Var2CSA Minimal CSA Binding Region Is Located within the N-Terminal Region

Abstract

Var2CSA, a key molecule linked with pregnancy-associated malaria (PAM), causes sequestration of Plasmodium falciparum infected erythrocytes (PEs) in the placenta by adhesion to chondroitin sulfate A (CSA). Var2CSA possesses a 300 kDa extracellular region composed of six Duffy-binding like (DBL) domains and a cysteine-rich interdomain region (CIDRpam) module. Although initial studies implicated several individual var2CSA DBL domains as important for adhesion of PEs to CSA, new studies revealed that these individual domains lack both the affinity and specificity displayed by the full-length extracellular region. Indeed, recent evidence suggests the presence of a single CSA-binding site formed by a higher-order domain organization rather than several independent binding sites located on the different domains. Here, we search for the minimal binding region within var2CSA that maintains high affinity and specificity for CSA binding, a characteristic feature of the full-length extracellular region. Accordingly, truncated recombinant var2CSA proteins comprising different domain combinations were expressed and their binding characteristics assessed against different sulfated glycosaminoglycans (GAGs). Our results indicate that the smallest region within var2CSA with similar binding properties to those of the full-length var2CSA is DBL1X-3X. We also demonstrate that inhibitory antibodies raised in rabbit against the full-length DBL1X-6ε target principally DBL3X and, to a lesser extent, DBL5ε. Taken together, our results indicate that efforts should focus on the DBL1X-3X region for developing vaccine and therapeutic strategies aimed at combating PAM.

Structural and Functional Insight into How the Plasmodium falciparum VAR2CSA Protein Mediates Binding to Chondroitin Sulfate A in Placental Malaria

“…Core CSA-binding Site Lies within DBL2X Domain-It has been suggested that the minimal CSA-binding region in VAR2CSA lies within DBL2X-ID2b, with the need of flanking domains for full affinity binding (19,20). Here we have analyzed shorter fragments of VAR2CSA to further map the regions required for CSA binding (Fig.…”

“…Previous work has suggested that the minimal CSA binding region in VAR2CSA is DBL2X-ID2b, with the need for DBL1X or DBL3X for full affinity binding (19,20). In continuation of this work we made further truncations of VAR2CSA, focusing on the DBL2X region.…”

“…The large size and complex structure, along with genetic variability among isolates complicates current large scale production strategies. The focus in vaccine development is therefore to produce a small fragment of VAR2CSA that has the ability to induce functional cross-inhibitory antibodies (19,20). In this, the obvious approach is to define the binding regions within VAR2CSA and examine the mechanism underlying the interaction.…”

“…The expression of the full-length VAR2CSA ectodomain (FV2) from FCR3 (13,19) and 3D7 (14,20) type parasites has enabled us to study both the binding properties and the structural characteristics of the VAR2CSA protein. Such studies revealed that the full-length VAR2CSA ectodomain binds specifically to CSA with an affinity in the nanomolar range (13,14,19).…”

Cytoadhesion of Plasmodium falciparum–infected erythrocytes to chondroitin-4-sulfate is cooperative and shear enhanced

“…VAR2CSA harbors 6 DBL domains, several of which have been implicated in CSA binding, although no consensus has been reached as to whether all or only individual DBL domains interact with CSA. 32,[39][40][41][42][43] To investigate the putative cooperative nature of the VAR2CSA/CSA interaction, we attempted to inhibit domain-specific regions of VAR2CSA using antibodies against the individual DBL domains. 26 We reasoned that these DBL domainspecific antibodies might interfere with substrate-induced allosteric changes and, thus, with the degree of cooperativity.…”