Genome-scale metabolic model analysis indicates low energy production efficiency in marine ammonia-oxidizing archaea

Abstract

Marine ammonia-oxidizing archaea (AOA) play an important role in the global nitrogen cycle by obtaining energy for biomass production from CO₂ via oxidation of ammonium. The isolation of Candidatus "Nitrosopumilus maritimus" strain SCM1, which represents the globally distributed AOA in the ocean, provided an opportunity for uncovering the contributions of those AOA to carbon and nitrogen cycles in ocean. Although several ammonia oxidation pathways have been proposed for SCM1, little is known about its ATP production efficiency. Here, based on the published genome of Nitrosopumilus maritimus SCM1, a genome-scale metabolic model named NmrFL413 was reconstructed. Based on the model NmrFL413, the estimated ATP/NH₄⁺ yield (0.149-0.276 ATP/NH₄⁺) is tenfold lower than the calculated theoretical yield of the proposed ammonia oxidation pathways in marine AOA (1.5-1.75 ATP/NH₄⁺), indicating a low energy production efficiency of SCM1. Our model also suggested the minor contribution of marine AOA to carbon cycle comparing with their significant contribution to nitrogen cycle in the ocean.

Keywords: Ammonia oxidation pathway; Ammonia-oxidizing archaea; Energy production efficiency; Genome-scale metabolic model; Nitrosopumilus maritimus SCM1.

Fig. 1

The workflow of genome-scale metabolic model reconstruction of strain SCM1

Fig. 2

The Thaumarchaeal HP/HB cycle for carbon fixation. Four missing reactions in the strain SCM1 draft model were marked in red-dashed line. Enzymes numbered in brackets are: 1, acetyl-CoA carboxylase (EC 6.4.1.2); 2, malonyl-CoA reductase (NADPH) (EC 1.2.1.75); 3, malonic semialdehyde reductase (NADPH) (EC 1.1.1.-); 4, 3-hydroxypropionyl-CoA synthetase (ADP-forming) (EC 6.2.1.-); 5, 3-hydroxypropionyl-CoA dehydratase (EC 4.2.1.116); 6, acryloyl-CoA reductase (EC 1.3.1.84); 7, propionyl-CoA carboxylase (EC 6.4.1.3); 8, methylmalonyl-CoA epimerase (EC 5.1.99.1); 9, methylmalonyl-CoA mutase (EC 5.4.99.2); 10, succinyl-CoA reductase (NADPH) (EC 1.2.1.76); 11, succinic semialdehyde reductase (NADPH) (EC 1.1.1-); 12, 4-hydroxybutyryl-CoA synthetase (ADP-forming) (EC 6.2.1.-); 13, 4-hydroxybutyryl-CoA dehydratase (EC 4.2.1.20); 14, crotonyl-CoA hydratase [(S)-3-hydroxybutyryl-CoA forming] (EC 4.2.1.17) 15, (S)-3-hydroxybutyryl-CoA dehydrogenase (NAD+) (EC 1.1.1.35); 16, acetoacetyl-CoA β-ketothiolase (EC 2.3.1.9)

Fig. 3

The arginine biosynthesis pathways in the strain SCM1 model. Genes for the first reaction (EC 2.3.1.1) in the normal arginine synthesis pathway (up part) is missing in the genome and thus a gap exists in the pathway. The existence of gene Nmar_1288 encoding ArgX that catalyzes the first step of the LysW-mediated arginine pathway (bottom part) suggests that strain SCM1 uses this pathway for arginine synthesis. Glu: l-glutamate; acglu: N-acetyl-l-glutamate; acg5p: N-acetyl-l-glutamate 5-phosphate; acg5sa: N-acetyl-l-glutamate 5-semialdehyde; acorn: N-acetylornithine; orn: l-ornithine; citr: l-citrulline; argsuc: N-(l-Arginino) succinate; arg: l-arginine; lysW-glu: LysW-l-glutamate; lysW-g5p: LysW-l-glutamyl 5-phosphate; lysW-g5sa: LysW-l-glutamate 5-semialdehyde; lysW-orn: LysW-l-ornithine

Fig. 4

Three proposed ammonia oxidation pathways in AOA. Pathway 1 was proposed by Walker et al. In this pathway, ammonia is oxidized to hydroxylamine by the membrane enzyme complex AMO/CuMMO (Walker et al. 2010). Subsequently, hydroxylamine is oxidized to nitrite in the periplasm by a heme-rich hydroxylamine oxidoreductase (CuHAO) complex. Four electrons from this oxidation are transferred to the quinone pool. Two electrons from the reduced quinone pool return to AMO (marked by red) and are required to initiate ammonia oxidation. The remaining two electrons enter the electron transport chain composed of pcy protein to generate the proton motive force (PMF) necessary for ATP synthesis and NADH synthesis. In Pathway 2 (Schleper and Nicol ; Stahl and de la Torre 2012), NO is speculated functioning as a redox shuttle to deliver electrons to the AMO (marked by green) since measurable amounts of NO are produced during ammonia oxidation. In pathway 3, iterative production and consumption of NO is involved in conversion of hydroxylamine to nitrite facilitated by a proposed novel copper enzyme capable of performing known P460 activity (CuP460) (Kozlowski et al. 2016). N₂O was formed abiotically from NO by interaction with media components or with debris in killed cell. AMO/CuMMO, ammonia monooxygenase; CuHAO, hydroxylamine dehydrogenase; NIR, Cu-containing NO-forming nitrite reductase; pcy, plastocyanin; Q/QH₂, quinone/quinol pool

Fig. 5

Outline of the reconstructed strain SCM1 metabolic network NmrFL413

Fig. 6

Comparison of theoretical and estimated ATP/NH₄⁺ yield based on Table 4