Enzymatic and physico-chemical characteristics of recombinant cMDH and mMDH of Clonorchis sinensis.

The cytosol and mitochondrial malate dehydrogenases (MDHs, EC 1.1.1.37) of Clonorchis sinensis were expressed in Escherichia coli as a fusion protein with a 6xHis and GST tag, respectively. The cytosol MDH of Clonorchis sinensis (Cs-cMDH) has higher resistibility to acid than mitochondrial MDH (Cs-mMDH). The Cs-cMDH also has higher heat resistibility and thermal stability than Cs-mMDH. Although there is only 22.8% identity between the amino acid sequences of Cs-cMDH and Cs-mMDH, they share several conserved residues. There are some differences between the circular dichroism spectra of Cs-cMDH and Cs-mMDH, but they have approximate percentages of helix. 4,4'-Bisdimethylamino diphenylcarbinol can decrease the Cs-mMDH activity but not the Cs-cMDH activity. Paraziquantel, metronidazole and albendazole did not inhibit the enzymes' activity, but adenosine 5'-monophosphate showed competitive inhibition to enzyme, with the Ki for Cs-cMDH and Cs-mMDH being 2.81 and 0.49 mM, respectively.