Methanotrophic Bacteria and Environmental Genomics/Transcriptomics Research Group, Max Planck Institute for Terrestrial Microbiologygrid.419554.8, Marburg, Germany.
Core Facility for Mass Spectrometry and Proteomics, Max Planck Institute for Terrestrial Microbiologygrid.419554.8, Marburg, Germany.
mSystems. 2022 Oct 26;7(5):e0040322. doi: 10.1128/msystems.00403-22. Epub 2022 Sep 26.
A high NH load is known to inhibit bacterial methane oxidation. This is due to a competition between CH and NH for the active site of particulate methane monooxygenase (pMMO), which converts CH to CHOH. Here, we combined global proteomics with amino acid profiling and nitrogen oxides measurements to elucidate the cellular acclimatization response of sp. strain SC2 to high NH levels. Relative to 1 mM NH, a high (50 mM and 75 mM) NH load under CH-replete conditions significantly increased the lag phase duration required for proteome adjustment. The number of differentially regulated proteins was highly significantly correlated with an increasing NH load. The cellular responses to increasing ionic and osmotic stress involved a significant upregulation of stress-responsive proteins, the K "salt-in" strategy, the synthesis of compatible solutes (glutamate and proline), and the induction of the glutathione metabolism pathway. A significant increase in the apparent value for CH oxidation during the growth phase was indicative of increased pMMO-based oxidation of NH to toxic hydroxylamine. The detoxifying activity of hydroxlyamine oxidoreductase (HAO) led to a significant accumulation of NO and, upon decreasing O tension, NO. Nitric oxide reductase and hybrid cluster proteins (Hcps) were the candidate enzymes for the production of NO. In summary, strain SC2 has the capacity to precisely rebalance enzymes and osmolyte composition in response to increasing NH exposure, but the need to simultaneously combat both ionic-osmotic stress and the toxic effects of hydroxylamine may be the reason why its acclimatization capacity is limited to 75 mM NH. In addition to reducing CH emissions from wetlands and landfills, the activity of alphaproteobacterial methane oxidizers of the genus contributes to the sink capacity of forest and grassland soils for atmospheric methane. The methane-oxidizing activity of spp. is, however, sensitive to high NH concentrations. This is due to the competition of CH and NH for the active site of particulate methane monooxygenase, thereby resulting in the production of toxic hydroxylamine with an increasing NH load. An understanding of the physiological and molecular response mechanisms of spp. is therefore of great importance. Here, we combined global proteomics with amino acid profiling and NOx measurements to disentangle the cellular mechanisms underlying the acclimatization of sp. strain SC2 to an increasing NH load.
高氨负荷已知会抑制细菌甲烷氧化。这是因为 CH 和 NH 竞争颗粒态甲烷单加氧酶（pMMO）的活性位点，pMMO 将 CH 转化为 CHOH。在这里，我们将全局蛋白质组学与氨基酸分析和氮氧化物测量相结合，阐明了 sp。菌株 SC2 在高 NH 水平下的细胞适应反应。与 1 mM NH 相比，CH 充足条件下的高（50 mM 和 75 mM）NH 负荷显著增加了调整蛋白质组所需的迟滞期。差异调节蛋白的数量与 NH 负荷的增加呈高度显著相关。对增加的离子和渗透压应激的细胞反应涉及应激响应蛋白的显著上调、K“盐进”策略、相容溶质（谷氨酸和脯氨酸）的合成以及谷胱甘肽代谢途径的诱导。在生长阶段 CH 氧化的表观值显著增加表明 NH 以有毒的羟胺形式被 pMMO 氧化增加。羟胺氧化还原酶（HAO）的解毒活性导致 NO 的显著积累，并且当 O 张力降低时，NO。一氧化氮还原酶和混合簇蛋白（Hcps）是产生 NO 的候选酶。总之，菌株 SC2 具有根据不断增加的 NH 暴露重新平衡酶和渗透物组成的能力，但需要同时对抗离子-渗透压应激和羟胺的毒性影响可能是其适应能力限于 75 mM NH 的原因。除了减少湿地和垃圾填埋场的 CH 排放外，属中的α变形菌甲烷氧化菌的活性有助于森林和草原土壤对大气甲烷的汇能力。然而， spp 的甲烷氧化活性。对高 NH 浓度敏感。这是因为 CH 和 NH 竞争颗粒态甲烷单加氧酶的活性位点，从而导致随着 NH 负荷的增加产生有毒的羟胺。因此，了解 spp 的生理和分子反应机制非常重要。在这里，我们将全局蛋白质组学与氨基酸分析和 NOx 测量相结合，以阐明 sp 的细胞适应机制。菌株 SC2 对不断增加的 NH 负荷的适应。
