Community Research and Development Information Service - CORDIS

Purified enzyme - nitrate reductase

The metalloprotein nitrate reductase has been extracted and purified from a genetically modified strain of the bacterium Paracocus denitrificans (formerly Thiosphaera pantrotropha). The bacterium was cultured in a 200 I bioreactor and was harvested using a cross-flow filtration system. The bacterium was fractionated and the soluble nitrate reductase enzyme was purified from the periplasmic fraction using a combination of anion-exchange chromatography, ammonium sulphate precipitation and nickel affinity chromatography.
The final yield of the nitrate reductase was ca. 30mg. Solution studies have shown that this enzyme specifically interacts with nitrate ions even in the presence of high concentrations of other potentially interfering ions. The interaction of the nitrate anion with the enzyme is indicated by changes in both the spectroscopic and redox properties of the haem centre of the protein. These changes can be related to the concentration of the nitrate ions present in the solution.

Reported by

University of East Anglia
University Plain
NR4 7TJ Norwich
United Kingdom
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