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Purified enzyme - cytochrome cd1 nitrite reductase

The metalloprotein cytochrome cd1 nitrite reductase has been extracted and purified from the bacterium Paracoccus 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 nitrite 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 cytochrome cd1 nitrite reductase was 1g. Solutions studies have shown that this enzyme specifically interacts with nitrite ions even in the presence of high concentrations of other potentially interfering ions. The interaction of the nitrite 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 directly to the concentration of the nitrite ions present in the solution.

Reported by

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