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Promoting uranium immobilization by the activities of microbial phophatases

Description: The first objective of this project is to determine the relationship of phosphatase activity to metal resistance in subsurface strains and the role of lateral gene transfer (LGT) in dissemination of nonspecific acid phosphatase genes. Nonspecific acid phosphohydrolases are a broad group of secreted microbial phosphatases that function in acidic-to-neutral pH ranges and utilize a wide range of organophosphate substrates. We have previously shown that PO43- accumulation during growth on a model organophosphorus compound was attributable to the overproduction of alkaline phosphatase by genetically modified subsurface pseudomonads [Powers et al. (2002) FEMS Microbiol. Ecol. 41:115-123]. During this report period, we have extended these results to include indigenous metal resistant subsurface microorganisms cultivated from the Field Research Center (FRC), in Oak Ridge Tennessee.
Date: June 1, 2005
Creator: Sobecky, Patricia A.
Partner: UNT Libraries Government Documents Department

Final Technical Report

Description: This final technical report describes results and findings from a research project to examine the role of microbial phosphohydrolase enzymes in naturally occurring subsurface microorganisms for the purpose of promoting the immobilization of the radionuclide uranium through the production of insoluble uranium phosphate minerals. The research project investigated the microbial mechanisms and the physical and chemical processes promoting uranium biomineralization and sequestration in oxygenated subsurface soils. Uranium biomineralization under aerobic conditions can provide a secondary biobarrier strategy to immobilize radionuclides should the metal precipitates formed by microbial dissimilatory mechanisms remobilize due to a change in redox state.
Date: March 29, 2013
Creator: Sobecky, Patricia A & Taillefert, Martial
Partner: UNT Libraries Government Documents Department

Promoting uranium immobilization by the activities of microbial phophatases

Description: The following is a summary of progress in our project ''Promoting uranium immobilization by the activities of microbial phosphatases'' during the second year of the project. (1). Assignment of microbial phosphatases to molecular classes. One objective of this project is to determine the relationship of phosphatase activity to metal resistance in subsurface strains and possible contributions of horizontal gene transfer (HGT) to the dissemination of nonspecific acid phosphatase genes. Non-specific acid phosphohydrolases are a broad group of secreted microbial phosphatases that function in acidic-to-neutral pH ranges and utilize a wide range of organophosphate substrates. To address this objective we have designed a collection of PCR primer sets based on known microbial acid phosphatase sequences. Genomic DNA is extracted from subsurface FRC isolates and amplicons of the expected sizes are sequenced and searched for conserved signature motifs. During this reporting period we have successfully designed and tested a suite of PCR primers for gram-positive and gram-negative groups of the following phosphatase classes: (1) Class A; (2) Class B; and (3) Class C (gram negative). We have obtained specific PCR products for each of the classes using the primers we have designed using control strains as well as with subsurface isolates.
Date: June 1, 2006
Creator: Sobecky, Patricia A. & Taillefert, Martial
Partner: UNT Libraries Government Documents Department

Promoting Uranium Immobilization by the Activities of Microbial Phosphatases

Description: The overall objective of this project is to examine the activity of nonspecific phosphohydrolases present in naturally occurring subsurface microorganisms for the purpose of promoting the immobilization of radionuclides through the production of uranium [U(VI)] phosphate precipitates. Specifically, we hypothesize that the precipitation of U(VI) phosphate minerals may be promoted through the microbial release and/or accumulation of PO4 3- as a means to detoxify radionuclides and heavy metals. An experimental approach was designed to determine the extent of phosphatase activity in bacteria previously isolated from contaminated subsurface soils collected at the ERSP Field Research Center (FRC) in Oak Ridge, TN. Screening of 135 metal resistant isolates for phosphatase activity indicated the majority (75 of 135) exhibited a phosphatase-positive phenotype. During this phase of the project, a PCR based approach has also been designed to assay FRC isolates for the presence of one or more classes of the characterized non-specific acid phophastase (NSAP) genes likely to be involved in promoting U(VI) precipitation. Testing of a subset of Pb resistant (Pbr) Arthrobacter, Bacillus and Rahnella strains indicated 4 of the 9 Pbr isolates exhibited phosphatase phenotypes suggestive of the ability to bioprecipitate U(VI). Two FRC strains, a Rahnella sp. strain Y9602 and a Bacillus sp. strain Y9-2, were further characterized. The Rahnella sp. exhibited enhanced phosphatase activity relative to the Bacillus sp. Whole-cell enzyme assays identified a pH optimum of 5.5, and inorganic phosphate accumulated in pH 5.5 synthetic groundwater (designed to mimic FRC conditions) incubations of both strains in the presence of a model organophosphorus substrate provided as the sole C and P source. Kinetic experiments showed that these two organisms can grow in the presence of 200 μM dissolved uranium and that Rahnella is much more efficient in precipitating U(VI) than Bacillus sp. The precipitation of U(VI) must be mediated ...
Date: April 19, 2007
Creator: Martinez, Robert J.; Beazley, Melanie J.; Webb, Samuel M.; Taillefert, Martial & Sobecky, Patricia A.
Partner: UNT Libraries Government Documents Department