Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.184 (LasR)
 897 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The use of cell-cell communication or "quorum sensing (QS)" elements from Gram-negative Proteobacteria has enabled synthetic biologists to begin engineering systems composed of multiple interacting organisms. However, additional tools are necessary if we are to progress toward synthetic microbial consortia that exhibit more complex, dynamic behaviors. EsaR from Pantoea stewartii subsp. stewartii is a QS regulator that binds to DNA as an apoprotein and releases the DNA when it binds to its cognate signal molecule, 3-oxohexanoyl-homoserine lactone (3OC6HSL). In the absence of 3OC6HSL, EsaR binds to DNA and can act as either an activator or a repressor of transcription. Gene expression from P(esaR), which is repressed by wild-type EsaR, requires 100- to 1000-fold higher concentrations of signal than commonly used QS activators, such as LuxR and LasR. Here we have identified EsaR variants with increased sensitivity to 3OC6HSL using directed evolution and a dual ON/OFF screening strategy. Although we targeted EsaR-dependent derepression of P(esaR), our EsaR variants also showed increased 3OC6HSL sensitivity at a second promoter, P(esaS), which is activated by EsaR in the absence of 3OC6HSL. Here, the increase in AHL sensitivity led to gene expression being turned off at lower concentrations of 3OC6HSL. Overall, we have increased the signal sensitivity of EsaR more than 70-fold and generated a set of EsaR variants that recognize 3OC6HSL concentrations ranging over 4 orders of magnitude. QS-dependent transcriptional regulators that bind to DNA and are active in the absence of a QS signal represent a new set of tools for engineering cell-cell communication-dependent gene expression.
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PMID:Directed evolution of the quorum-sensing regulator EsaR for increased signal sensitivity. 2336 22

Many members of the LuxR family of quorum sensing (QS) transcriptional activators, including LasR of Pseudomonas aeruginosa, are believed to require appropriate acyl-homoserine lactone (acyl-HSL) ligands to fold into an active conformation. The failure to purify ligand-free LuxR homologues in nonaggregated form at the high concentrations required for their structural characterization has limited the understanding of the mechanisms by which QS receptors are activated. Surface-enhanced Raman scattering (SERS) is a vibrational spectroscopy technique that can be applied to study proteins at extremely low concentrations in their active state. The high sensitivity of SERS has allowed us to detect molecular interactions between the ligand-binding domain of LasR (LasRLBD) as a soluble apoprotein and modulators of P. aeruginosa QS. We found that QS activators and inhibitors produce differential SERS fingerprints in LasRLBD, and in combination with molecular docking analysis provide insight into the relevant interaction mechanism. This study reveals signal-specific structural changes in LasR upon ligand binding, thereby confirming the applicability of SERS to analyze ligand-induced conformational changes in proteins.
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PMID:Using surface enhanced Raman scattering to analyze the interactions of protein receptors with bacterial quorum sensing modulators. 2592 41