Publications

• Structure of PqsD, a Pseudomonas quinolone signal biosynthetic enzyme, in complex with anthranilate.

  Bera A.K., Atanasova V., Robinson H., Eisenstein E., Coleman J.P., Pesci E.C., Parsons J.F.

  Pseudomonas quinolone signal (PQS), 2-heptyl-3-hydroxy-4-quinolone, is an intercellular alkyl quinolone signaling molecule produced by the opportunistic pathogen Pseudomonas aeruginosa. Alkyl quinolone signaling is an atypical system that, in P. aeruginosa, controls the expression of numerous viru ... >> More

  Pseudomonas quinolone signal (PQS), 2-heptyl-3-hydroxy-4-quinolone, is an intercellular alkyl quinolone signaling molecule produced by the opportunistic pathogen Pseudomonas aeruginosa. Alkyl quinolone signaling is an atypical system that, in P. aeruginosa, controls the expression of numerous virulence factors. PQS is synthesized from the tryptophan pathway intermediate, anthranilate, which is derived either from the kynurenine pathway or from an alkyl quinolone specific anthranilate synthase encoded by phnAB. Anthranilate is converted to PQS by the enzymes encoded by the pqsABCDE operon and pqsH. PqsA forms an activated anthraniloyl-CoA thioester that shuttles anthranilate to the PqsD active site where it is transferred to Cys112 of PqsD. In the only biochemically characterized reaction, a condensation then occurs between anthraniloyl-PqsD and malonyl-CoA or malonyl-ACP, a second PqsD substrate, forming 2,4-dihydroxyquinoline (DHQ). The role PqsD plays in the biosynthesis of other alkyl quinolones, such as PQS, is unclear, though it has been reported to be required for their production. No evidence exists that DHQ is a PQS precursor, however. Here we present a structural and biophysical characterization of PqsD that includes several crystal structures of the enzyme, including that of the PqsD-anthranilate covalent intermediate and the inactive Cys112Ala active site mutant in complex with anthranilate. The structure reveals that PqsD is structurally similar to the FabH and chalcone synthase families of fatty acid and polyketide synthases. The crystallographic asymmetric unit contains a PqsD dimer. The PqsD monomer is composed of two nearly identical approximately 170-residue alphabetaalphabetaalpha domains. The structures show anthranilate-liganded Cys112 is positioned deep in the protein interior at the bottom of an approximately 15 A long channel while a second anthraniloyl-CoA molecule is waiting in the cleft leading to the protein surface. Cys112, His257, and Asn287 form the FabH-like catalytic triad of PqsD. The C112A mutant is inactive, although it still reversibly binds anthraniloyl-CoA. The covalent complex between anthranilate and Cys112 clearly illuminates the orientation of key elements of the PqsD catalytic machinery and represents a snapshot of a key point in the catalytic cycle. << Less

  Biochemistry 48:8644-8655(2009) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• PqsE of Pseudomonas aeruginosa acts as pathway-specific thioesterase in the biosynthesis of alkylquinolone signaling molecules.

  Drees S.L., Fetzner S.

  Pseudomonas aeruginosa uses the alkylquinolones PQS (2-heptyl-3-hydroxy-4(1H)-quinolone) and HHQ (2-heptyl-4(1H)-quinolone) as quorum-sensing signal molecules, controlling the expression of many virulence genes as a function of cell population density. The biosynthesis of HHQ is generally accepted ... >> More

  Pseudomonas aeruginosa uses the alkylquinolones PQS (2-heptyl-3-hydroxy-4(1H)-quinolone) and HHQ (2-heptyl-4(1H)-quinolone) as quorum-sensing signal molecules, controlling the expression of many virulence genes as a function of cell population density. The biosynthesis of HHQ is generally accepted to require the pqsABCD gene products. We now reconstitute the biosynthetic pathway in vitro, and demonstrate that in addition to PqsABCD, PqsE has a role in HHQ synthesis. PqsE acts as thioesterase, hydrolyzing the biosynthetic intermediate 2-aminobenzoylacetyl-coenzyme A to form 2-aminobenzoylacetate, the precursor of HHQ and 2-aminoacetophenone. The role of PqsE can be taken over to some extent by the broad-specificity thioesterase TesB, explaining why the pqsE deletion mutant of P. aeruginosa still synthesizes HHQ. Interestingly, the pqsE mutant produces increased levels of 2,4-dihydroxyquinoline, resulting from intramolecular cyclization of 2-aminobenzoylacetyl-coenzyme A. Overall, our data suggest that PqsE promotes the efficiency of alkylquinolone signal molecule biosynthesis in P. aeruginosa and balances the levels of secondary metabolites deriving from the alkylquinolone biosynthetic pathway. << Less

  Chem. Biol. 22:611-618(2015) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• The end of an old hypothesis: the Pseudomonas signaling molecules 4-hydroxy-2-alkylquinolines derive from fatty acids, not 3-ketofatty acids.

  Dulcey C.E., Dekimpe V., Fauvelle D.A., Milot S., Groleau M.C., Doucet N., Rahme L.G., Lepine F., Deziel E.

  Groups of pathogenic bacteria use diffusible signals to regulate their virulence in a concerted manner. Pseudomonas aeruginosa uses 4-hydroxy-2-alkylquinolines (HAQs), including 4-hydroxy-2-heptylquinoline (HHQ) and 3,4-dihydroxy-2-heptylquinoline (PQS), as unique signals. We demonstrate that octa ... >> More

  Groups of pathogenic bacteria use diffusible signals to regulate their virulence in a concerted manner. Pseudomonas aeruginosa uses 4-hydroxy-2-alkylquinolines (HAQs), including 4-hydroxy-2-heptylquinoline (HHQ) and 3,4-dihydroxy-2-heptylquinoline (PQS), as unique signals. We demonstrate that octanoic acid is directly incorporated into HHQ. This finding rules out the long-standing hypothesis that 3-ketofatty acids are the precursors of HAQs. We found that HAQ biosynthesis, which requires the PqsABCD enzymes, proceeds by a two-step pathway: (1) PqsD mediates the synthesis of 2-aminobenzoylacetate (2-ABA) from anthraniloyl-coenzyme A (CoA) and malonyl-CoA, then (2) the decarboxylating coupling of 2-ABA to an octanoate group linked to PqsC produces HHQ, the direct precursor of PQS. PqsB is tightly associated with PqsC and required for the second step. This finding uncovers promising targets for the development of specific antivirulence drugs to combat this opportunistic pathogen. << Less

  Chem. Biol. 20:1481-1491(2013) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.