Cloning and expression of mitochondrial malate dehydrogenase of Clonorchis sinensis.

The NAD-dependent mitochondrial malate dehydrogenase (mMDH, EC1.1.1.37) plays pivotal roles in tricarboxylic acid and is crucial for the survival and pathogenecity of parasites. A cDNA, which was identified by high throughput sequencing from the cDNA library constructed from adult Clonorchis sinensis, encoded a putative peptide of 341 amino acids with more than 50% identity with mMDHs from other organisms. The mMDH was expressed in Escherichia coli as the recombinant protein with a GST tag and purified by glutathione-Sepharose 4B column. The recombinant mMDH showed MDH activity of 63.6 U/mg, without lactate dehydrogenase activity and NADPH selectivity. The kinetic constants of recombinant mMDH were determined.