Ammonia-oxidizing bacteria (AOB) and archaea (AOA) derive total energy for life from the multi-electron, proton-coupled oxidation of ammonia (NH3) to nitrite (NO2-). The relevant pathway—nitrification—constitutes a major sink of NH3 applied to soil to improve crop yields. Additional consequences of unchecked nitrification include production of harmful wastewater and gaseous pollutants. Enzymes known to participate in biological nitrification include the copper-containing membrane monooxygenase ammonia monooxygenase (AMO) and the multi-heme containing hydroxylamine oxidoreductase (HAO). Mechanistic study of the enzymes driving this chemistry would inform more efficient and sustainable agricultural practices. Our lab has studied cytochrome (cyt) P460, which contains the same unique active site as HAO, but without the additional electron transfer hemes that can interfere with spectroscopic analysis. Recently, our lab has shown a direct link between cyt P460 and the agriculture-based production of the potent greenhouse gas nitrous oxide (N2O) through the oxidation of hydroxylamine (NH2OH). Our reported mechanism of NH2OH oxidation by cyt P460 invokes an FeII intermediate that is rapidly oxidized to close the catalytic cycle. However, we now show that this FeII intermediate exhibits unique reactivity with NH2OH. We now present a detailed kinetic and spectroscopic study of the FeII intermediate and its reactivity with compounds germane to nitrification. These studies expand our understanding of the reactivity of P460-heme containing enzymes, thus contributing fresh insights to the vibrant field of porphyrinoid biochemistry.

Elucidating the reactivity of ferrous heme-P460 cofactors

254th National Meeting (2017)

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