A major surface antigen of procyclic stage Trypanosoma congolense.
Five monoclonal antibodies (mAb) were raised that bound to the surface of procyclic stage Trypanosoma congolense with high intensity in immunofluorescence. Immunoblot analysis of trypanosome lysates using 3 of these mAb revealed a diffuse SDS-PAGE band of 36-40 kDa. The purified antigen did not react with Coomassie Blue or silver stains, but did stain blue with Stains-all, indicating acidity. For the one mAb tested, the epitope was periodate-sensitive and therefore probably glycan. Although this antigen shares properties with procyclin/PARP, which forms a surface coat on procyclic Trypanosoma brucei, a search in T. congolense for homologues of a procyclin/PARP gene revealed only non-coding sequence of partial similarity. Using a differential screen, a procyclic stage T. congolense cDNA clone was isolated that encoded a putative 256-amino acid protein containing 2 peptides chemically sequenced independently by Beecroft et al. [36]. The protein, termed glutamate and alanine-rich protein (GARP), has potential hydrophobic leader and tail sequences (the latter with potential for replacement by a glycosyl phosphoinositol anchor) and no potential N-linked glycosylation sites. It has no significant sequence homology with known proteins. Antibodies against a translational fusion of GARP bound specifically in Western blots to a band very similar to that detected by the mAb and also to the purified antigen. Immunogold electron microscopy revealed a dense packing of the antigen on the cell surface. It appears that procyclic T. brucei and T. congolense have major surface proteins with structural analogy, but with no sequence homology.
Bayne
R A
RA
Wellcome Unit of Molecular Parasitology and Institute of Genetics, University of Glasgow, Scotland, UK.
Kilbride
E A
EA
Lainson
F A
FA
Tetley
L
L
Barry
J D
JD
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan / Q000032:analysis* / Q000096:biosynthesis*; D000954:Antigens, Surface / Q000032:analysis* / Q000096:biosynthesis*; D001483:Base Sequence; D003062:Codon; D017931:DNA Primers; D016054:DNA, Protozoan / Q000302:isolation & purification / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D015698:Genomic Library; D015151:Immunoblotting; D008969:Molecular Sequence Data; D012326:RNA Splicing; D012333:RNA, Messenger / Q000302:isolation & purification / Q000378:metabolism; D016053:RNA, Protozoan / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D014348:Trypanosoma congolense / Q000276:immunology / Q000502:physiology*
D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D000954:Antigens, Surface; D003062:Codon; D017931:DNA Primers; D016054:DNA, Protozoan; D012333:RNA, Messenger; D016053:RNA, Protozoan