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Determination of the molecular basis for a limited dimorphism, N417K, in the Plasmodium vivax Duffy-binding protein.

McHenry AM, Barnes SJ, Ntumngia FB, King CL, Adams JH - PLoS ONE (2011)

Bottom Line: In natural isolates only two residues are found at this site, asparagine (N) and lysine (K).Our results suggest that the observed dimorphism likely arose due to both structural requirements and immune selection pressure.To our knowledge, this is the first exhaustive examination of this kind of the role of a single amino acid residue in antigenic character and binding ability.

View Article: PubMed Central - PubMed

Affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, Indiana, United States of America.

ABSTRACT
Invasion of human red blood cells by Plasmodium merozoites is vital for replication and survival of the parasite and, as such, is an attractive target for therapeutic intervention. Merozoite invasion is mediated by specific interactions between parasite ligands and host erythrocyte receptors. The P. vivax Duffy-binding protein (PvDBP) is heavily dependent on the interaction with the human Duffy blood group antigen/receptor for chemokines (DARC) for invasion. Region II of PvDBP contains many allelic polymorphisms likely to have arisen by host immune selection. Successful vaccine development necessitates a deeper understanding of the role of these polymorphisms in both parasite function and evasion of host immunity. A 3D structure of the homologous P. knowlesi DBP predicts that most variant residues are surface-exposed, including N417K, which is a dimorphic residue change that has previously been shown to be part of a linked haplotype that alters DBP sensitivity to inhibitory antibody. In natural isolates only two residues are found at this site, asparagine (N) and lysine (K). Site-directed mutagenesis of residue 417 was used to create a panel of 20 amino acid variants that were then examined for their binding phenotype and response to immune sera. Our results suggest that the observed dimorphism likely arose due to both structural requirements and immune selection pressure. To our knowledge, this is the first exhaustive examination of this kind of the role of a single amino acid residue in antigenic character and binding ability. Our results demonstrate that a single amino acid substitution can dramatically alter both the ability of the PvDBP to bind to human erythrocytes and its antigenic character.

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Related in: MedlinePlus

Percentage erythrocyte binding to DBP variants containing single amino acid substitutions at the 417 site.Binding is shown as a percentage compared to the naturally occurring variant N. All constructs are on a Sal I strain background. Polar residues are shown in black. Nonpolar residues are shown in gray. § indicates naturally occurring variant containing N or K at the 417 site. * indicates DBP variants which have significantly lower binding than one or both of the naturally occurring residues (N and K) (p<0.05). Statistical differences were analyzed using a 1-way analysis of variance (ANOVA) and a Tukey's posttest.
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pone-0020192-g001: Percentage erythrocyte binding to DBP variants containing single amino acid substitutions at the 417 site.Binding is shown as a percentage compared to the naturally occurring variant N. All constructs are on a Sal I strain background. Polar residues are shown in black. Nonpolar residues are shown in gray. § indicates naturally occurring variant containing N or K at the 417 site. * indicates DBP variants which have significantly lower binding than one or both of the naturally occurring residues (N and K) (p<0.05). Statistical differences were analyzed using a 1-way analysis of variance (ANOVA) and a Tukey's posttest.

Mentions: We analyzed the binding phenotype of variant constructs containing each of the 20 amino acids at site 417 using an in vitro COS7 cell assay for DBP-erythrocyte binding function (Fig. 1). Nine of the constructs bound significantly less than the naturally occurring residues, N and K (p<0.05). Most of these residues were nonpolar although Y was also a poor binder. Nine of the variants, including two nonpolar residues (A and G), were not significantly lower than the naturally occurring residues in their binding phenotype (p>0.05) and did not bind to Duffy negative cells. We further analyzed these nine ‘normal binding’ variants for their inhibition phenotypes using polyclonal rabbit sera raised against the Sal I strain DBPII containing N at the 417 site (Fig. 2). This well characterized serum inhibits in vitro binding of DBPII to erythrocytes and the DARC receptor, as well as inhibits invasion of human erythrocytes by P. vivax parasites [26].


Determination of the molecular basis for a limited dimorphism, N417K, in the Plasmodium vivax Duffy-binding protein.

McHenry AM, Barnes SJ, Ntumngia FB, King CL, Adams JH - PLoS ONE (2011)

Percentage erythrocyte binding to DBP variants containing single amino acid substitutions at the 417 site.Binding is shown as a percentage compared to the naturally occurring variant N. All constructs are on a Sal I strain background. Polar residues are shown in black. Nonpolar residues are shown in gray. § indicates naturally occurring variant containing N or K at the 417 site. * indicates DBP variants which have significantly lower binding than one or both of the naturally occurring residues (N and K) (p<0.05). Statistical differences were analyzed using a 1-way analysis of variance (ANOVA) and a Tukey's posttest.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3101244&req=5

pone-0020192-g001: Percentage erythrocyte binding to DBP variants containing single amino acid substitutions at the 417 site.Binding is shown as a percentage compared to the naturally occurring variant N. All constructs are on a Sal I strain background. Polar residues are shown in black. Nonpolar residues are shown in gray. § indicates naturally occurring variant containing N or K at the 417 site. * indicates DBP variants which have significantly lower binding than one or both of the naturally occurring residues (N and K) (p<0.05). Statistical differences were analyzed using a 1-way analysis of variance (ANOVA) and a Tukey's posttest.
Mentions: We analyzed the binding phenotype of variant constructs containing each of the 20 amino acids at site 417 using an in vitro COS7 cell assay for DBP-erythrocyte binding function (Fig. 1). Nine of the constructs bound significantly less than the naturally occurring residues, N and K (p<0.05). Most of these residues were nonpolar although Y was also a poor binder. Nine of the variants, including two nonpolar residues (A and G), were not significantly lower than the naturally occurring residues in their binding phenotype (p>0.05) and did not bind to Duffy negative cells. We further analyzed these nine ‘normal binding’ variants for their inhibition phenotypes using polyclonal rabbit sera raised against the Sal I strain DBPII containing N at the 417 site (Fig. 2). This well characterized serum inhibits in vitro binding of DBPII to erythrocytes and the DARC receptor, as well as inhibits invasion of human erythrocytes by P. vivax parasites [26].

Bottom Line: In natural isolates only two residues are found at this site, asparagine (N) and lysine (K).Our results suggest that the observed dimorphism likely arose due to both structural requirements and immune selection pressure.To our knowledge, this is the first exhaustive examination of this kind of the role of a single amino acid residue in antigenic character and binding ability.

View Article: PubMed Central - PubMed

Affiliation: Eck Institute for Global Health, University of Notre Dame, Notre Dame, Indiana, United States of America.

ABSTRACT
Invasion of human red blood cells by Plasmodium merozoites is vital for replication and survival of the parasite and, as such, is an attractive target for therapeutic intervention. Merozoite invasion is mediated by specific interactions between parasite ligands and host erythrocyte receptors. The P. vivax Duffy-binding protein (PvDBP) is heavily dependent on the interaction with the human Duffy blood group antigen/receptor for chemokines (DARC) for invasion. Region II of PvDBP contains many allelic polymorphisms likely to have arisen by host immune selection. Successful vaccine development necessitates a deeper understanding of the role of these polymorphisms in both parasite function and evasion of host immunity. A 3D structure of the homologous P. knowlesi DBP predicts that most variant residues are surface-exposed, including N417K, which is a dimorphic residue change that has previously been shown to be part of a linked haplotype that alters DBP sensitivity to inhibitory antibody. In natural isolates only two residues are found at this site, asparagine (N) and lysine (K). Site-directed mutagenesis of residue 417 was used to create a panel of 20 amino acid variants that were then examined for their binding phenotype and response to immune sera. Our results suggest that the observed dimorphism likely arose due to both structural requirements and immune selection pressure. To our knowledge, this is the first exhaustive examination of this kind of the role of a single amino acid residue in antigenic character and binding ability. Our results demonstrate that a single amino acid substitution can dramatically alter both the ability of the PvDBP to bind to human erythrocytes and its antigenic character.

Show MeSH
Related in: MedlinePlus