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Discovering Selective Diguanylate Cyclase Inhibitors: From PleD to Discrimination of the Active Site of Cyclic-di-GMP Phosphodiesterases.
One of the most important signals involved in controlling biofilm formation is represented by the intracellular second messenger 3',5'-cyclic diguanylic acid (c-di-GMP). Since the pathways involved in c-di-GMP biosynthesis and breakdown are found only in bacteria, targeting c-di-GMP metabolism represents an attractive strategy for the development of biofilm-disrupting drugs. Here, we present the workflow required to perform a structure-based design of inhibitors of diguanylate cyclases, the enzymes responsible for c-di-GMP biosynthesis. Downstream of the virtual screening process, detailed in the first part of the chapter, we report the step-by-step protocols required to test the positive hits in vitro and to validate their selectivity, thus minimizing possible off-target effects.
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