The Physiology and Biochemistry of Archaeal Ammonia Oxidisers

Wright, Chloe (2021) The Physiology and Biochemistry of Archaeal Ammonia Oxidisers. Doctoral thesis, University of East Anglia.

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Archaeal and bacterial ammonia oxidisers (AOA and AOB) initiate nitrification by oxidising ammonia (NH3) to hydroxylamine, catalysed by ammonia monooxygenase (AMO). Archaeal ammonia oxidation was first confirmed 16 years ago with the cultivation of Nitrosopumilus maritimus SCM1. Since then, environmental gene surveys have revealed AOA are ubiquitous and often outnumber their bacterial counterparts in many nitrifying environments. Whilst ecological studies have demonstrated that AOA play a significant role in the global N cycle, little is known about the underpinning physiology and biochemistry. Here, culture-dependent research was conducted using phylogenetically distinct AOA isolates to gain insights about AOA energy metabolism. This began with exploring the structure and function of the archaeal AMO enzyme, which is not yet amenable to purification. By characterising the inhibition of archaeal AMOs to specific inhibitors and comparing with other members of the copper-dependent membrane monooxygenase (CuMMO) family, this study provided insights into the structure of the archaeal AMO active site(s) and its potential substrate range. Specifically, archaeal AMO has a narrower hydrocarbon substrate range compared to bacterial AMO and is restricted to oxidising short-chain-length hydrocarbons based on 1-alkynes inhibition profiles. Phenylacetylene inhibited the archaeal and bacterial AMO at different thresholds and by different mechanisms, highlighting structural differences between the two monooxygenases. Further work explored the oxidation and metabolism of methane and methanol by AOA using 13C-tracer experiments. Results suggested that methane and methanol were oxidised and metabolised, but this was dependent on the concentration of ammonia present. Ammonia competes with methane/methanol for the same AMO binding site and provides the only source of reductant for AMO activity. Subsequently, hydrazine was tested as an external source of reductant for AMO driven oxidations. These findings prompted an exploration into the similarities and differences between the archaeal and bacterial ammonia oxidation pathways, principally the role of nitric oxide (NO).

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Biological Sciences
Depositing User: Nicola Veasy
Date Deposited: 30 Mar 2022 13:15
Last Modified: 30 Mar 2022 13:15


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