Cloning and sxpression of mitochondrial heat shock protein 70 of Trypanosoma congolense and potential use as a diagnostic antigen.
The ability to use mitochondrial heat shock protein 70 (MTP) of Trypanosoma congolense as a diagnostic antigen was examined. One cDNA clone was obtained by immunoscreening of a T. congolense procyclic form (PCF) cDNA library with monoclonal antibody (MAb) 10F9. The cDNA clone contained an open reading frame of 1,977 bp encoding a polypeptide consisting of 659 amino acids. Southern blotting analysis indicated that there were at least three copies of the MTP gene in the haploid genome. Interference of the MTP RNA resulted in complete inhibition, which indicated that MTP is essential at the PCF stage. Northern and Western blotting analyses revealed that MTP is expressed both in the bloodstream form (BSF) and in PCF. The B-cell epitope recognized by MAb 10F9 was located within 206 amino acids from the C terminus. Depending on the conditions of protein extraction, MTP was cleaved into smaller polypeptides by endogenous proteases. However, the C-terminal epitope of MTP was preserved with a high degree of antigenicity, even after cleavage. Antibody detection by enzyme-linked immunosorbent assay with the truncated recombinant MTP revealed that anti-MTP antibodies exist in experimentally infected mouse sera. Thus, MTP may be useful as an antigen for the serodiagnosis of primary T. congolense infection.
Bannai
Hiroshi
H
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
Sakurai
Tatsuya
T
Inoue
Noboru
N
Sugimoto
Chihiro
C
Igarashi
Ikuo
I
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000953:Antigens, Protozoan / Q000235:genetics* / Q000276:immunology; D015152:Blotting, Northern; D015139:Blotting, Southern; D015153:Blotting, Western; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D005260:Female; D015723:Gene Library; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics* / Q000276:immunology*; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D014348:Trypanosoma congolense / Q000235:genetics* / Q000276:immunology; D014353:Trypanosomiasis, African / Q000276:immunology
D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D018840:HSP70 Heat-Shock Proteins