Refolding and characterization of a diabody against Pfs25, a vaccine candidate of Plasmodium falciparum

Jagannath, D. K., Valiyaparambil, A., Viswanath, V. K., Hurakadli, M. A., Kamariah, N., Jafer, A. C., Patole, C., Pradhan, S., Kumar, N. and Lakshminarasimhan, A. (2022) Refolding and characterization of a diabody against Pfs25, a vaccine candidate of Plasmodium falciparum. Analytical Biochemistry, 655, 114830. (doi: 10.1016/j.ab.2022.114830) (PMID:35944694)

Abstract

Pfs25, a vaccine candidate, expressed on the surface of the malarial parasite, plays an important role in the development of Plasmodium falciparum. 1269, a monoclonal antibody targeting the epidermal growth factor-like domain 1 and epidermal growth factor-like domain 3 of Pfs25, blocks the transmission of parasites in mosquitoes. In this study, we refolded 1269-Db, a dimeric antibody fragment referred as diabody, designed from 1269, with a yield of 3 mg/litre of bacterial culture. Structural integrity of the protein was validated with thermal stability, disulphide bond analysis and glutaraldehyde crosslinking experiments. To evaluate the functionality of 1269-Db, recombinant monomeric MBP-Pfs25 was produced from bacteria. Qualitative binding assays demonstrated that 1269-Db recognized the epitopes on Pfs25 in its native, but not the denatured state. An apparent KD of 2.6 nM was determined for 1269-Db with monomeric MBP-Pfs25, using isothermal titration calorimetry. 1269-Db recognized the periphery of zygotes/ookinetes, demonstrating recognition of Pfs25, expressed on the surface of the parasite. As the established refolding method resulted in a functional diabody, the optimized method pipeline for 1269-Db can potentially facilitate engineering of antibody fragments with desired properties.

• Item Type: Articles
• Additional Information: The authors would like to thank Tata trusts for funding this study.
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Pradhan, Dr Sabyasachi

Creator Roles

Pradhan, S.: Methodology

• Authors: Jagannath, D. K., Valiyaparambil, A., Viswanath, V. K., Hurakadli, M. A., Kamariah, N., Jafer, A. C., Patole, C., Pradhan, S., Kumar, N., and Lakshminarasimhan, A.
• College/School: College of Medical Veterinary and Life Sciences > School of Biodiversity, One Health & Veterinary Medicine
• Journal Name: Analytical Biochemistry
• Publisher: Elsevier
• ISSN: 0003-2697
• ISSN (Online): 1096-0309
• Published Online: 06 August 2022