Abstract
Denitrification is a respiratory process by which nitrate is reduced to dinitrogen. Incomplete denitrification results in the emission of the greenhouse gas nitrous oxide and this is potentiated in acidic soils, which display reduced denitrification rates and high N2O/N2 ratios compared to alkaline soils. In this work, impact of pH on the proteome of the soil denitrifying bacterium Paracoccus denitrificans PD1222 was analysed with nitrate as sole energy and nitrogen source under anaerobic conditions at pH ranging from 6.5 to 7.5. Quantitative proteomic analysis revealed that the highest difference in protein representation was observed when the proteome at pH 6.5 was compared to the reference proteome at pH 7.2. However, this difference in the extracellular pH was not enough to produce modification of intracellular pH, which was maintained at 6.5 ± 0.1. The biosynthetic pathways of several cofactors relevant for denitrification and nitrogen assimilation like cobalamin, riboflavin, molybdopterin and nicotinamide were negatively affected at pH 6.5. In addition, peptide representation of reductases involved in nitrate assimilation and denitrification were reduced at pH 6.5. Data highlight the strong negative impact of pH on NosZ synthesis and intracellular copper content, thus impairing active NosZ assembly and, in turn, leading to elevated nitrous oxide emissions.
Highlights
A great variety of nitrogen cycle processes are highly active in soils, including denitrification, which contributes to 70% of global nitrous oxide e missions[1,2]
The aim of this work was to study the influence of pH on the proteome of the model soil denitrifier Paracoccus denitrificans PD1222, identifying proteins affected by changes in the pH with a particular interest on denitrification proteins and their cofactors and how these changes may impact on the variations in N2O emissions
To study the effect of pH on growth and the proteome of P. denitrificans PD1222, this strain was cultured in mineral salt media at different initial pHs (6.5, 7.0, 7.2 or 7.5) under anaerobic denitrifying conditions with nitrate (30 mM) as both nitrogen source and electron acceptor
Summary
A great variety of nitrogen cycle processes are highly active in soils, including denitrification, which contributes to 70% of global nitrous oxide e missions[1,2]. The model soil bacterium Paracoccus denitrificans performs the complete denitrification pathway, using a molybdenum-dependent membrane-bound nitrate reductase (Nar), a cytochrome cd1-type nitrite reductase (NirS), a heme c/b nitric oxide reductase (Nor), and a copper-dependent nitrous oxide reductase (NosZ). All these reductases can be coupled to the core electron transport pathway at the level of the ubiquinol pool in the membrane and the periplasmic heme-containing cytochrome c550 or Cu-containing pseudoazurin p ool[3,5]. Key processes leading to the passage through the soil of N-fertilizers, and formation, consumption and emission of N2O, are not fully understood in the natural context[14]
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