Confocal microscopic findings of cysteine protease calpain in Plasmodium falciparum.
Pf-calpain, a cysteine protease of Plasmodium falciparum, is believed to be one of the central mediators for essential parasitic activity. However, the roles of calpain on parasitic activity have not been determined in P. falciparum. In the present study, the localization of Pf-calpain was investigated using polyclonal antibodies (anti-Pf-calpain antibody A and B) against peptides that distinguished it from human calpain-7 and rat calpain-10 protein. Recombinant Pf-calpain (rPf-calpain) was identified as a 46 kDa protein using an anti-Pf-calpain antibody A, which can recognize the Pf-calpain binding site. Confocal microscopy revealed calpain within cytoplasmic localized parasites in the erythrocytic cycle. The findings suggested that the expression of Pf-calpain would be proportional to all different parasites in the erythrocytic cycle. On the other hand, anti-human calpain-7 antibody detected Pf-calpain in schizonts, and the immunofluorescence was stronger than with anti-rat calpain-10 antibody. However, the antibodies reacted with calpains in human red blood cells. These results show that anti-Pf-calpain antibody A and B specifically recognize only Pf-calpain. Taken together, the results suggest that Pf-calpain is expressed in all erythrocytic stages. In particular, the expression of Pf-calpain is increased much more when the late ring matures into the early trophozoite. Moreover, anti-Pf-calpain antibody A and B against synthetic peptides of the catalytic domain of Pf-calpain are useful to specifically detect Pf-calpain in all erythrocytic stages, while human and rat calpain antibody are not useful.
Choi
Yun-Young
YY
Department of Infection Biology, Zoonosis Research Center, Wonkwang University School of Medicine, Chonbuk 570-749, South Korea.
Jung
Suk-Yul
SY
Cho
Pyo Yun
PY
Soh
Byoung Yul
BY
Zheng
Bing
B
Kim
Sung Yeon
SY
Park
Kie-In
KI
Park
Hyun
H
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002154:Calpain / Q000032:analysis* / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D006801:Humans; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000235:genetics; D051381:Rats; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D002154:Calpain