Cloning and expression of Plasmodium falciparum lactate dehydrogenase (PfLDH) in Escherichia coli BL21(DE3)

  • Fifi Fitriyah Masduki Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung
  • Yeni Hotimah Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Bandung
  • Rosalia Rani Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Bandung
  • Arsyam Mawardi Department of Biology, Faculty of Mathematics and Natural Sciences, Cendrawasih University, Jayapura
  • Euniche R.P.F. Ramandey Department of Biology, Faculty of Mathematics and Natural Sciences, Cendrawasih University, Jayapura
  • Azzania Fibriani School of Life Sciences and Technology, Institut Teknologi Bandung, Bandung
  • Sony Suhandono School of Life Sciences and Technology, Institut Teknologi Bandung, Bandung
Keywords: Plasmodium falciparum, lactate dehydrogenase, Malaria Rapid Diagnostic Test (RDT), recombinant protein

Abstract

Background: Immediate and accurate diagnosis of malaria is essential for effective control of this disease. Immunochromatographic based rapid diagnostic tests (RDTs)  are economical, simple to perform, and provide results in a relative short time, can be useful to assist effective management of malaria. The commercially available malaria RDT in Indonesia is still imported. Therefore, an effort to produce malaria RDT independently is necessary. One of the biomarkers used in RDTs is Plasmodium lactate dehydrogenase pLDH. The production and accumulation of pLDH during asexual stage or blood-stage in all human infected malaria parasites can be used to indicate parasites viability, which is correlated with the number of parasites present in the plasma of infected patients.  

Objective: The aim of this research is to produce recombinant PfLDH in Escherichia coli BL21(DE3).

MethodsPfLDH gene was cloned into pET30a expression vector to obtain a 6.2 kbp recombinant plasmid pET30a-PfLDH. E. coli BL21(DE3) was transformed with pET30a-PfLDH using the heat shock method. Then, E. coli BL21(DE3)- pET30a-PfLDH was cultured in LB broth containing 50 mg/mL kanamycin and was induced by 1mM IPTG at 37oC.

Results: SDS-PAGE and Western Blot analysis showed that recombinant PfLDH was expressed with molecular mass ~30 kDa.

Conclusion: Recombinant PfLDH is expressed in E. coli BL21(DE3) and can be used in further research for producing rPfLDH as a biomarker for malaria RDT development.

References

World Health Organization. World Malaria Report 2019 [Internet]. 2019. 1-210 p. Available from: https://www.who.int/malaria/publications/world-malaria-report-2019/en/

Chanda P, Hamainza B, Mulenga S, Chalwe V, Msiska C, Chizema-Kawesha E. Early results of integrated malaria control and implications for the management of fever in under-five children at a peripheral health facility: a case study of Chongwe rural health centre in Zambia. Malar J. 2009;8:49. https://doi.org/10.1186/1475-2875-8-49

Makler MT, Hinrichs DJ. Measurement of the lactate dehydrogenase activity of Plasmodium falciparum as an assessment of parasitemia. Am. J. Trop. Med. Hyg. 1993;48:205-210. https://doi.org/10.4269/ajtmh.1993.48.205

Piper R1, Lebras J, Wentworth L, Hunt-Cooke A, Houzé S, Chiodini P, et al. Immunocapture diagnostic assays for malaria using Plasmodium lactate dehydrogenase (pLDH). Am J Trop Med Hyg. 1999;60(1):109-18. https://doi.org/10.4269/ajtmh.1999.60.109

Sherman IW. Heterogeneity of lactic dehydrogenase in avian malaria (Plasmodium lophurae). J Exp Med. 1961;114: 1049-1062. https://doi.org/10.1084/jem.114.6.1049

Das S, Dash HR. Microbial biotechnology-A laboratory manual for bacterial systems. Springer. 2015 https://doi.org/10.1007/978-81-322-2095-4

Schägger, H., Tricine-SDS-PAGE. Nature Protocols. 2006;1(1):16-22. https://doi.org/10.1038/nprot.2006.4

Murray CK, Gasser RA Jr, Magill AJ, Miller RS. Update on rapid diagnostic testing for malaria. Clin Microbiol Rev. 2008;21(1):97-110. https://doi.org/10.1128/CMR.00035-07

Jain P, Chakma B, Patra S, Goswami P. Potential biomarkers and their applications for rapid and reliable detection of malaria. Biomed Res Int. 2014;2014:852645. https://doi.org/10.1155/2014/852645

SnapGene software (from GSL Biotech; available at snapgene.com)

Published
2019-12-31
How to Cite
Masduki, F. F., Hotimah, Y., Rani, R., Mawardi, A., Ramandey, E. R., Fibriani, A., & Suhandono, S. (2019). Cloning and expression of Plasmodium falciparum lactate dehydrogenase (PfLDH) in Escherichia coli BL21(DE3). Acta Biochimica Indonesiana, 2(2), 52-57. https://doi.org/10.32889/actabioina.v2i2.39