Cloning and characterisation of Lactate Dehydrogenase Gene from Plasmodium knowlesi in bacterial system

• Main Authors: Ogu Salim, Nurhainis, Ahmad Fuad, Fazia Adyani
• Format: Conference or Workshop Item
• Language: English
• Published: 2016
• Subjects: RA Public aspects of medicine
• Online Access: http://irep.iium.edu.my/97180/1/J3%202016%20BBS%20KK%20Abstract-HB8.pdf; http://irep.iium.edu.my/97180/

Glycolysis is essential for Plasmodium survival during its intra-erythrocytic stage in the human host. As a consequence, enzymes in the glycolytic pathway have been proposed as ideal therapeutic targets for malaria pharmaceuticals. Specifically, lactate dehydrogenase, which is the final enzyme in glycolysis, has been validated as a good drug target. We have cloned and characterised recombinant lactate dehydrogenase from Plasmodium knowlesi in a bacterial system. Synthetic P. knowlesi lactate dehydrogenase (Pk-LDH) gene was obtained from GenScript®. Pk-LDH gene was successfully amplified from the pUC57 vector and a PCR product with the size of 951bp was cloned into pEASY-Blunt E1 expression vector. The ligated product was subsequently transformed into Trans1-T1 Phage Resistant Chemically Competent Cell. A sequence alignment analysis, which was conducted to compare the sequence similarity of Pk-LDH to LDH from other human malaria parasites revealed open reading frame of 316 amino acids of Pk-LDH and showed 97.8% homology to P. vivax LDH and 90% homology to P. malariae, P. falciparum, and P. ovale LDHs, respectively. The purified recombinant Pk-LDH will later be utilised for inhibition studies in future antimalarial drug design and discovery research, specifically for P. knowlesi.