Depending on the environmental stimuli, bacteria that normally live in a planktonic form adhere to a surface, forming cellular communities called biofilms. Despite the complexity and the involvement of several bacterial systems, it is known that the intracellular level of a small signaling molecule (c-di-GMP) controls this phenotypic transition. High concentrations of c-di-GMP usually implies in biofilms formation. Bacterial communities are highly resistant to conventional treatment with antibiotics and represent the predominant phenotype in most chronic infections, resulting in serious medical problems. The c-di-GMP is synthesized from two GTP molecules by enzymes diguanilate cyclases (DGC) belonging to GGDEF family, which are, therefore, potential targets for developing new therapies that interfere with the process of biofilm formation. In this project, we propose the application of computational methods for identifying ligands of enzymes diguanilate cyclases, evaluate the molecular recognition of compounds identified by kinetic methods, thermodynamic and structural characterization. For this, two already characterized DGC (WSPR from Pseudomonas aeruginosa and YdeH from Escherichia coli) and two enzymes not studied yet from Xantomonas axonopodis (XAC2482 and XAC0614) will be cloned and tested for kinetic and structural studies with the identified inhibitors. It is expected that analysis of interactions between selected compounds and DGCS provide the basis for the development of specific inhibitors of this enzyme family and allow understanding the molecular mechanisms involved in signaling by c-di-GMP.
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