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JBC, Vol. 252, Issue 1, 212-218, Jan, 1977
R. T. St John and T. C. Hollocher
The pathway of anaerobic reduction of nitrite to nitrogen gas (N2) by cell
suspensions of the denitrifier, Pseudomonas aeruginosa, was studied using
the techniques of gas chromatography and mass spectrometry. While release
of nitrous oxide (N2O) is not normally detected during the reduction of
nitrite to N2 by this organism, 15N from [15N]nitrite nevertheless can be
trapped quantitatively as 15N2O in a pool of added N2O. In such experiments
the abundance of 15N in N2O always exceeds that in product N2, consistent
with the absence of a major reductive route from nitrite to N2 which
by-passes N2O. During the reduction of a mixture of [15N]nitrite and nitric
oxide (NO), 15NO produced at most only in trace amounts. The final products
are chiefly 15N2 and 14N2 with only a small fraction of the scrambled
product, 14N15N. Much of the 14N15N can be accounted for as an artifact
caused by traces of molecular oxygen, which promote the conversion of NO to
nitrite by autooxidation and thereby degrade slightly the isotopic purity
of [15N]nitrite. Nitrous oxide shows all the properties of a free
obligatory intermediate during the denitrification of nitrite to N2 by P.
aeruginosa, whereas NO does not. The inability to trap 15NO in a pool of NO
indicates that NO is not a free obligatory intermediate in the reduction of
nitrite. The small mole fractions of 14N15N produced from a mixture of
[15N]nitrite and NO require that the main reductive pathways for these
nitrogen oxides cannot share any freely diffusible mono-nitrogen
intermediate in common. The simplest interpretation is that nitrite and NO
are denitrified by separate pathways, at least prior to the formation of
the first bi-nitrogen compound.
Nitrogen 15 tracer studies on the pathway of denitrification in Pseudomonas aeruginosa
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