Isolation and preliminary characterization of a respiratory nitrate reductase from hydrocarbon-degrading bacterium Gordonia alkanivorans S7.

Gordonia alkanivorans S7 is an efficient degrader of fuel oil hydrocarbons that can simultaneously utilize oxygen and nitrate as electron acceptors. The respiratory nitrate reductase (Nar) from this organism has been isolated using ion exchange chromatography and gel filtration, and then preliminarily characterized. PAGE, SDS-PAGE and gel filtration chromatography revealed that Nar consisted of three subunits of 103, 53 and 25 kDa. The enzyme was optimally active at pH 7.9 and 40 degrees C. K(m) values for NO(3)(-) (110 microM) and for ClO(3)(-) (138 microM) were determined for a reduced viologen as an electron donor. The purified Nar did not use NADH as the electron donor to reduce nitrate or chlorate. Azide was a strong inhibitor of its activity. Our results imply that enzyme isolated from G. alkanivorans S7 is a respiratory membrane-bound nitrate reductase. This is the first report of purification of a nitrate reductase from Gordonia species.