Recombinant expression, characterization and application of a dihydrolipoamide dehydrogenase with diaphorase activity from Bacillus sphaericus
作者： Anvarsadat Kianmehr , Rahman Mahdizadeh , Morteza Oladnabi , Javad Ansari
DOI: 10.1007/S13205-017-0763-0
关键词:
摘要: Diaphorases are flavin-containing enzymes with potential applications in biotransfomation reactions, biosensor design and vitro diagnostic tests. In this communication, we describe recombinant expression, characterization application of a lipoamide dehydrogenase (DLD) diaphorase activity from strain Bacillus sphaericus. The DLD gene consisting 1413 bp encoding protein 470 amino acids was expressed Escherichia coli BL21 (DE3) the enzyme characterized. B. sphaericus catalyzed reduction NAD+ by dihydrolipoamide exhibited NADH-dependent activity. molecular weight purified about 50 kDa, determined to be monomeric protein. Diaphorase active stable pH 7.0 9.0 an optimal at 8.5. It showed its maximal temperature 30 °C almost temperatures between 25 30 °C. Different metal ions inhibitors no influence on target enzyme. K m V max values for NADH were estimated 0.33 mM 200.0 U/ml, respectively. Moreover, considerable used as component tests quantification metabolites. conclusion, considering properties PAD-91, it can have
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