- Introduction
- Materials and Methods
- Serum samples.
- Depletion of albumin and IgG.
- Concentration of pooled serum from multiple depletions.
- Buffer exchange.
- First dimension separation.
- Second dimension separation.
- Silver staining.
- Purification of total IgG.
- Western blot.
- Colloidal blue stain.
- Protein digestion, mass spectrometry analyses and database search.
- Results
- Discussion
- Conclusion
ProCite
RefWorks
Reference Manager
BibTeX
Zotero
EndNote
-
1.
World Health Organization, 2013. World Malaria Report. Available at: http://www.who.int/malaria/publications/world_malaria_report_2013/report/en/. Accessed February 24, 2014.
-
2.
Gelhaus C, Fritsch J, Krause E, Leippe M, 2005. Fractionation and identification of proteins by 2-DE and MS: towards a proteomic analysis of Plasmodium falciparum. Proteomics 5: 4213–4222.
-
3.
Parra ME, Evans CB, Taylor DW, 1991. Identification of Plasmodium falciparum histidine-rich protein 2 in the plasma of humans with malaria. J Clin Microbiol 29: 1629–1634.
-
4.
Shevchenko A, Wilm M, Vorm O, Mann M, 1996. Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal Chem 68: 850–858.
-
5.
Ahmed N, Barker G, Oliva K, Garfin D, Talmadge K, Georgiou H, Quinn M, Rice G, 2003. An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum. Proteomics 3: 1980–1987.
-
6.
Wiria AE, Prasetyani MA, Hamid F, Wammes LJ, Lell B, Ariawan I, Uh HW, Wibowo H, Djuardi Y, Wahyuni S, Sutanto I, May L, Luty AJ, Verweij JJ, Sartono E, Yazdanbakhsh M, Supali T, 2010. Does treatment of intestinal helminth infections influence malaria? Background and methodology of a longitudinal study of clinical, parasitological and immunological parameters in Nangapanda, Flores, Indonesia (ImmunoSPIN Study). BMC Infect Dis 10: 77.
-
7.
MALDI SYNAPT G2 HDMS, 2010. System Overview and Maintenance Guide. 2010. Waters Rev B: 3–10, 3–13.
-
8.
Smit S, Stoychev S, Louw AI, Birkholtz LM, 2010. Proteomic profiling of Plasmodium falciparum through improved, semiquantitative two-dimensional gel electrophoresis. J Proteome Res 9: 2170–2181.
-
9.
Florens L, Washburn MP, Raine JD, Anthony RM, Grainger M, Haynes JD, Moch JK, Muster N, Sacci JB, Tabb DL, Witney AA, Wolters D, Wu Y, Gardner MJ, Holder AA, Sinden RE, Yates JR, Carucci DJ, 2002. A proteomic view of the Plasmodium falciparum life cycle. Nature 419: 520–526.
-
10.
Acharya P, Pallavi R, Chandran S, Chakravarti H, Middha S, Acharya J, Kochar S, Kochar D, Subudhi A, Boopathi AP, Garg S, Das A, Tatu U, 2009. A glimpse into the clinical proteome of human malaria parasites Plasmodium falciparum and Plasmodium vivax. Proteomics Clin Appl 3: 1314–1325.
-
11.
Vignali M, Armour CD, Chen J, Morrison R, Castle JC, Biery MC, Bouzek H, Moon W, Babak T, Fried M, Raymond CK, Duffy PE, 2011. NSR-seq transcriptional profiling enables identification of a gene signature of Plasmodium falciparum parasites infecting children. J Clin Invest 121: 1119–1129.
-
12.
Howard RJ, Lyon JA, Uni S, Saul AJ, Aley SB, Klotz F, Panton LJ, Sherwood JA, Marsh K, Aikawa M, 1987. Transport of an Mr approximately 300,000 Plasmodium falciparum protein (Pf EMP 2) from the intraerythrocytic asexual parasite to the cytoplasmic face of the host cell membrane. J Cell Biol 104: 1269–1280.
-
13.
Kun JF, Waller KL, Coppel RL, 1999. Plasmodium falciparum: structural and functional domains of the mature-parasite-infected erythrocyte surface antigen. Exp Parasitol 91: 258–267.
-
14.
Maier AG, Cooke BM, Cowman AF, Tilley L, 2009. Malaria parasite proteins that remodel the host erythrocyte. Nat Rev Microbiol 7: 341–354.
-
15.
Waller KL, Nunomura W, An X, Cooke BM, Mohandas N, Coppel RL, 2003. Mature parasite-infected erythrocyte surface antigen (MESA) of Plasmodium falciparum binds to the 30-kDa domain of protein 4.1 in malaria-infected red blood cells. Blood 102: 1911–1914.
-
16.
Magowan C, Coppel RL, Lau AO, Moronne MM, Tchernia G, Mohandas N, 1995. Role of the Plasmodium falciparum mature-parasite-infected erythrocyte surface antigen (MESA/PfEMP-2) in malarial infection of erythrocytes. Blood 86: 3196–3204.
-
17.
Biswas S, Tomar D, Rao DN, 2005. Investigation of the kinetics of histidine-rich protein 2 and of the antibody responses to this antigen, in a group of malaria patients from India. Ann Trop Med Parasitol 99: 553–562.
| Past two years | Past Year | Past 30 Days | |
|---|---|---|---|
| Abstract Views | 183 | 164 | 25 |
| Full Text Views | 298 | 7 | 0 |
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Mature Erythrocyte Surface Antigen Protein Identified in the Serum of Plasmodium falciparum-Infected Patients
Nurul Shazalina Zainudin
Nurul Shazalina Zainudin
Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia, Penang, Malaysia; Kuching Health Office, Ministry of Health, Kuching, Sarawak, Malaysia; Malaysia Genome Institute, Jalan Bangi, 43000 Kajang, Selangor, Malaysia
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Nurulhasanah Othman
Nurulhasanah Othman
Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia, Penang, Malaysia; Kuching Health Office, Ministry of Health, Kuching, Sarawak, Malaysia; Malaysia Genome Institute, Jalan Bangi, 43000 Kajang, Selangor, Malaysia
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Jamail Muhi
Jamail Muhi
Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia, Penang, Malaysia; Kuching Health Office, Ministry of Health, Kuching, Sarawak, Malaysia; Malaysia Genome Institute, Jalan Bangi, 43000 Kajang, Selangor, Malaysia
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Asmahani Azira Abdu Sani
Asmahani Azira Abdu Sani
Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia, Penang, Malaysia; Kuching Health Office, Ministry of Health, Kuching, Sarawak, Malaysia; Malaysia Genome Institute, Jalan Bangi, 43000 Kajang, Selangor, Malaysia
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Rahmah Noordin
Rahmah Noordin
Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia, Penang, Malaysia; Kuching Health Office, Ministry of Health, Kuching, Sarawak, Malaysia; Malaysia Genome Institute, Jalan Bangi, 43000 Kajang, Selangor, Malaysia
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This study was performed to identify circulating Plasmodium falciparum proteins in patient serum, which may be useful as diagnostic markers. Depletion of highly abundant proteins from each pooled serum sample obtained from P. falciparum-infected patients and healthy individuals was performed using the Proteoseek Antibody-Based Albumin/IgG Removal Kit (Thermo Scientific, Rockford, IL). In analysis 1, the depleted serum was analyzed directly by NanoLC-MS/MS. In analysis 2, the depleted serum was separated by two-dimensional electrophoresis followed by western blot analysis. Subsequently, the selected band was analyzed by NanoLC-MS/MS. The result of analysis 1 revealed the presence of two mature erythrocyte surface antigen (MESA) proteins and chloroquine resistance transporter protein (PfCRT). In addition, analysis 2 revealed an antigenic 75-kDa band when the membrane was probed with purified IgG from the pooled serum obtained from P. falciparum-infected patients. MS/MS analysis of this protein band revealed fragments of P. falciparum MESA proteins. Thus, in this study, two different analyses revealed the presence of Plasmodium MESA protein in pooled serum from malaria patients; thus, this protein should be further investigated to determine its usefulness as a diagnostic marker.
Author Notes
Financial support: This research was funded by USM RU grant, No. 1001/CIPPM/812046 and EU grant No. INCO-CT-2006-031714.
Authors' addresses: Nurul Shazalina Zainudin, Nurulhasanah Othman, and Rahmah Noordin, Institute for Research in Molecular Medicine, Universiti Sains Malaysia, 11800 Penang, Malaysia, E-mails: shazalina87@gmail.com, nho80@yahoo.co.uk, and rahmah8485@gmail.com. Jamail Muhi, Sabah Health Office, Kota Kinabalu, Sabah, Malaysia, E-mail: jamail@moh.gov.my. Asmahani Azira Abdu Sani, Malaysia Genome Institute, Jalan Bangi, 43000 Kajang, Selangor, Malaysia, E-mail: azira@genomemalaysia.gov.my.
- Introduction
- Materials and Methods
- Serum samples.
- Depletion of albumin and IgG.
- Concentration of pooled serum from multiple depletions.
- Buffer exchange.
- First dimension separation.
- Second dimension separation.
- Silver staining.
- Purification of total IgG.
- Western blot.
- Colloidal blue stain.
- Protein digestion, mass spectrometry analyses and database search.
- Results
- Discussion
- Conclusion
ProCite
RefWorks
Reference Manager
BibTeX
Zotero
EndNote
-
1.
World Health Organization, 2013. World Malaria Report. Available at: http://www.who.int/malaria/publications/world_malaria_report_2013/report/en/. Accessed February 24, 2014.
-
2.
Gelhaus C, Fritsch J, Krause E, Leippe M, 2005. Fractionation and identification of proteins by 2-DE and MS: towards a proteomic analysis of Plasmodium falciparum. Proteomics 5: 4213–4222.
-
3.
Parra ME, Evans CB, Taylor DW, 1991. Identification of Plasmodium falciparum histidine-rich protein 2 in the plasma of humans with malaria. J Clin Microbiol 29: 1629–1634.
-
4.
Shevchenko A, Wilm M, Vorm O, Mann M, 1996. Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal Chem 68: 850–858.
-
5.
Ahmed N, Barker G, Oliva K, Garfin D, Talmadge K, Georgiou H, Quinn M, Rice G, 2003. An approach to remove albumin for the proteomic analysis of low abundance biomarkers in human serum. Proteomics 3: 1980–1987.
-
6.
Wiria AE, Prasetyani MA, Hamid F, Wammes LJ, Lell B, Ariawan I, Uh HW, Wibowo H, Djuardi Y, Wahyuni S, Sutanto I, May L, Luty AJ, Verweij JJ, Sartono E, Yazdanbakhsh M, Supali T, 2010. Does treatment of intestinal helminth infections influence malaria? Background and methodology of a longitudinal study of clinical, parasitological and immunological parameters in Nangapanda, Flores, Indonesia (ImmunoSPIN Study). BMC Infect Dis 10: 77.
-
7.
MALDI SYNAPT G2 HDMS, 2010. System Overview and Maintenance Guide. 2010. Waters Rev B: 3–10, 3–13.
-
8.
Smit S, Stoychev S, Louw AI, Birkholtz LM, 2010. Proteomic profiling of Plasmodium falciparum through improved, semiquantitative two-dimensional gel electrophoresis. J Proteome Res 9: 2170–2181.
-
9.
Florens L, Washburn MP, Raine JD, Anthony RM, Grainger M, Haynes JD, Moch JK, Muster N, Sacci JB, Tabb DL, Witney AA, Wolters D, Wu Y, Gardner MJ, Holder AA, Sinden RE, Yates JR, Carucci DJ, 2002. A proteomic view of the Plasmodium falciparum life cycle. Nature 419: 520–526.
-
10.
Acharya P, Pallavi R, Chandran S, Chakravarti H, Middha S, Acharya J, Kochar S, Kochar D, Subudhi A, Boopathi AP, Garg S, Das A, Tatu U, 2009. A glimpse into the clinical proteome of human malaria parasites Plasmodium falciparum and Plasmodium vivax. Proteomics Clin Appl 3: 1314–1325.
-
11.
Vignali M, Armour CD, Chen J, Morrison R, Castle JC, Biery MC, Bouzek H, Moon W, Babak T, Fried M, Raymond CK, Duffy PE, 2011. NSR-seq transcriptional profiling enables identification of a gene signature of Plasmodium falciparum parasites infecting children. J Clin Invest 121: 1119–1129.
-
12.
Howard RJ, Lyon JA, Uni S, Saul AJ, Aley SB, Klotz F, Panton LJ, Sherwood JA, Marsh K, Aikawa M, 1987. Transport of an Mr approximately 300,000 Plasmodium falciparum protein (Pf EMP 2) from the intraerythrocytic asexual parasite to the cytoplasmic face of the host cell membrane. J Cell Biol 104: 1269–1280.
-
13.
Kun JF, Waller KL, Coppel RL, 1999. Plasmodium falciparum: structural and functional domains of the mature-parasite-infected erythrocyte surface antigen. Exp Parasitol 91: 258–267.
-
14.
Maier AG, Cooke BM, Cowman AF, Tilley L, 2009. Malaria parasite proteins that remodel the host erythrocyte. Nat Rev Microbiol 7: 341–354.
-
15.
Waller KL, Nunomura W, An X, Cooke BM, Mohandas N, Coppel RL, 2003. Mature parasite-infected erythrocyte surface antigen (MESA) of Plasmodium falciparum binds to the 30-kDa domain of protein 4.1 in malaria-infected red blood cells. Blood 102: 1911–1914.
-
16.
Magowan C, Coppel RL, Lau AO, Moronne MM, Tchernia G, Mohandas N, 1995. Role of the Plasmodium falciparum mature-parasite-infected erythrocyte surface antigen (MESA/PfEMP-2) in malarial infection of erythrocytes. Blood 86: 3196–3204.
-
17.
Biswas S, Tomar D, Rao DN, 2005. Investigation of the kinetics of histidine-rich protein 2 and of the antibody responses to this antigen, in a group of malaria patients from India. Ann Trop Med Parasitol 99: 553–562.
| Past two years | Past Year | Past 30 Days | |
|---|---|---|---|
| Abstract Views | 183 | 164 | 25 |
| Full Text Views | 298 | 7 | 0 |
| PDF Downloads | 67 | 8 | 0 |