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From Proteopedia

Quorum-sensing in Pseudomonas aeruginosa

Since their initial discovery when it was found that E. coli could inactivate penicillin, antibiotic-resistant bacteria have posed concerns to many. Recently, hospital acquired infections have increased with the increasingly common lactose-fermenting gram-negative bacteria, and this has resulted in increased attention among clinicians. One such pathogen, Pseudomonas aeruginosa, is a dangerous bacterium that is resistant to many classes of antibiotics. (Bonomo and Szabo 2006) This bacterium is the most significant pathogen that inhabits the lungs of patients with the disease cystic fibrosis (CF). Once P. aeruginosa has colonized the lungs of these patients, antibiotics have little efficacy in eradicating the pathogen, which frequently leads to eventual to lung damage, respiratory failure, and death. (Singh et al., 2000) One reason that colonization is so harmful to patients infected with the pathogen is due to the quorum-sensing systems las and rhl, which contribute to the negative health effects of infection by this bacterium. Quorum sensing in bacteria involves the production and detection of acyl homoserine lactones (AHLs). These molecules can freely diffuse across bacterial membranes, which means that the concentration of these quorum-sensing molecules is the same both inside the cell that produces them and in the extracellular medium of that cell. As these molecules are produced by an increasing number of bacterial cells, their extracellular concentrations increase accordingly, where they form a complex with a transcriptional activator. This complex induces gene expression, resulting in the bacterium’s response to the factors produced by other bacterial cells. (Davies and Bilton, 2009) One quorum-sensing system in P. aeruginosa is the las system. The AHL in this system is N-(3-oxododecanoyl) homoserine lactone (3O-C12-HSL), which is produced by the synthase protein lasI. As concentration of 3O-C12-HSL increases, it binds to and activates the transcriptional activator protein LasR. The activated LasR/3OC12-HSL complex then binds to the lux-box, which is a chromosomal DNA sequence upstream of various target genes, including LasI and perhaps LasR. (Anguige, King, Ward, 2006)

LasI

The 3O-C12-HSL synthase uses 3-oxo-C12-acyl-carrier protein (acyl-ACP) and S-adenosyl-L-methionine to synthesize 3O-C12-HSL. The protein can be seen in pink with its ligands in lime green. The residues Arg23, Phe27, and Trp33 are presumed to form the , which can be seen here in dark blue. The residues Arg154 and Lys150 are thought to be the , seen here in light blue. Additionally, the residues Trp33, Trp69, Met79, Leu102, Phe105, Thr121, Leu122, Met125, Leu140, Thr142, Thr144, Val148, Met151, Met152, Ala155, Leu157, Ile178 and Leu188 form a that is the acyl-chain binding pocket. (Gould, Schweizer, and Churchill, 2004)

LasR

(Unless otherwise noted, protein structures are shown in green and ligands are shown in red.)

The transcriptional activator LasR binds to 3O-C12-HSL and the complex then activates transcription. In solution the binding domain forms a , which can be seen here with one subunit in light blue, the other in light green, and the ligand 3O-C12-HSL in red. Each subunit consists of a 5-stranded antiparallel (shown here in yellow) with two sets of three (shown here in green) on either side of the sheets. (Zou and Nair, 2009)

The loop (shown here in royal blue) covers the binding pocket of the binding domain and prevents it from being accessible to solvent. A number of including Tyr56, Trp60 , Asp73, and Ser129 (shown here in light blue) are involved in hydrogen-bonding with the polar head of the ligand. The (shown here in orange) Leu36, Leu40, Ile52, Val76, and Leu125 also interact with the ligand, but they interact with its hydrophobic tail through van der Waals interactions. (Zou and Nair, 2009)

An important factor that determines specificity of the transcriptional activator for its ligand (as opposed to other acyl homoserine lactone with a different chain length) is the residue (shown here in dark blue), which is a part of the L3 loop. This residue compacts the hydrocarbon tail of the ligand against the protein and shields it against unfavorable interaction with the solvent. Presumably the absence of this interaction would result in exposure of the hydrophobic pocket and protein aggregation. A smaller sized ligand should also destabilize the interaction, meaning that the protein is most stable only when bound to its specific ligand. (Zou and Nair, 2009)

However, molecules other than 3O-C12-HSL are able to bind to the transcriptional activator. For example some triphenyl mimics have been found to activate the transcriptional activator, such as . Although this mimic behaves similarly to the natural ligand 3O-C12-HSL, one notable difference is that it forces the protein residue (shown here in blue) to interact with the solvent instead of facing toward the ligand binding pocket as is the case when 3O-C12-HSL is bound. (Zou and Nair, 2009)

References

Anguige, K., King, J.R., Ward, J.P, 2006. A multi-phase mathematical model of quorum sensing in a maturing Pseudomonas aeruginosa biofilm. Math Biosci.;203(2):240-76.

Bonomo, R.A., Szabo, D, 2006. Mechanisms of multidrug resistance in Acinetobacter species and Pseudomonas aeruginosa. Clin Infect Dis;43:Suppl 2:S49-S56.

Davies, J.C., Bilton, D, 2009. Bugs, biofilms, and resistance in cystic fibrosis. Respir Care;54(5):628–40.

Gould TA, Schweizer HP, Churchill ME, 2004. Structure of the Pseudomonas aeruginosa acyl-homoserinelactone synthase LasI. Mol Microbiol. 53(4):1135-46.

Singh, P.K., Schaefer, A.L., Parsek, M.R., et al., 2000. Quorum-sensing signals indicate that cystic fibrosis lungs are infected with bacterial biofilms. Nature;407(6805):762–4.

Zou Y, Nair SK, 2009. Molecular basis for the recognition of structurally distinct autoinducer mimics by the Pseudomonas aeruginosa LasR quorum-sensing signaling receptor. Chem Biol;16(9):961-70.