title: Molecular characterizations of three distinct Babesia gibsoni rhoptry-associated protein-1s (RAP-1s).
abstract: Three cDNAs encoding rhoptry-associated protein 1 (RAP-1) homologues were found in the Babesia gibsoni EST database. Based on similarities to BgRAP-1a, which was identified previously by serological screening of a cDNA merozoite library, the two new genes were designated BgRAP-1b (33.7%) and BgRAP-1c (57%). Mice antiserum raised against each recombinant protein reacted specifically with B. gibsoni parasites as determined by Western blotting, which showed native molecular sizes of the BgRAP-1a (51 kDa), BgRAP-1b (53 kDa) and BgRAP-1c (47 kDa) consistent with predictable molecular weights. Immunofluoresence using these antibodies revealed localization of all BgRAP-1s within the matrix of merozoites; however, BgRAP-1a appeared to diverge from the other two when it was found secreted into the cytoplasm of infected erythrocytes. Apical localization of all 3 BgRAP-1s during the extracellular stage of the parasite combined with their ability to bind a canine erythrocyte membrane fraction was suggestive of a role for these proteins in erythrocyte attachment. Lastly, the ability of these recombinant proteins to be used as diagnostic reagents was tested by ELISA and the sensitivities of BgRAP-1a and BgRAP-1c were found increased through N-terminal truncation. Taken together, our data suggest divergent roles for the 3 BgRAP-1s in the merozoite stage of B. gibsoni.
authors.affiliation: National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido 080-8555, Japan.
authors.forename: M A
authors.initials: MA
authors.lastname: Terkawi
authors.forename: A
authors.initials: A
authors.lastname: Amornthep
authors.forename: H
authors.initials: H
authors.lastname: Ooka
authors.forename: G
authors.initials: G
authors.lastname: Aboge
authors.forename: H
authors.initials: H
authors.lastname: Jia
authors.forename: Y-K
authors.initials: YK
authors.lastname: Goo
authors.forename: B
authors.initials: B
authors.lastname: Nelson
authors.forename: J
authors.initials: J
authors.lastname: Yamagishi
authors.forename: Y
authors.initials: Y
authors.lastname: Nishikawa
authors.forename: I
authors.initials: I
authors.lastname: Igarashi
authors.forename: S-I
authors.initials: SI
authors.lastname: Kawazu
authors.forename: K
authors.initials: K
authors.lastname: Fujisaki
authors.forename: X
authors.initials: X
authors.lastname: Xuan
mesh_terms: D000818:Animals; D001403:Babesia / Q000145:classification / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D001404:Babesiosis / Q000175:diagnosis / Q000469:parasitology / Q000662:veterinary*; D015153:Blotting, Western; D016054:DNA, Protozoan / Q000032:analysis; D004283:Dog Diseases / Q000175:diagnosis* / Q000469:parasitology; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D051379:Mice; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D017422:Sequence Analysis, DNA
chemical_list: D016054:DNA, Protozoan; D015800:Protozoan Proteins; C097551:rhoptry-associated protein-1, Babesia