FC0107
Merozoite surface protein 2 (MSP2) 3D7  -  mAb6D8

Biological function
MSP2 is an abundant and polymorphic antigen of the invasive blood-stage form of the malaria parasite, which is glycosylphosphatidylinositol (GPI)-anchored to the membrane surface. MSP2 elicits strain-specific response in humans, but antibodies bind quite poorly to its conserved N- and C-terminal regions. The monoclonal antibody (mAb) 6D8 recognizes a conserved N-terminal epitope on recombinant MSP2.

Domain organization/sequence features
All variants of MSP2 share conserved N- and C-terminal regions. Two allelic families, 3D7 and FC27, differ in tandem repeats and dimorphic flanking sequences within the central region of the protein. The polymorphic region is immunodominant in the natural immune response to MSP2.

Structural evidence
In complex with mAb 6D8, residues beyond MSP2 residue 24 do not make contacts with the antibody. In the complex of 3D7- MSP2_14-30 - 6D8 Fv, residues 24-28 were visible, yet with higher B-factors. The C-terminal extension of the peptide induced significant differences in the ¹H,¹⁵N HSQC spectrum of 6D8 bound to the extended peptide 3D7-MSP2_14-30 shows significant differences as compared to the spectrum bound to the crystallographically defined epitope, suggesting that MSP2 residues 24 to 30 interact with 6D8 scFv in solution, even though such interactions were not observed crystallographically. The spectra of 6D8 scFv bound to MSP2_14-30 from different strains (FC27-MSP2_14-30 and 3D7-MSP2_14-30) exhibit differences in intensity and/or chemical shifts corresponding to interactions exchanging in a timescale of ms or slower.

Biochemical evidence
Based on SPR, the affinity of the 11-23 residue segment of MSP2 for mAb 6D8 is 6 nM, while that of 15-22 segment is 87 nM. N-terminal extensions (14-22 segment) restored affinity, while C-terminal extensions only slightly improved binding. Based on ITC 3D7-MSP2_14-30 has K_d of 64±12 nM, comparable to the full-length 3D7 MSP2 K_d of 61±9 nM.

Structure/Mechanism
Additionally, the N-terminal 10-22 region adopts an α-helix upon binding to the membrane (DPC micelles), which is incompatible with mAb 6D8. Most likely, C-terminal extension could balance between the two binding partners via modulating the population of the mAb 6D8 competent state and tuning the entropic contribution of the mAb 6D8 binding.

Mechanism category
competitive binding

Significance
Fuzziness enables strain-specific binding to the antibody owing to transient interactions established by the variable, C-terminal extension of the antigen epitope.

Submitted by
Chris MacRaild   chris.macraild@monash.edu