Compartmentalization of biosynthetic enzymes in bacterial cells: the histidine metabolic pathway case

It is known that the inner concentration of proteins within the cell cytoplasm is so high that limits the diffusion of enzymes and metabolic intermediates, leading to a loss of time and energy. Therefore, the organization of genes in operons would have enabled to have enzymes involved in the same metabolic pathway physically close to each other. The aim of this study is to demonstrate the compartmentalization of proteins involved in the same metabolic pathway, and the formation of multi-enzymatic complexes in bacterial cells, using as model system the histidine biosynthesis. In Escherichia coli the genes involved in this metabolic pathway are organized in a compact operon. To evaluate the possible interactions between the enzymes of this pathway a combination of different approaches has been used: the Bacterial Adenylate Cyclase Two-Hybrid (BACTH) system, the β-galactosidase assay, the His-tag method, and microscopy analyses. So far, using the BACTH system, the interaction between the enzymes His-H and His-F has been validate, as a positive control of the system. The testing of all the other possible interactions between the enzymes of the histidine biosynthetic pathway, through the proposed multidisciplinary approach, are in progress to evaluate if they form a single big multi-enzymatic complex.