Characterization of a potential diguanylate cyclase from Pseudomonas aeruginosa

Abstract

The second messenger cyclic bis-(3',5')-di-guanosine monophosphate (c-di-GMP) influences the regulation of several bacterial processes involved in virulence, such as biofilm formation. The synthesis and degradation of c-di-GMP occur due to two classes of proteins: diguanylate cyclases (DGCs), with active GGDEF domains; and phosphodiesterases (PDEs) with EAL or HD-GYP domains. The opportunistic bacterium Pseudomonas aeruginosa harbors about 40 proteins with such domains, however, many of these proteins remain uncharacterized. Among these, is the PA14_04420 protein, which has the GGDEF domain and the PAS sensory domain, with its auxiliary domain PAC. Previous results have shown a putative interaction of PA14_04420 with proteins involved in cell division, cell wall formation, and lipopolysaccharide synthesis. Therefore, this project aims to evaluate the potential catalytic activity of diguanylate cyclase of the PA14_04420 protein by expressing and purifying the recombinant protein and also evaluating concentrations of c-di-GMP produced in vivo, and verifying the enzymatic activity in vitro. Thus, the characterization of PA14_04420 may help to better understand the roles of c-di-GMP signaling pathways, which may provide a new target for therapeutic development against bacterial infections.(AU)