title: Structure of the Plasmodium falciparum circumsporozoite protein, a leading malaria vaccine candidate.
abstract: The Plasmodium falciparum circumsporozoite protein (CSP) is critical for sporozoite function and invasion of hepatocytes. Given its critical nature, a phase III human CSP malaria vaccine trial is ongoing. The CSP is composed of three regions as follows: an N terminus that binds heparin sulfate proteoglycans, a four amino acid repeat region (NANP), and a C terminus that contains a thrombospondin-like type I repeat (TSR) domain. Despite the importance of CSP, little is known about its structure. Therefore, recombinant forms of CSP were produced by expression in both Escherichia coli (Ec) and then refolded (EcCSP) or in the methylotrophic yeast Pichia pastoris (PpCSP) for structural analyses. To analyze the TSR domain of recombinant CSP, conformation-dependent monoclonal antibodies that recognized unfixed P. falciparum sporozoites and inhibited sporozoite invasion of HepG2 cells in vitro were identified. These monoclonal antibodies recognized all recombinant CSPs, indicating the recombinant CSPs contain a properly folded TSR domain structure. Characterization of both EcCSP and PpCSP by dynamic light scattering and velocity sedimentation demonstrated that both forms of CSP appeared as highly extended proteins (R(h) 4.2 and 4.58 nm, respectively). Furthermore, high resolution atomic force microscopy revealed flexible, rod-like structures with a ribbon-like appearance. Using this information, we modeled the NANP repeat and TSR domain of CSP. Consistent with the biochemical and biophysical results, the repeat region formed a rod-like structure about 21-25 nm in length and 1.5 nm in width. Thus native CSP appears as a glycosylphosphatidylinositol-anchored, flexible rod-like protein on the sporozoite surface.
authors.affiliation: Malaria Vaccine Development Branch, NIAID, National Institutes of Health, Rockville, Maryland 20852, USA.
authors.forename: Matthew L
authors.initials: ML
authors.lastname: Plassmeyer
authors.forename: Karine
authors.initials: K
authors.lastname: Reiter
authors.forename: Richard L
authors.initials: RL
authors.lastname: Shimp
authors.forename: Svetlana
authors.initials: S
authors.lastname: Kotova
authors.forename: Paul D
authors.initials: PD
authors.lastname: Smith
authors.forename: Darrell E
authors.initials: DE
authors.lastname: Hurt
authors.forename: Brent
authors.initials: B
authors.lastname: House
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Zou
authors.forename: Yanling
authors.initials: Y
authors.lastname: Zhang
authors.forename: Merrit
authors.initials: M
authors.lastname: Hickman
authors.forename: Onyinyechukwu
authors.initials: O
authors.lastname: Uchime
authors.forename: Raul
authors.initials: R
authors.lastname: Herrera
authors.forename: Vu
authors.initials: V
authors.lastname: Nguyen
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Glen
authors.forename: Jacob
authors.initials: J
authors.lastname: Lebowitz
authors.forename: Albert J
authors.initials: AJ
authors.lastname: Jin
authors.forename: Louis H
authors.initials: LH
authors.lastname: Miller
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: MacDonald
authors.forename: Yimin
authors.initials: Y
authors.lastname: Wu
authors.forename: David L
authors.initials: DL
authors.lastname: Narum
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000953:Antigens, Protozoan / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D001665:Binding Sites; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006493:Heparin / Q000031:analogs & derivatives / Q000378:metabolism; D022781:Hepatocytes / Q000276:immunology / Q000469:parasitology / Q000473:pathology; D006801:Humans; D015151:Immunoblotting; D017780:Malaria Vaccines / Q000276:immunology*; D008958:Models, Molecular; D010963:Plasmodium falciparum / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011509:Proteoglycans / Q000378:metabolism; D015800:Protozoan Proteins / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D034101:Sporozoites / Q000276:immunology* / Q000378:metabolism; D013696:Temperature; D014461:Ultracentrifugation
chemical_list: D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D017261:Glycosylphosphatidylinositols; D017780:Malaria Vaccines; D011509:Proteoglycans; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C041595:circumsporozoite protein, Protozoan; C036090:heparin proteoglycan; D006493:Heparin