Identification of a serodiagnostic antigen, legumain, by immunoproteomic analysis of excretory-secretory products of Clonorchis sinensis adult worms.
Clonorchis sinensis, the Chinese liver fluke, is the causative agent of clonorchiasis as well as liver and biliary diseases. The excretory-secretory products (ESPs) of the parasites play important roles in host-parasite interactions. In this study, we have investigated the proteome of ESPs obtained from C. sinensis adult worms. Although the full genome database of C. sinensis is not yet available, we have successfully identified 62 protein spots using 2-DE-based mass analysis and EST database of C. sinensis. The proteins identified include detoxification enzymes, such as glutathione S-transferase and thioredoxin peroxidase, myoglobin and a number of cysteine proteases that are expressed abundantly. In order to identify potential targets for the diagnosis and therapy of clonorchiasis, we conducted immunoblot analysis of the ESPs proteome using the sera obtained from clonorchiasis patients and identified legumains and cysteine proteases as antigens present in the ESPs. Although the cysteine proteases were previously reported to elicit antigenicity, the legumains are found herein for the first time as a serological antigen of C. sinensis. To confirm these findings, we expressed recombinant legumain in Escherichia coli and verified that recombinant legumain also functions as a potent antigen against the sera of clonorchiasis patients. Our results illustrate the validity of immuno-proteomic approaches in the identification of serodiagnostic antigens in the parasites.
Ju
Jung-Won
JW
Division of Malaria and Parasitic Diseases, National Institute of Health, Seoul, Korea.
Joo
Hyun-Na
HN
Lee
Myoung-Ro
MR
Cho
Shin-Hyeong
SH
Cheun
Hyeng-Il
HI
Kim
Jung-Yeon
JY
Lee
Young-Hee
YH
Lee
Kwang-Jun
KJ
Sohn
Woon-Mok
WM
Kim
Dong-Min
DM
Kim
Il-Chul
IC
Park
Byoung Chul
BC
Kim
Tong-Soo
TS
D000595:Amino Acid Sequence; D000818:Animals; D000947:Antigens, Helminth / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D003003:Clonorchiasis / Q000175:diagnosis / Q000469:parasitology; D003004:Clonorchis sinensis / Q000201:enzymology* / Q000235:genetics / Q000276:immunology; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D015801:Helminth Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015151:Immunoblotting; D008969:Molecular Sequence Data; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D016415:Sequence Alignment; D012698:Serologic Tests / Q000379:methods*; D000099240:Asparaginyl Endopeptidase
D000947:Antigens, Helminth; D003546:Cysteine Endopeptidases; D015801:Helminth Proteins; D020543:Proteome; D011994:Recombinant Proteins; D000099240:Asparaginyl Endopeptidase