Conserved cysteine residues in malaria chorismate synthase indicate their important role in protein structure and function

161-168Predicted protein sequences of Chorismate synthase (Cs) from different Plasmodium species have shown high number of conserved cysteine residues when compared to the predicted Cs protein sequences from other bacterial, fungal or plant species. To better understand the structure and function of malaria Cs, we have cloned, expressed and purified recombinant Cs from Plasmodium falciparum (rPfCs) in E.coli. the rPfCs exhibited disulfide linkages as indicated by mobility shifts observed for this protein when compared between non-reduced and reduced-alkylated conditions on SDS-PAGE. Antibodies generated against rPfCs also detected similar mobility shift for the native parasite protein from asexual blood stage culture indicating that the conserved cysteine residues in native parasite Cs play an important role in the protein structure and function

Nano-differential scanning fluorimetry as a tool for the assessment of refolded antibody fragments: a case study for anti-Pfs25 single-chain antibodies

Differential Scanning Fluorimetry (DSF) is a valuable and versatile tool in the field of protein and antibody stability studies, providing valuable insights into their thermal stability, and aiding in the optimization of experimental conditions for various applications. Antibody fragments provide armamentarium for researchers and practitioners across various fields, enabling advancements in medicine, diagnostics, research, and industrial applications. In the present study, single-chain antibodies derived from mAb 4B7 and mAb 1245 were expressed in E. coli, refolded, and checked for their binding with the antigen - Pfs25 (transmission-blocking malaria vaccine candidate). We used nano-DSF as a valuable tool to assess thermal stability, consequently aiding in predicting the correct folding of single-chain antibodies.. Employing nano-DSF as a checkpoint enables the determination of whether to proceed with functional and binding studies on the refolded single-chain antibodies