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PfRH5: a novel reticulocyte-binding family homolog of plasmodium falciparum that binds to the erythrocyte, and an investigation of its receptor.

Rodriguez M, Lustigman S, Montero E, Oksov Y, Lobo CA - PLoS ONE (2008)

Bottom Line: Attachment is inhibited if the target cells are exposed to high concentrations of trypsin, but not to lower concentrations or to chymotrypsin or neuraminidase.We have determined the affinity, copy number and apparent molecular mass of the receptor protein.Thus, we have shown that PfRH5 is a novel erythrocyte-binding ligand and the identification and partial characterization of the new RBC receptor may indicate the existence of an unrecognized P. falciparum invasion pathway.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Blood-Borne Parasites, Lindsley Kimball Research Institute, The New York Blood Center, New York, New York, USA.

ABSTRACT
Multiple interactions between parasite ligands and their receptors on the human erythrocyte are a condition of successful Plasmodium falciparum invasion. The identification and characterization of these receptors presents a major challenge in the effort to understand the mechanism of invasion and to develop the means to prevent it. We describe here a novel member of the reticulocyte-binding family homolog (RH) of P. falciparum, PfRH5, and show that it binds to a previously unrecognized receptor on the RBC. PfRH5 is expressed as a 63 kDa protein and localized at the apical end of the invasive merozoite. We have expressed a fragment of PfRH5 which contains the RBC-binding domain and exhibits the same pattern of interactions with the RBC as the parent protein. Attachment is inhibited if the target cells are exposed to high concentrations of trypsin, but not to lower concentrations or to chymotrypsin or neuraminidase. We have determined the affinity, copy number and apparent molecular mass of the receptor protein. Thus, we have shown that PfRH5 is a novel erythrocyte-binding ligand and the identification and partial characterization of the new RBC receptor may indicate the existence of an unrecognized P. falciparum invasion pathway.

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Erythrocyte-binding activity of native PfRH5.A Binding of the native PfRH5 protein in the HB3 culture supernatant incubated with untreated (WT) erythrocytes, variously enzyme-treated erythrocytes (N: neuraminidase; HT, high trypsin; C, chymotrypsin). The PfRH5 parasite protein was detected in the eluate fractions by immunoprecipitation with anti-RH5 antibodies in WT, N and C lanes but not HT. B The same eluate samples from (A) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to wild type and chymotrypsin-treated erythrocytes but not to neuraminidase and high trypsin-treated erythrocytes C Binding of the native RH5 protein to erythrocytes treated with lower concentrations of trypsin (LT low trypsin and MT: moderate trypsin). D The eluate samples from (C) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to untreated erythrocytes but not to low-trypsin- and moderate-trypsin-treated erythrocytes.
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pone-0003300-g003: Erythrocyte-binding activity of native PfRH5.A Binding of the native PfRH5 protein in the HB3 culture supernatant incubated with untreated (WT) erythrocytes, variously enzyme-treated erythrocytes (N: neuraminidase; HT, high trypsin; C, chymotrypsin). The PfRH5 parasite protein was detected in the eluate fractions by immunoprecipitation with anti-RH5 antibodies in WT, N and C lanes but not HT. B The same eluate samples from (A) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to wild type and chymotrypsin-treated erythrocytes but not to neuraminidase and high trypsin-treated erythrocytes C Binding of the native RH5 protein to erythrocytes treated with lower concentrations of trypsin (LT low trypsin and MT: moderate trypsin). D The eluate samples from (C) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to untreated erythrocytes but not to low-trypsin- and moderate-trypsin-treated erythrocytes.

Mentions: Having established its location at the invasive apical end of the merozoite and homology with other known RBC binding proteins, we performed assays to determine whether native PfRH5 binds to RBCs. The native full-length PfRH5 was isolated from [35S]methionine/cysteine-labeled culture supernatants (HB3 strain) that contained merozoites released from infected erythrocytes in the absence of target erythrocytes. Studies have shown that extracellular merozoites release parasite proteins into the culture, and such culture supernatants can be a source of parasite ligands that bind erythrocytes. Thus, [35S]methionine/cysteine-labeled spent merozoite supernatants were used as the source of RBC-binding proteins, and when the eluate was immunoprecipitated with anti-RH5 antiserum, a dominant band at ∼63 kDa was seen (Fig. 3A, lane WT). Thus, PfRH5 appears to be an adhesin that participates in invasion by binding to the RBC surface.


PfRH5: a novel reticulocyte-binding family homolog of plasmodium falciparum that binds to the erythrocyte, and an investigation of its receptor.

Rodriguez M, Lustigman S, Montero E, Oksov Y, Lobo CA - PLoS ONE (2008)

Erythrocyte-binding activity of native PfRH5.A Binding of the native PfRH5 protein in the HB3 culture supernatant incubated with untreated (WT) erythrocytes, variously enzyme-treated erythrocytes (N: neuraminidase; HT, high trypsin; C, chymotrypsin). The PfRH5 parasite protein was detected in the eluate fractions by immunoprecipitation with anti-RH5 antibodies in WT, N and C lanes but not HT. B The same eluate samples from (A) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to wild type and chymotrypsin-treated erythrocytes but not to neuraminidase and high trypsin-treated erythrocytes C Binding of the native RH5 protein to erythrocytes treated with lower concentrations of trypsin (LT low trypsin and MT: moderate trypsin). D The eluate samples from (C) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to untreated erythrocytes but not to low-trypsin- and moderate-trypsin-treated erythrocytes.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2553180&req=5

pone-0003300-g003: Erythrocyte-binding activity of native PfRH5.A Binding of the native PfRH5 protein in the HB3 culture supernatant incubated with untreated (WT) erythrocytes, variously enzyme-treated erythrocytes (N: neuraminidase; HT, high trypsin; C, chymotrypsin). The PfRH5 parasite protein was detected in the eluate fractions by immunoprecipitation with anti-RH5 antibodies in WT, N and C lanes but not HT. B The same eluate samples from (A) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to wild type and chymotrypsin-treated erythrocytes but not to neuraminidase and high trypsin-treated erythrocytes C Binding of the native RH5 protein to erythrocytes treated with lower concentrations of trypsin (LT low trypsin and MT: moderate trypsin). D The eluate samples from (C) were used for the detection of EBA-175 by immunoprecipitation. EBA-175 binds to untreated erythrocytes but not to low-trypsin- and moderate-trypsin-treated erythrocytes.
Mentions: Having established its location at the invasive apical end of the merozoite and homology with other known RBC binding proteins, we performed assays to determine whether native PfRH5 binds to RBCs. The native full-length PfRH5 was isolated from [35S]methionine/cysteine-labeled culture supernatants (HB3 strain) that contained merozoites released from infected erythrocytes in the absence of target erythrocytes. Studies have shown that extracellular merozoites release parasite proteins into the culture, and such culture supernatants can be a source of parasite ligands that bind erythrocytes. Thus, [35S]methionine/cysteine-labeled spent merozoite supernatants were used as the source of RBC-binding proteins, and when the eluate was immunoprecipitated with anti-RH5 antiserum, a dominant band at ∼63 kDa was seen (Fig. 3A, lane WT). Thus, PfRH5 appears to be an adhesin that participates in invasion by binding to the RBC surface.

Bottom Line: Attachment is inhibited if the target cells are exposed to high concentrations of trypsin, but not to lower concentrations or to chymotrypsin or neuraminidase.We have determined the affinity, copy number and apparent molecular mass of the receptor protein.Thus, we have shown that PfRH5 is a novel erythrocyte-binding ligand and the identification and partial characterization of the new RBC receptor may indicate the existence of an unrecognized P. falciparum invasion pathway.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Blood-Borne Parasites, Lindsley Kimball Research Institute, The New York Blood Center, New York, New York, USA.

ABSTRACT
Multiple interactions between parasite ligands and their receptors on the human erythrocyte are a condition of successful Plasmodium falciparum invasion. The identification and characterization of these receptors presents a major challenge in the effort to understand the mechanism of invasion and to develop the means to prevent it. We describe here a novel member of the reticulocyte-binding family homolog (RH) of P. falciparum, PfRH5, and show that it binds to a previously unrecognized receptor on the RBC. PfRH5 is expressed as a 63 kDa protein and localized at the apical end of the invasive merozoite. We have expressed a fragment of PfRH5 which contains the RBC-binding domain and exhibits the same pattern of interactions with the RBC as the parent protein. Attachment is inhibited if the target cells are exposed to high concentrations of trypsin, but not to lower concentrations or to chymotrypsin or neuraminidase. We have determined the affinity, copy number and apparent molecular mass of the receptor protein. Thus, we have shown that PfRH5 is a novel erythrocyte-binding ligand and the identification and partial characterization of the new RBC receptor may indicate the existence of an unrecognized P. falciparum invasion pathway.

Show MeSH