Overexpression and one‐step renaturation‐purification of the tagged creatinine deiminase of Corynebacterium glutamicum in Escherichia coli cells |
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| Authors: | Andriy E. Zakalskiy Nataliya Ye. Stasyuk Oksana M. Zakalska Yuriy R. Boretsky Mykhailo V. Gonchar |
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| Abstract: | 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. |
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| Keywords: | Corynebacterium glutamicum creatinine deiminase Escherichia coli gene cloning inclusion bodies metal‐affinity chromatography |
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