profile differed on average by 2 standard deviations from all other identical residues within the crystal structure.

Mentions: Calculation of Gibbs free energies on exhaustively mutagenized structures can provide information on structural constraints of a protein. Given the newly evolved fold-flap and pocket in the P. falciparum CS, polymorphic changes could reflect a lack of structural constraint in this region. To study the energetic constraints and effects of point mutations, we performed a comprehensive point mutation analysis of the structure using MUMBO software. This yields an estimate of Gibbs free energy required for each of the possible alternate states, indicating the favorability of making each of the 19 residue substitutions theoretically possible at each position in the reference sequence (Table S4). After quality control checks to validate the appropriateness of the method to the CS structure, we searched for residues which behaved anomalously (differing by at least 2 standard deviations) when changed from the reference state relative to similar residues at other positions. Five such residues were identified. Substitution of Asn 340 with Leu, Ile, and Val was predicted to be particularly favorable on the grounds of energetics (Figure S3), suggesting that mutation towards an aliphatic residue from a negatively charged residue was highly permissible. Substitution of Gln 343 to the aromatics residues His, Tyr, Trp, and Phe was strongly disfavored in this analysis and a similar trend was observed for Ser 332 and Ile 342. Substitution of Gly 341 by any other amino acid generates a substantial energy increase. This is supported by comparison between species of malaria, in which Gly 341 is conserved among all species, while the other residues have one alternate state (S332T, N340V, I342V, and Q343R) [19]. Upon mapping these residues to the structure (Figure 7), they clustered behind the conserved hydrophobic pocket, falling on β strand 2 except Ser332, which packs with β2. The observed tight packaging within the structure probably causes spatial constraints, disfavoring the incorporation of large amino acid substitutions. Gly appears to be selected for its small size, given the large van der Waals forces likely to be generated by substitutions from this smallest amino acid. The location of these restricted residues in relation to the pocket suggests that both the pocket and the packed core need to be highly conserved for stabilization of the molecule.


Diversity of T cell epitopes in Plasmodium falciparum circumsporozoite protein likely due to protein-protein interactions.

Aragam NR, Thayer KM, Nge N, Hoffman I, Martinson F, Kamwendo D, Lin FC, Sutherland C, Bailey JA, Juliano JJ - PLoS ONE (2013)

Significantly energetically-constrained amino acid positions identified by MUMBO Analysis.The 5 amino acids identified by MUMBO as having constrained ΔΔG mutational profiles relative to other identical amino acids within the crystal structure are color coded: ASN340 (red), Gly341 (orange) Ile342 (yellow) Glu343 (green), and Ser332 (cyan), are shown with respect to the TH2 and TH3 domains’ surface area (colored as in Figure 4). These residues cluster behind the conserved hydrophobic pocket and were identified because there mutational profile differed on average by 2 standard deviations from all other identical residues within the crystal structure.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3646838&req=5

pone-0062427-g007: Significantly energetically-constrained amino acid positions identified by MUMBO Analysis.The 5 amino acids identified by MUMBO as having constrained ΔΔG mutational profiles relative to other identical amino acids within the crystal structure are color coded: ASN340 (red), Gly341 (orange) Ile342 (yellow) Glu343 (green), and Ser332 (cyan), are shown with respect to the TH2 and TH3 domains’ surface area (colored as in Figure 4). These residues cluster behind the conserved hydrophobic pocket and were identified because there mutational profile differed on average by 2 standard deviations from all other identical residues within the crystal structure.
Mentions: Calculation of Gibbs free energies on exhaustively mutagenized structures can provide information on structural constraints of a protein. Given the newly evolved fold-flap and pocket in the P. falciparum CS, polymorphic changes could reflect a lack of structural constraint in this region. To study the energetic constraints and effects of point mutations, we performed a comprehensive point mutation analysis of the structure using MUMBO software. This yields an estimate of Gibbs free energy required for each of the possible alternate states, indicating the favorability of making each of the 19 residue substitutions theoretically possible at each position in the reference sequence (Table S4). After quality control checks to validate the appropriateness of the method to the CS structure, we searched for residues which behaved anomalously (differing by at least 2 standard deviations) when changed from the reference state relative to similar residues at other positions. Five such residues were identified. Substitution of Asn 340 with Leu, Ile, and Val was predicted to be particularly favorable on the grounds of energetics (Figure S3), suggesting that mutation towards an aliphatic residue from a negatively charged residue was highly permissible. Substitution of Gln 343 to the aromatics residues His, Tyr, Trp, and Phe was strongly disfavored in this analysis and a similar trend was observed for Ser 332 and Ile 342. Substitution of Gly 341 by any other amino acid generates a substantial energy increase. This is supported by comparison between species of malaria, in which Gly 341 is conserved among all species, while the other residues have one alternate state (S332T, N340V, I342V, and Q343R) [19]. Upon mapping these residues to the structure (Figure 7), they clustered behind the conserved hydrophobic pocket, falling on β strand 2 except Ser332, which packs with β2. The observed tight packaging within the structure probably causes spatial constraints, disfavoring the incorporation of large amino acid substitutions. Gly appears to be selected for its small size, given the large van der Waals forces likely to be generated by substitutions from this smallest amino acid. The location of these restricted residues in relation to the pocket suggests that both the pocket and the packed core need to be highly conserved for stabilization of the molecule.

Bottom Line: Using the information from these parasite populations, structural analysis reveals that polymorphic amino acids within TH2 and TH3 colocalize to one side of the protein, surround, but do not involve, the hydrophobic pocket in CS, and predominately involve charge switches.In addition, free energy analysis shows polymorphic residues tend to be populated by energetically unfavorable amino acids.In combination, these findings suggest the diversity of T cell epitopes in CS may be primarily an evolutionary response to intermolecular interactions at the surface of the protein potentially counteracting antibody-mediated immune recognition or evolving host receptor diversity.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, University of North Carolina School of Medicine, Chapel Hill, North Carolina, United States of America.

ABSTRACT
Circumsporozoite protein (CS) is a leading vaccine antigen for falciparum malaria, but is highly polymorphic in natural parasite populations. The factors driving this diversity are unclear, but non-random assortment of the T cell epitopes TH2 and TH3 has been observed in a Kenyan parasite population. The recent publication of the crystal structure of the variable C terminal region of the protein allows the assessment of the impact of diversity on protein structure and T cell epitope assortment. Using data from the Gambia (55 isolates) and Malawi (235 isolates), we evaluated the patterns of diversity within and between epitopes in these two distantly-separated populations. Only non-synonymous mutations were observed with the vast majority in both populations at similar frequencies suggesting strong selection on this region. A non-random pattern of T cell epitope assortment was seen in Malawi and in the Gambia, but structural analysis indicates no intramolecular spatial interactions. Using the information from these parasite populations, structural analysis reveals that polymorphic amino acids within TH2 and TH3 colocalize to one side of the protein, surround, but do not involve, the hydrophobic pocket in CS, and predominately involve charge switches. In addition, free energy analysis suggests residues forming and behind the novel pocket within CS are tightly constrained and well conserved in all alleles. In addition, free energy analysis shows polymorphic residues tend to be populated by energetically unfavorable amino acids. In combination, these findings suggest the diversity of T cell epitopes in CS may be primarily an evolutionary response to intermolecular interactions at the surface of the protein potentially counteracting antibody-mediated immune recognition or evolving host receptor diversity.

Show MeSH
Related in: MedlinePlus