Overexpression and one-step renaturation-purification of the tagged creatinine deiminase of Corynebacterium glutamicum in Escherichia coli cells

Cell Biol Int. 2020 May;44(5):1204-1211. doi: 10.1002/cbin.11320. Epub 2020 Feb 19.

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.

Keywords: Corynebacterium glutamicum; Escherichia coli; creatinine deiminase; gene cloning; inclusion bodies; metal-affinity chromatography.

MeSH terms

  • Aminohydrolases / isolation & purification*
  • Bacterial Proteins / isolation & purification*
  • Cloning, Molecular
  • Corynebacterium glutamicum / enzymology*
  • Escherichia coli / genetics

Substances

  • Bacterial Proteins
  • Aminohydrolases
  • creatinine deiminase