Molecular basis of HHQ biosynthesis: molecular dynamics simulations, enzyme kinetic and surface plasmon resonance studies

2.50
Hdl Handle:
http://hdl.handle.net/10033/620948
Title:
Molecular basis of HHQ biosynthesis: molecular dynamics simulations, enzyme kinetic and surface plasmon resonance studies
Authors:
Steinbach, Anke; Maurer, Christine K; Weidel, Elisabeth; Henn, Claudia; Brengel, Christian; Hartmann, Rolf W; Negri, Matthias
Abstract:
Abstract Background PQS (P seudomonas Quinolone Signal) and its precursor HHQ are signal molecules of the P. aeruginosa quorum sensing system. They explicate their role in mammalian pathogenicity by binding to the receptor PqsR that induces virulence factor production and biofilm formation. The enzyme PqsD catalyses the biosynthesis of HHQ. Results Enzyme kinetic analysis and surface plasmon resonance (SPR) biosensor experiments were used to determine mechanism and substrate order of the biosynthesis. Comparative analysis led to the identification of domains involved in functionality of PqsD. A kinetic cycle was set up and molecular dynamics (MD) simulations were used to study the molecular bases of the kinetics of PqsD. Trajectory analysis, pocket volume measurements, binding energy estimations and decompositions ensured insights into the binding mode of the substrates anthraniloyl-CoA and β-ketodecanoic acid. Conclusions Enzyme kinetics and SPR experiments hint at a ping-pong mechanism for PqsD with ACoA as first substrate. Trajectory analysis of different PqsD complexes evidenced ligand-dependent induced-fit motions affecting the modified ACoA funnel access to the exposure of a secondary channel. A tunnel-network is formed in which Ser317 plays an important role by binding to both substrates. Mutagenesis experiments resulting in the inactive S317F mutant confirmed the importance of this residue. Two binding modes for β-ketodecanoic acid were identified with distinct catalytic mechanism preferences.
Citation:
BMC Biophysics. 2013 Aug 01;6(1):10
Issue Date:
1-Aug-2013
URI:
http://dx.doi.org/10.1186/2046-1682-6-10; http://hdl.handle.net/10033/620948
Type:
Journal Article
Appears in Collections:
publications of the department drug design and optimization (HIPS]DDOP)

Full metadata record

DC FieldValue Language
dc.contributor.authorSteinbach, Ankeen
dc.contributor.authorMaurer, Christine Ken
dc.contributor.authorWeidel, Elisabethen
dc.contributor.authorHenn, Claudiaen
dc.contributor.authorBrengel, Christianen
dc.contributor.authorHartmann, Rolf Wen
dc.contributor.authorNegri, Matthiasen
dc.date.accessioned2017-06-15T08:44:04Z-
dc.date.available2017-06-15T08:44:04Z-
dc.date.issued2013-08-01en
dc.identifier.citationBMC Biophysics. 2013 Aug 01;6(1):10en
dc.identifier.urihttp://dx.doi.org/10.1186/2046-1682-6-10en
dc.identifier.urihttp://hdl.handle.net/10033/620948-
dc.description.abstractAbstract Background PQS (P seudomonas Quinolone Signal) and its precursor HHQ are signal molecules of the P. aeruginosa quorum sensing system. They explicate their role in mammalian pathogenicity by binding to the receptor PqsR that induces virulence factor production and biofilm formation. The enzyme PqsD catalyses the biosynthesis of HHQ. Results Enzyme kinetic analysis and surface plasmon resonance (SPR) biosensor experiments were used to determine mechanism and substrate order of the biosynthesis. Comparative analysis led to the identification of domains involved in functionality of PqsD. A kinetic cycle was set up and molecular dynamics (MD) simulations were used to study the molecular bases of the kinetics of PqsD. Trajectory analysis, pocket volume measurements, binding energy estimations and decompositions ensured insights into the binding mode of the substrates anthraniloyl-CoA and β-ketodecanoic acid. Conclusions Enzyme kinetics and SPR experiments hint at a ping-pong mechanism for PqsD with ACoA as first substrate. Trajectory analysis of different PqsD complexes evidenced ligand-dependent induced-fit motions affecting the modified ACoA funnel access to the exposure of a secondary channel. A tunnel-network is formed in which Ser317 plays an important role by binding to both substrates. Mutagenesis experiments resulting in the inactive S317F mutant confirmed the importance of this residue. Two binding modes for β-ketodecanoic acid were identified with distinct catalytic mechanism preferences.en
dc.titleMolecular basis of HHQ biosynthesis: molecular dynamics simulations, enzyme kinetic and surface plasmon resonance studiesen
dc.typeJournal Articleen
dc.language.rfc3066enen
dc.rights.holderSteinbach et al.; licensee BioMed Central Ltd.en
dc.date.updated2015-09-04T08:24:33Zen
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