Updated project metadata. High NH4+ load is known to competitively inhibit bacterial methane oxidation. This is due to a competition between CH4 and NH4+/NH3 for the active site of particulate methane monooxygenase (pMMO), which converts CH4 to CH3OH. Here, we combined growth experiments with global proteomics to elucidate the capability of the methanotroph Methylocystis sp. strain SC2 in acclimatizing to increased NH4+ levels. Our experimental approach also involved amino acid profiling and measurement of NOx compounds. Relative to 1 mM NH4+, high (50 mM and 75 mM) NH4+ load under CH4 replete conditions significantly increased lag phase duration required for proteome adjustment. The proteomic and metabolic responses to increasing ionic and osmotic stress involved significant upregulation of stress-responsive proteins, K+ “salt in” strategy, synthesis of compatible solutes (glutamate and proline), and induction of the glutathione metabolism pathway. A significant increase in the apparent Km value for CH4 oxidation during the growth phase was indicative of increased pMMO-based oxidation of NH4+/NH3 to toxic hydroxylamine. The detoxifying activity of hydroxlyamine oxidoreductase (HAO) led to a significant accumulation of NO2- and, upon decreasing O2 tension, N2O. Putative free intermediate of HAO activity was NO, with NO reductase and hybrid cluster proteins (Hcps) being the candidate enzymes for the reduction of NO to N2O. In summary, strain SC2 has the capacity to precisely rebalance enzymes and osmolyte composition, 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 NH4+.