Thiol-benzo-triazolo-quinazolinone Inhibits Alg44 Binding to c-di-GMP and Reduces Alginate Production by Pseudomonas aeruginosa.

Abstract

Pseudomonas aeruginosa is an opportunistic pathogen that affects a large proportion of cystic fibrosis (CF) patients. CF patients have dehydrated mucus within the airways that leads to the inability of the mucociliary escalator to expel inhaled microbes. Once inhaled, P. aeruginosa can persist in the lungs of the CF patients for the remainder of their lives. During this chronic infection, a phenomenon called mucoid conversion can occur in which P. aeruginosa can mutate and inactivate their mucA gene. As a consequence, transcription of the alg operon is highly expressed, leading to the copious secretion of the alginate exopolysaccharide, which is associated with decreased lung function and increased CF patient morbidity and mortality. Alginate biosynthesis by P. aeruginosa is post-translationally regulated by bis(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP), which binds to the receptor protein Alg44 to activate alginate production. The identification of small molecules that disrupt the binding of c-di-GMP to Alg44 could inhibit the ability of P. aeruginosa to produce alginate. In this work, a class of thiol-benzo-triazolo-quinazolinone compounds that inhibited Alg44 binding to c-di-GMP in vitro was identified after screening chemical libraries consisting of ∼50 000 chemical compounds. Thiol-benzo-triazolo-quinazolinones were shown to specifically inhibit Alg44-c-di-GMP interactions by forming a disulfide bond with the cysteine residue in the PilZ domain of Alg44. The more potent thiol-benzo-triazolo-quinazolinone had the ability to reduce P. aeruginosa alginate secretion by up to 30%. These compounds serve as leads in the development of novel inhibitors of alginate production by P. aeruginosa after mucoid conversion.