Abstract
Plasmodium lactate dehydrogenase (pLDH), owing to unique structural and kinetic properties, is a well known target for antimalarial compounds. To explore a new approach for high level soluble expression of Plasmodium falciparum lactate dehydrogenase ( PfLDH) in E. coli, PfLDH encoding sequence was cloned into pQE-30 Xa vector. When transformed E. coli SG13009 cells were induced at 37 °C with 0.5 mM isopropyl β- d-thiogalactoside (IPTG) concentration, the protein was found to be exclusively associated with inclusion bodies. By reducing cell growth temperature to 15 °C and IPTG concentration to 0.25 mM, it was possible to get approximately 82% of expressed protein in soluble form. Recombinant PfLDH (r PfLDH) was purified to homogeneity yielding 18 mg of protein/litre culture. r PfLDH was found to be biologically active with specific activity of 453.8 μmol/min/mg. The enzyme exhibited characteristic reduced substrate inhibition and enhanced k cat [(3.2 ± 0.02) × 10 4] with 3-acetylpyridine adenine dinucleotide (APAD +). The procedure described in this study may provide a reliable and simple method for production of large quantities of soluble and biologically active PfLDH.
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