1.Identification of Immunodominant B-cell Epitope Regions of Reticulocyte Binding Proteins in Plasmodium vivax by Protein Microarray Based Immunoscreening.
Jin Hee HAN ; Jian LI ; Bo WANG ; Seong Kyun LEE ; Myat Htut NYUNT ; Sunghun NA ; Jeong Hyun PARK ; Eun Taek HAN
The Korean Journal of Parasitology 2015;53(4):403-411
Plasmodium falciparum can invade all stages of red blood cells, while Plasmodium vivax can invade only reticulocytes. Although many P. vivax proteins have been discovered, their functions are largely unknown. Among them, P. vivax reticulocyte binding proteins (PvRBP1 and PvRBP2) recognize and bind to reticulocytes. Both proteins possess a C-terminal hydrophobic transmembrane domain, which drives adhesion to reticulocytes. PvRBP1 and PvRBP2 are large (> 326 kDa), which hinders identification of the functional domains. In this study, the complete genome information of the P. vivax RBP family was thoroughly analyzed using a prediction server with bioinformatics data to predict B-cell epitope domains. Eleven pvrbp family genes that included 2 pseudogenes and 9 full or partial length genes were selected and used to express recombinant proteins in a wheat germ cell-free system. The expressed proteins were used to evaluate the humoral immune response with vivax malaria patients and healthy individual serum samples by protein microarray. The recombinant fragments of 9 PvRBP proteins were successfully expressed; the soluble proteins ranged in molecular weight from 16 to 34 kDa. Evaluation of the humoral immune response to each recombinant PvRBP protein indicated a high antigenicity, with 38-88% sensitivity and 100% specificity. Of them, N-terminal parts of PvRBP2c (PVX_090325-1) and PvRBP2 like partial A (PVX_090330-1) elicited high antigenicity. In addition, the PvRBP2-like homologue B (PVX_116930) fragment was newly identified as high antigenicity and may be exploited as a potential antigenic candidate among the PvRBP family. The functional activity of the PvRBP family on merozoite invasion remains unknown.
Epitopes, B-Lymphocyte/*chemistry/genetics/*immunology
;
Female
;
Humans
;
Immunodominant Epitopes/chemistry/genetics/*immunology
;
Malaria, Vivax/immunology/*parasitology
;
Middle Aged
;
Plasmodium vivax/chemistry/genetics/*immunology
;
Protein Structure, Tertiary
;
Protozoan Proteins/chemistry/genetics/*immunology
;
Reticulocytes/*parasitology
2.Usefulness of the recombinant liver stage antigen-3 for an early serodiagnosis of Plasmodium falciparum infection.
Hyeong Woo LEE ; Sung Ung MOON ; Hye Sun RYU ; Yeon Joo KIM ; Shin Hyeong CHO ; Gyung Tae CHUNG ; Khin LIN ; Byoung Kuk NA ; Yoon KONG ; Kyung Suk CHUNG ; Tong Soo KIM
The Korean Journal of Parasitology 2006;44(1):49-54
In order to develop tools for an early serodiagnosis of Plasmodium falciparum infection, we evaluated the usefulness of P. falciparum liver stage antigen-3 (LSA-3) as a serodiagnostic antigen. A portion of LSA-3 gene was cloned, and its recombinant protein (rLSA-3) was expressed in Escherichia coli and purified by column chromatography. The purified rLSA-3 and 120 test blood/serum samples collected from inhabitants in malaria-endemic areas of Mandalay, Myanmar were used for this study. In microscopic examinations of blood samples, P. falciparum positive rate was 39.1% (47/120) in thin smear trials, and 33.3% (40/120) in thick smear trials. Although the positive rate associated with the rLSA-3 (30.8%) was lower than that of the blood stage antigens (70.8%), rLSA-3 based enzyme-linked immunosorbent assay could detect 12 seropositive cases (10.0%), in which blood stage antigens were not detected. These results indicate that the LSA-3 is a useful antigen for an early serodiagnosis of P. falciparum infection.
Recombinant Proteins/biosynthesis/genetics/*immunology
;
Plasmodium vivax/isolation & purification
;
Plasmodium falciparum/*immunology
;
Molecular Sequence Data
;
Malaria, Falciparum/blood/*diagnosis
;
Humans
;
Genes, Protozoan/genetics/immunology
;
Fluorescent Antibody Technique, Direct/methods
;
Escherichia coli/genetics
;
Enzyme-Linked Immunosorbent Assay/methods
;
Early Diagnosis
;
DNA, Protozoan/chemistry
;
DNA Primers/chemistry
;
Cloning, Molecular/methods
;
Base Sequence
;
Antigens, Protozoan/biosynthesis/chemistry/genetics/*immunology
;
Animals
;
Amino Acid Sequence
3.Efficacy of the Merozoite Surface Protein 1 of Plasmodium Vivax as an Antigen for ELISA to Diagnose Malaria.
Yong Man KIM ; Hyun Ah HWANG ; Woo Sang YUN ; Suk Il KIM ; Kil Whoan LEE ; Seung Kyu PARK ; Young Jin LEE ; Tae Kyun KIM ; Chansuda WONGSRICHANALAI ; Judy A SAKANARI ; Hyun PARK
Yonsei Medical Journal 2004;45(1):129-134
Malaria is still a major health problem in Thailand and its incidence is currently rising in Korea. To identify a useful antigen for the diagnosis of malaria patients, a cDNA expression library from malaria parasites was constructed and screened out immunologically. One clone was selected in view of its predominant reactivity with the patient sera. The recombinant malaria parasite antigen (Pv30) with 27 kDa as a C-terminal His-tag fusion protein that was produced in Escherichia coli was identified through immunoblot analysis. The deduced amino acid sequence had the sequence homology with the merozoite surface protein 1 (MSP1) genes of Plasmodium falciparum and P. yoelii, each by 41% and 42%, respectively. Measurement of serum IgG and IgM antibody to Pv30 by enzyme-linked immunosorbent assay (ELISA) was evaluated as a serodiagnostic test for malaria patients in Thailand (endemic area) and Korea (recently reemerging area). The sensitivity of P. vivax, P. falciparum, and P. malariae was 96.3% (26 /27), 90.6% (29/32), and 100% (6/6), respectively, and the specificity was 63.5% (40/63) in Thailand samples. The sensitivity of P. vivax was 98.8% (88/89), and the specificity was 96.6% (86/89) in Korean samples. Pv30 appears to be a good and reliable recombinant antigen for serodiagonosis of malaria in a nonendemic area.
Amino Acid Sequence
;
Animals
;
Antibodies, Protozoan
;
Enzyme-Linked Immunosorbent Assay/*methods
;
Human
;
Korea
;
Malaria, Vivax/*diagnosis/immunology
;
Merozoite Surface Protein 1/*analysis/genetics/immunology
;
Molecular Sequence Data
;
Plasmodium vivax/chemistry/immunology/*isolation & purification
;
Sensitivity and Specificity
;
Serologic Tests
;
Support, Non-U.S. Gov't