In the present study the polycyclic aromatic hydrocarbon removal and metabolic adaptation of Amycolatopsis tucumanensis DSM 45259 were investigated. Analysis of one-dimensional gel electrophoresis of crude cell extracts revealed differential synthesis of proteins which were identified by MALDI-TOF. To elucidate the phenanthrene metabolic pathway in A. tucumanensis DSM45259, two-dimensional electrophoresis and detection of phenanthrene degradation intermediates by GS-MS were performed. The presence of aromatic substrates resulted in changes in the abundance of proteins involved in the metabolism of aromatic compounds, oxidative stress response, energy production and protein synthesis. The obtained results allowed us to clarify the phenanthrene catabolic pathway, by confirming the roles of several proteins involved in the degradation process and comprehensive adaptation. This may clear the way for more efficient engineering of bacteria in the direction of more effective bioremediation applications.