title: Characterization of a phosphotriesterase-like lactonase from Sulfolobus solfataricus and its immobilization for disruption of quorum sensing.
abstract: SsoPox, a bifunctional enzyme with organophosphate hydrolase and N-acyl homoserine lactonase activities from the hyperthermophilic archaeon Sulfolobus solfataricus, was overexpressed and purified from recombinant Pseudomonas putida KT2440 with a yield of 9.4 mg of protein per liter of culture. The enzyme has a preference for N-acyl homoserine lactones (AHLs) with acyl chain lengths of at least 8 carbon atoms, mainly due to lower K(m) values for these substrates. The highest specificity constant obtained was for N-3-oxo-decanoyl homoserine lactone (k(cat)/K(m) = 5.5 × 10(3) M(-1)·s(-1)), but SsoPox can also degrade N-butyryl homoserine lactone (C(4)-HSL) and N-oxo-dodecanoyl homoserine lactone (oxo-C(12)-HSL), which are important for quorum sensing in our Pseudomonas aeruginosa model system. When P. aeruginosa PAO1 cultures were grown in the presence of SsoPox-immobilized membranes, the production of C(4)-HSL- and oxo-C(12)-HSL-regulated virulence factors, elastase, protease, and pyocyanin were significantly reduced. This is the first demonstration that immobilized quorum-quenching enzymes can be used to attenuate the production of virulence factors controlled by quorum-sensing signals.
authors.affiliation: Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada. sseah@uoguelph.ca.
authors.forename: Filomena S W
authors.initials: FS
authors.lastname: Ng
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Wright
authors.forename: Stephen Y K
authors.initials: SY
authors.lastname: Seah
mesh_terms: D015107:4-Butyrolactone / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004800:Enzymes, Immobilized / Q000378:metabolism; D015870:Gene Expression; D007783:Lactones / Q000737:chemistry / Q000378:metabolism; D010196:Pancreatic Elastase / Q000096:biosynthesis; D010447:Peptide Hydrolases / Q000096:biosynthesis; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism; D044345:Phosphoric Triester Hydrolases / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011550:Pseudomonas aeruginosa / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism* / Q000472:pathogenicity; D011710:Pyocyanine / Q000096:biosynthesis; D053038:Quorum Sensing; D017422:Sequence Analysis, DNA; D015398:Signal Transduction; D048229:Sulfolobus solfataricus / Q000201:enzymology* / Q000235:genetics; D037521:Virulence Factors / Q000096:biosynthesis
chemical_list: D004800:Enzymes, Immobilized; D007783:Lactones; C092312:N-butyrylhomoserine lactone; D037521:Virulence Factors; D011710:Pyocyanine; D002265:Carboxylic Ester Hydrolases; C432003:N-acyl homoserine lactonase; C045290:phosphorylphosphatase; D010744:Phosphoric Monoester Hydrolases; D044345:Phosphoric Triester Hydrolases; D010447:Peptide Hydrolases; D010196:Pancreatic Elastase; D015107:4-Butyrolactone