title: Refolding, purification, and crystallization of apical membrane antigen 1 from Plasmodium falciparum.
abstract: Extracellular domains of malaria antigens almost invariably contain disulphide linkages but lack N- and O-linked glycosylation. The best practical approach to generating recombinant extracellular Plasmodium proteins is not established and the problems encountered when using a bacterial expression/refolding approach are discussed in detail. Limited proteolysis experiments were used to identify a relatively non-flexible core region of the Plasmodium falciparum protein apical membrane antigen 1 (AMA1), and refolding/purification was used to generate two fragments of AMA1. Several chromatographically distinct AMA1 variants were identified that are presumably differentially refolded proteins. One of these AMA1 preparations proved to be crystallizable and generated two crystal forms that diffracted X-rays to 2 A resolution.
authors.affiliation: University of Maryland School of Pharmacy, 20 Penn Street, Baltimore, MD 21201, USA.
authors.forename: Aditi
authors.initials: A
authors.lastname: Gupta
authors.forename: Tao
authors.initials: T
authors.lastname: Bai
authors.forename: Vince
authors.initials: V
authors.lastname: Murphy
authors.forename: Phillip
authors.initials: P
authors.lastname: Strike
authors.forename: Robin F
authors.initials: RF
authors.lastname: Anders
authors.forename: Adrian H
authors.initials: AH
authors.lastname: Batchelor
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan / Q000737:chemistry* / Q000302:isolation & purification*; D001483:Base Sequence; D003460:Crystallization; D018360:Crystallography, X-Ray; D016054:DNA, Protozoan / Q000235:genetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D010957:Plasmids / Q000235:genetics; D010963:Plasmodium falciparum / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
chemical_list: D000911:Antibodies, Monoclonal; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D016054:DNA, Protozoan; D008565:Membrane Proteins; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C090077:apical membrane antigen I, Plasmodium