• metabolic compartimentalization

The idea behind the project was to imagine what happen if we consider in the same reaction mix, different metabolic pathways that usually don’t take place in the same cell space. With spectrophotometric analysis and mass spectrometry analysis different enzymatic activities were tested in presence of metabolites belonging to a different metabolic pathway. In this way inhibition responses were detected. The analyses were conducted using computational tools, as R studio statistical software and Graph Pad prism software. For the spectrophotometric assay, the plate reader was used, while for the mass spectrometry assays a triple Quad MS was used, combined with high performance liquid chromatography (HPLC) was used. The experiments have considered separately the reactions of different enzymes in presence and absence of the metabolites of another metabolic pathway. All the experiments were performed in vitro. The second and last part of the project was a computational analysis, using one of the software for simulation and modeling of biochemical networks: COPASI. A previous biological model was modified using the biochemical parameters calculated from the experimental data. The inhibitions calculated, has changed the metabolic fluxes of the model, confirming the initial idea that a biochemical barrier could take place in the cellular environment in absence of the membrane barriers that already keep unconnected some metabolic pathways.