The Chondroitin Sulfate A-binding Site of the VAR2CSA Protein Involves Multiple N-terminal Domains

Abstract

Malaria during pregnancy is a major health problem for African women. The disease is caused by Plasmodium falciparum malaria parasites, which accumulate in the placenta by adhering to chondroitin sulfate A (CSA). The interaction between infected erythrocytes and the placental receptor is mediated by a parasite expressed protein named VAR2CSA. A vaccine protecting pregnant women against placental malaria should induce antibodies inhibiting the interaction between VAR2CSA and CSA. Much effort has been put into defining the part of the 350 kDa VAR2CSA protein that is responsible for binding. It has been shown that full-length recombinant VAR2CSA binds specifically to CSA with high affinity, however to date no sub-fragment of VAR2CSA has been shown to interact with CSA with similar affinity or specificity. In this study, we used a biosensor technology to examine the binding properties of a panel of truncated VAR2CSA proteins. The experiments indicate that the core of the CSA-binding site is situated in three domains, DBL2X-CIDRPAM and a flanking domain, located in the N-terminal part of VAR2CSA. Furthermore, recombinant VAR2CSA subfragments containing this region elicit antibodies with high parasite adhesion blocking activity in animal immunization experiments.

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

“…In addition, we have recently shown that the core CSAbinding site lies within the DBL2X-CIDR PAM region. Specifically, we found two overlapping VAR2CSA fragments (DBL1X-CIDR PAM and ID1-DBL3X) showing CSA affinity comparable with that of the full-length protein (19). Srivastava et al (20) showed the same binding characteristics for DBL1X-DBL3X.…”

“…Solid Phase Binding Assay (ELISA)-The solid phase binding assay was performed as previously described (19). Briefly, microtiter plates were incubated with chondroitin sulfate proteoglycan (CSPG) (bovine decorin, D8428, Sigma) and heparan sulfate proteoglycan (HSPG) (H4777, Sigma) overnight at 4°C.…”

“…Such studies revealed that the full-length VAR2CSA ectodomain binds specifically to CSA with an affinity in the nanomolar range (13,14,19). In addition, we have recently shown that the core CSAbinding site lies within the DBL2X-CIDR PAM 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).…”