Overexpression and one-step renaturation-purification of the tagged creatinine deiminase of Corynebacterium glutamicum in Escherichia coli cells.
Andriy E ZakalskiyNataliya Ye StasyukOksana M ZakalskaYuriy R BoretskyMykhailo V GoncharPublished in: Cell biology international (2020)
The codA gene of Corynebacterium glutamicum PCM 1945 coding for a creatinine deiminase (CDI) (EC 3.5.4.21) has been amplified and cloned. The recombinant strain of Escherichia coli that overproduces the (His)6 -tagged inactive CDI of C. glutamicum as inclusion bodies has been constructed. After solubilization of inclusion bodies in the presence of 0.3% N-lauroylsarcosine, the enzyme was renaturated and purified by a single-step procedure using metal-affinity chromatography. The yield of the (His)6 -tagged CDI is ~30 mg from 1 L culture. The purified enzyme is sufficiently stable under the conditions designed and possesses an activity of 10-20 U/mg. The main characteristics of the tagged enzyme remained similar to that of the natural enzyme.
Keyphrases
- escherichia coli
- induced apoptosis
- uric acid
- mass spectrometry
- klebsiella pneumoniae
- gene expression
- cell cycle arrest
- wastewater treatment
- high speed
- copy number
- transcription factor
- tandem mass spectrometry
- biofilm formation
- pseudomonas aeruginosa
- cystic fibrosis
- cell death
- high performance liquid chromatography
- multidrug resistant