Multimeric assembly, clustered interactions, and molecular complexes between parasite ligands and host-cell receptors for invasion.

<p>(<b>A</b>) PfTRAP engagement with heparan sulphate proteoglycans (HSPGs) on the hepatocyte surface; (<b>B</b>) proteolytic processing and shedding of PfMSP1 exposes the 19 kDa fragment (MSP1₁₉) that forms an invasion complex with MSP9 and the band 3 homodimer; (<b>C</b>) assembly of two PfEBA-175 monomers around dimeric glycophorin A of erythrocytes; (<b>D</b>) stepwise multimeric assembly of two PvDBP with two Duffy antigen/receptor for chemokines on reticulocyte surface; (<b>E</b>) monomeric interaction between PfEBA-140 and glycophorin C on erythrocytes; (<b>F</b>) proposed complexes of TgMIC2 and TgM2AP and of TgMIC1, TgMIC4, and TgMIC6 on the parasite surface; (<b>G</b>) variations in oligomeric states of GPI-anchored surface antigens (SAGs) create distinct interaction sites.</p

Multimeric assembly, clustered interactions, and molecular complexes between parasite ligands and host-cell receptors for invasion.

<p>(<b>A</b>) PfTRAP engagement with heparan sulphate proteoglycans (HSPGs) on the hepatocyte surface; (<b>B</b>) proteolytic processing and shedding of PfMSP1 exposes the 19 kDa fragment (MSP1₁₉) that forms an invasion complex with MSP9 and the band 3 homodimer; (<b>C</b>) assembly of two PfEBA-175 monomers around dimeric glycophorin A of erythrocytes; (<b>D</b>) stepwise multimeric assembly of two PvDBP with two Duffy antigen/receptor for chemokines on reticulocyte surface; (<b>E</b>) monomeric interaction between PfEBA-140 and glycophorin C on erythrocytes; (<b>F</b>) proposed complexes of TgMIC2 and TgM2AP and of TgMIC1, TgMIC4, and TgMIC6 on the parasite surface; (<b>G</b>) variations in oligomeric states of GPI-anchored surface antigens (SAGs) create distinct interaction sites.</p

ELISA with anti-DBP conformational specific antibodies.

<p>ELISA assays for conformational anti-DBP antibodies with BSA as a negative control, Sal1 DBP-II as a positive control, and DEKnull protein. Four inhibitory (3C9, 2H2, 2C6, 2D10) and two non-inhibitory (3D10, 2F12) antibodies were tested.</p

DEKnull is structurally similar to Sal1 DBP-II.

<p>(A) DEKnull separated into three sub-domains, sub-domain 1 (S1—red), sub-domain 2 (S2—blue), and sub-domain 3 (S3—green). (B) S1 (red) contains a β-hairpin, S2 (blue) is a helix bundle, and S3 (green) is a helix bundle. Domain boundaries and disulfide bonding cysteines are labeled. (C) Structural alignment of DEKnull (solid colors) with Sal1 DBP-II (light tinted colors) with r.m.s. deviation of 0.435Å. (D) Structural alignment of individual DEKnull sub-domains (solid colors) with Sal1 DBP-II sub-domains (light tinted colors). S1 alignment has a r.m.s. deviation of 0.308 Å. S2 alignment has a r.m.s. deviation of 0.288 Å. S3 alignment has a r.m.s. deviation of 0.310 Å.</p

DEKnull mutations do not affect protein secondary structure.

<p>(A) and (B) Two views of a structural alignment of Sal1 DBP-II DEKAQQRRKQ polymorphic region (white) and DEKnull ASTAATSRTS mutant region (blue). Mutated residues are labeled and shown as sticks. (C) Structural alignment of DARC binding sites on Sal1 DBP-II (white) and DEKnull (blue). DARC binding residues are labeled and shown as sticks. (D) Structural alignment of DBL dimerization interfaces on Sal1 DBP-II (white) and DEKnull (blue). Dimerization residues are labeled and shown as sticks.</p

DiscoTope B-cell epitope prediction of Sal1 DBP-II and DEKnull.

<p>(A) Graphical representation of DiscoTope B-cell epitope scores for Sal1 DBP-II (blue line) and DEKnull (red line). Prediction threshold is shown in green. DEK residues are located at amino acids 339–348 and shown above the corresponding location in the graph. (B) Two views of Sal1 DBP-II predicted epitopes mapped onto crystal structure. (C) Two views of DEKnull predicted epitopes mapped onto crystal structure.</p

Data collection and refinement statistics for DEKnull.

<p>Data were collected from a single crystal.</p><p>*Highest resolution shell is shown in parenthesis</p><p>^†Does not include hydrogens</p><p>Data collection and refinement statistics for DEKnull.</p

Location of DEK polymorphisms on Sal1 DBP-II.

<p>DEK polymorphisms (DEKAQQRRKQ) mapped onto the Sal1 DBP-II and DARC heterotetramer. DEK residues are shown in brown and side chains are displayed as sticks. The two DBP-II molecules are in green and yellow. The two DARC peptides are in purple and blue.</p

Antibody epitope identification by immunofluorescence assay.

<p>DBL domain proteins were surface expressed on HEK293 cells and probed with R217 or R218 as primary and Alexafluor-546 labeled anti-IgG₁ as secondary. Green channel shows GFP tagged expressed protein. Red channel shows Alexafluor-546 labeled proteins on HEK293 cell surface. Merged channel shows overlap between green and red channels.</p

Crystal structure of F1/R218 Fab complex.

<p>(A) Overall structure of the F1/R218 Fab complex shown in ribbon representation. The F2 domain of RII is colored in green. The Fab heavy chain (VH) is in blue and the light chain (VL) in pink. (B) Ribbon representation of F1 mapping the R218 epitope. Residues contacted by the Fab are show in stick. Residues contacted by the heavy chain are colored blue, residues contacted by the Fab light chain are colored orange, and residues contacted by both chains are in beige. Residues not contacted by the antibody are in green. (C) Surface representation of F2 mapping the R217 epitope. Color scheme as in B. (D) Surface representation of the R218 Fab, mapping heavy chain residues (blue) that contact F2 (green). The light chain is shown in white. (E) Surface representation of the R218 Fab, mapping light chain residues (orange) that contact F2 (green). The heavy chain is shown in white.</p