title: Structure and Characterisation of a Key Epitope in the Conserved C-Terminal Domain of the Malaria Vaccine Candidate MSP2.
abstract: Merozoite surface protein 2 (MSP2) is an intrinsically disordered antigen that is abundant on the surface of the malaria parasite Plasmodium falciparum. The two allelic families of MSP2, 3D7 and FC27, differ in their central variable regions, which are flanked by highly conserved C-terminal and N-terminal regions. In a vaccine trial, full-length 3D7 MSP2 induced a strain-specific protective immune response despite the detectable presence of conserved region antibodies. This work focuses on the conserved C-terminal region of MSP2, which includes the only disulphide bond in the protein and encompasses key epitopes recognised by the mouse monoclonal antibodies 4D11 and 9H4. Although the 4D11 and 9H4 epitopes are overlapping, immunofluorescence assays have shown that the mouse monoclonal antibody 4D11 binds to MSP2 on the merozoite surface with a much stronger signal than 9H4. Understanding the structural basis for this antigenic difference between these antibodies will help direct the design of a broad-spectrum and MSP2-based malaria vaccine. 4D11 and 9H4 were reengineered into antibody fragments [variable region fragment (Fv) and single-chain Fv (scFv)] and were validated as suitable models for their full-sized IgG counterparts by surface plasmon resonance and isothermal titration calorimetry. An alanine scan of the 13-residue epitope 3D7-MSP2207-222identified the minimal binding epitope of 4D11 and the key residues involved in binding. A 2.2-Å crystal structure of 4D11 Fv bound to the eight-residue epitope NKENCGAA provided valuable insight into the possible conformation of the C-terminal region of MSP2 on the parasite. This work underpins continued efforts to optimise recombinant MSP2 constructs for evaluation as potential vaccine candidates.
authors.affiliation: Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, Australia.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Seow
authors.affiliation: Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, Australia.
authors.forename: Rodrigo A V
authors.initials: RA
authors.lastname: Morales
authors.affiliation: Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, Australia.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: MacRaild
authors.affiliation: Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, Australia.
authors.forename: Bankala
authors.initials: B
authors.lastname: Krishnarjuna
authors.affiliation: Department of Microbiology, Monash University, Clayton 3168, Australia.
authors.forename: Sheena
authors.initials: S
authors.lastname: McGowan
authors.affiliation: Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, Australia.
authors.forename: Tamir
authors.initials: T
authors.lastname: Dingjan
authors.affiliation: NMR Research Centre, Indian Institute of Science, Bangalore, 560012, India.
authors.forename: Garima
authors.initials: G
authors.lastname: Jaipuria
authors.affiliation: Garvan Institute of Medical Research, Darlinghurst 2010, Australia.
authors.forename: Romain
authors.initials: R
authors.lastname: Rouet
authors.affiliation: National Deuteration Facility, Australian Nuclear Science and Technology Organisation, Lucas Heights 2234, Australia.
authors.forename: Karyn L
authors.initials: KL
authors.lastname: Wilde
authors.affiliation: NMR Research Centre, Indian Institute of Science, Bangalore, 560012, India.
authors.forename: Hanudatta S
authors.initials: HS
authors.lastname: Atreya
authors.affiliation: Centre for Biomedical Research, The Burnet Institute, Melbourne 3004, Australia.
authors.forename: Jack S
authors.initials: JS
authors.lastname: Richards
authors.affiliation: Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne 3086, Australia.
authors.forename: Robin F
authors.initials: RF
authors.lastname: Anders
authors.affiliation: Garvan Institute of Medical Research, Darlinghurst 2010, Australia.
authors.forename: Daniel
authors.initials: D
authors.lastname: Christ
authors.affiliation: Department of Microbiology, Monash University, Clayton 3168, Australia.
authors.forename: Nyssa
authors.initials: N
authors.lastname: Drinkwater
authors.affiliation: Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, Australia. Electronic address: ray.norton@monash.edu.
authors.forename: Raymond S
authors.initials: RS
authors.lastname: Norton
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D000913:Antibodies, Protozoan / Q000737:chemistry / Q000276:immunology; D000953:Antigens, Protozoan / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D002151:Calorimetry; D018360:Crystallography, X-Ray; D000939:Epitopes / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D017780:Malaria Vaccines / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D051379:Mice; D008958:Models, Molecular; D010963:Plasmodium falciparum / Q000235:genetics* / Q000276:immunology*; D011485:Protein Binding; D011487:Protein Conformation; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D020349:Surface Plasmon Resonance
chemical_list: D000911:Antibodies, Monoclonal; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D000939:Epitopes; D017780:Malaria Vaccines; D015800:Protozoan Proteins; C065974:merozoite surface protein 2, Plasmodium
keywords: antibody; disordered protein; malaria; merozoite surface protein 2; structure