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.
Zheng
Nancai
N
Center for Disease Control and Prevention of JiangMen City, Guangdong Province, 529000, People's Republic of China.
Huang
Baoming
B
Xu
Jin
J
Huang
Shansheng
S
Chen
Jinzhong
J
Hu
Xuchu
X
Ji
Chaoneng
C
Ying
Kang
K
Yu
Xinbing
X
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003004:Clonorchis sinensis / Q000201:enzymology* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015723:Gene Library; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA
D011994:Recombinant Proteins; D009243:NAD; D008291:Malate Dehydrogenase