Affiliation:
1. Division of Parasitology and Tropical Veterinary Medicine, Department of Infectious Diseases and Immunology, Utrecht University, 3508 TD, Utrecht, The Netherlands
Abstract
ABSTRACT
Apical membrane antigen 1 (AMA-1) is a micronemal protein secreted to the surface of merozoites of
Plasmodium
species and
Toxoplasma gondii
tachyzoites in order to fulfill an essential but noncharacterized function in host cell invasion. Here we describe cloning and characterization of a
Babesia bovis
AMA-1 homologue designated BbAMA-1. The overall level of similarity of BbAMA-1 to
P. falciparum
AMA-1 was low (18%), but characteristic features like a transmembrane domain near the C terminus, a predicted short cytoplasmic C-terminal sequence with conserved sequence properties, and an extracellular domain containing 14 conserved cysteine residues putatively involved in disulfide bridge formation are typical of AMA-1. Rabbit polyclonal antisera were raised against three synthetic peptides derived from the N-terminal region and domains II and III of the putative extracellular domain and were shown to recognize specifically recombinant BbAMA-1 expressed in
Escherichia coli
. Immunofluorescence microscopy showed that there was labeling of the apical half of merozoites with these antisera. Preincubation of free merozoites with all three antisera reduced the efficiency of invasion of erythrocytes by a maximum of 65%. Antisera raised against the N-terminal peptide detected a 82-kDa protein on Western blots and a 69-kDa protein in the supernatant that was harvested after in vitro invasion, suggesting that proteolytic processing and secretion take place during or shortly after invasion. A combination of two-dimensional Western blotting and metabolic labeling allowing direct identification of spots reacting with the BbAMA-1 peptide antisera together with the very low silver staining intensity of these spots indicated that very low levels of BbAMA-1 are present in
Babesia
merozoites.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
Cited by
100 articles.
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