title: Characterization of a protective Escherichia coli-expressed Plasmodium falciparum merozoite surface protein 3 indicates a non-linear, multi-domain structure.
abstract: Immunization with a recombinant yeast-expressed Plasmodium falciparum merozoite surface protein 3 (MSP3) protected Aotus nancymai monkeys against a virulent challenge infection. Unfortunately, the production process for this yeast-expressed material was not optimal for human trials. In an effort to produce a recombinant MSP3 protein in a scaleable manner, we expressed and purified near-full-length MSP3 in Escherichia coli (EcMSP3). Purified EcMSP3 formed non-globular dimers as determined by analytical size-exclusion HPLC with in-line multi-angle light scatter and quasi-elastic light scatter detection and velocity sedimentation (R(h) 7.6+/-0.2nm and 6.9nm, respectively). Evaluation by high-resolution atomic force microscopy revealed non-linear asymmetric structures, with beaded domains and flexible loops that were recognized predominantly as dimers, although monomers and larger multimers were observed. The beaded substructure corresponds to predicted structural domains, which explains the velocity sedimentation results and improves the conceptual model of the protein. Vaccination with EcMSP3 in Freund's adjuvant-induced antibodies that recognized native MSP3 in parasitized erythrocytes by an immunofluorescence assay and gave delayed time to treatment in a group of Aotus monkeys in a virulent challenge infection with the FVO strain of P. falciparum. Three of the seven monkeys vaccinated with EcMSP3 had low peak parasitemias. EcMSP3, which likely mimics the native MSP3 structure located on the merozoite surface, is a viable candidate for inclusion in a multi-component malaria vaccine.
authors.affiliation: Malaria Vaccine Development Branch, National Institute of Allergy and Infectious Disease, National Institutes of Health, Rockville, MD 20852, United States.
authors.forename: Chiawei W
authors.initials: CW
authors.lastname: Tsai
authors.forename: Peter F
authors.initials: PF
authors.lastname: Duggan
authors.forename: Albert J
authors.initials: AJ
authors.lastname: Jin
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Macdonald
authors.forename: Svetlana
authors.initials: S
authors.lastname: Kotova
authors.forename: Jacob
authors.initials: J
authors.lastname: Lebowitz
authors.forename: Darrell E
authors.initials: DE
authors.lastname: Hurt
authors.forename: Richard L
authors.initials: RL
authors.lastname: Shimp
authors.forename: Lynn
authors.initials: L
authors.lastname: Lambert
authors.forename: Louis H
authors.initials: LH
authors.lastname: Miller
authors.forename: Carole A
authors.initials: CA
authors.lastname: Long
authors.forename: Allan
authors.initials: A
authors.lastname: Saul
authors.forename: David L
authors.initials: DL
authors.lastname: Narum
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D016645:Aotidae; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D017780:Malaria Vaccines / Q000276:immunology; D016778:Malaria, Falciparum / Q000276:immunology / Q000517:prevention & control; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000737:chemistry* / Q000235:genetics; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D011897:Random Allocation; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics; D014614:Vaccines, Synthetic / Q000276:immunology
chemical_list: D000953:Antigens, Protozoan; D017780:Malaria Vaccines; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D014614:Vaccines, Synthetic; C089133:merozoite surface protein 3, Plasmodium