Structural analysis of novel drug targets for mitigation of Pseudomonas aeruginosa biofilms.

Pseudomonas aeruginosa is an opportunistic human pathogen responsible for acute and chronic, hard to treat infections. Persistence of P. aeruginosa is due to its ability to develop into biofilms, which are sessile bacterial communities adhered to substratum and encapsulated in layers of self-produced exopolysaccharides. These biofilms provide enhanced protection from the host immune system and resilience towards antibiotics, which poses a challenge for treatment. Various strategies have been expended for combating biofilms, which involve inhibiting biofilm formation or promoting their dispersal. The current remediation approaches offer some hope for clinical usage, however, treatment and eradication of preformed biofilms is still a challenge. Thus, identifying novel targets and understanding the detailed mechanism of biofilm regulation becomes imperative. Structure-based drug discovery (SBDD) provides a powerful tool that exploits the knowledge of atomic resolution details of the targets to search for high affinity ligands. This review describes the available structural information on the putative target protein structures that can be utilized for high throughput in silico drug discovery against P. aeruginosa biofilms. Integrating available structural information on the target proteins in readily accessible format will accelerate the process of drug discovery.

The antiactivator FleN uses an allosteric mechanism to regulate σ54-dependent expression of flagellar genes in Pseudomonas aeruginosa.

Diverse sigma factors associate with the RNA polymerase (RNAP) core enzyme to initiate transcription of specific target genes in bacteria. σ54-Mediated transcription uses AAA+ activators that utilize their ATPase activity for transcription initiation. FleQ is a σ54-dependent master transcriptional regulator of flagellar genes in Pseudomonas aeruginosa. The ATPase activity of FleQ is regulated via a P-loop ATPase, FleN, through protein-protein interaction. We report a high-resolution crystal structure of the AAA+ domain of FleQ in complex with antiactivator FleN. The data reveal that FleN allosterically prevents ATP binding to FleQ. Furthermore, FleN remodels the region of FleQ essential for engagement with σ54 for transcription initiation. Disruption of the conserved protein-protein interface, by mutation, shows motility and transcription defects in vivo and multiflagellate phenotype. Our study provides a detailed mechanism used by monoflagellate bacteria to fine-tune the expression of flagellar genes to form and maintain a single flagellum.