title: Evidence that bacterial ABC-type transporter imports free EDTA for metabolism.
abstract: EDTA, a common chelating agent, is becoming a major organic pollutant in the form of metal-EDTA complexes in surface waters, partly due to its recalcitrance to biodegradation. Even an EDTA-degrading bacterium, BNC1, does not degrade stable metal-EDTA complexes. In the present study, an ABC-type transporter was identified for possible uptake of EDTA because the transporter genes and the EDTA monooxygenase gene were expressed from a single operon in BNC1. The ABC-type transporter had a periplasmic-binding protein (EppA) that should confer the substrate specificity for the transporter; therefore, EppA was produced in Escherichia coli, purified, and characterized. EppA was shown to bind free EDTA with a dissociation constant as low as 25 nM by using isothermal titration calorimetry. When unstable metal-EDTA complexes, e.g., (Mg-EDTA)(2-), were added to the EppA solution, binding was also observed. However, experimental data and theoretical analysis supported EppA binding only of free EDTA. When stable metal-EDTA complexes, e.g., (Cu-EDTA)(2-), were titrated into the EppA solution, no binding was observed. Since EDTA monooxygenase in the cytoplasm uses some of the stable metal-EDTA complexes as substrates, we suggest that the lack of EppA binding and EDTA uptake are responsible for the failure of BNC1 cells to degrade the stable complexes.
authors.affiliation: School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-4234, USA.
authors.forename: Hua
authors.initials: H
authors.lastname: Zhang
authors.forename: Jacob P
authors.initials: JP
authors.lastname: Herman
authors.forename: Harvey
authors.initials: H
authors.lastname: Bolton
authors.forename: Zhicheng
authors.initials: Z
authors.lastname: Zhang
authors.forename: Sue
authors.initials: S
authors.lastname: Clark
authors.forename: Luying
authors.initials: L
authors.lastname: Xun
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D004492:Edetic Acid / Q000737:chemistry / Q000378:metabolism*; D008670:Metals / Q000737:chemistry / Q000378:metabolism; D033902:Periplasmic Binding Proteins / Q000235:genetics / Q000378:metabolism; D012230:Rhizobiaceae / Q000378:metabolism*; D013997:Time Factors; D014874:Water Pollutants, Chemical / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D008670:Metals; D033902:Periplasmic Binding Proteins; D014874:Water Pollutants, Chemical; D004492:Edetic Acid