Molecular features of uranium-binding natural organic matter in a riparian wetland determined by ultrahigh resolution mass spectrometry.

Tims Branch riparian wetland located in South Carolina, USA has immobilized 94 % of the U released >50 years ago from a nuclear fuel fabrication facility. Sediment organic matter (OM) has been shown to play an important role in immobilizing U. Yet, uranium-OM-mineral interactions at the molecular scale have never been investigated at ambient concentrations. The objectives of this study were to extract, purify, and concentrate U-bound sediment OM along the stream water pathway and perform molecular characterization using Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS). Out of 9614 identified formulas, 715 contained U. These U-containing formulas were enriched with Fe, N, and/or S compared to the total OM. Lignin-like and protein-like molecules accounted for 40 % and 19 % of the U-containing formulas, respectively. Phosphorus-containing formulas were found to exert an insignificant influence on complexing U. U-containing formulas in the 'mobile' (groundwater extractable) OM fraction had lower (reduced) nominal oxidation states of carbon (NOSC); and less aromatic moieties than OM recovered from the 'immobile' (sodium pyrophosphate extractable) OM fraction. U-containing formulas in the redox interfacial zones (stream banks) compared to those in nearby up-slope zones tended to have smaller molecular weights; lower NOSC; higher contents of COO and/or CONO functional groups; and higher abundance of Fe-containing formulas. Fe was present in 38 % of the U-containing formulas but only 20 % of the total OM formulas. It is postulated that Fe played an important role in stabilizing the structure of sedimentary OM, especially U-containing compounds. The identification for the first time of hundreds of Fe-U-OM formulas demonstrates the complexity of such system is much greater than commonly believed and numerically predicting U binding behavior in OM-rich systems may require greater use of statistical or artificial intelligence approaches rather than deterministic approaches limited to measuring metal complexation with well-defined individual analogue organic ligands.

Biosensors for the detection of chorismate and cis,cis-muconic acid in Corynebacterium glutamicum.

Corynebacterium glutamicum ATCC 13032 is a promising microbial chassis for industrial production of valuable compounds, including aromatic amino acids derived from the shikimate pathway. In this work, we developed two whole-cell, transcription factor based fluorescent biosensors to track cis,cis-muconic acid (ccMA) and chorismate in C. glutamicum. Chorismate is a key intermediate in the shikimate pathway from which value-added chemicals can be produced, and a shunt from the shikimate pathway can divert carbon to ccMA, a high value chemical. We transferred a ccMA-inducible transcription factor, CatM, from Acinetobacter baylyi ADP1 into C. glutamicum and screened a promoter library to isolate variants with high sensitivity and dynamic range to ccMA by providing benzoate, which is converted to ccMA intracellularly. The biosensor also detected exogenously supplied ccMA, suggesting the presence of a putative ccMA transporter in C. glutamicum, though the external ccMA concentration threshold to elicit a response was 100-fold higher than the concentration of benzoate required to do so through intracellular ccMA production. We then developed a chorismate biosensor, in which a chorismate inducible promoter regulated by natively expressed QsuR was optimized to exhibit a dose-dependent response to exogenously supplemented quinate (a chorismate precursor). A chorismate-pyruvate lyase encoding gene, ubiC, was introduced into C. glutamicum to lower the intracellular chorismate pool, which resulted in loss of dose dependence to quinate. Further, a knockout strain that blocked the conversion of quinate to chorismate also resulted in absence of dose dependence to quinate, validating that the chorismate biosensor is specific to intracellular chorismate pool. The ccMA and chorismate biosensors were dually inserted into C. glutamicum to simultaneously detect intracellularly produced chorismate and ccMA. Biosensors, such as those developed in this study, can be applied in C. glutamicum for multiplex sensing to expedite pathway design and optimization through metabolic engineering in this promising chassis organism. ONE-SENTENCE SUMMARY: High-throughput screening of promoter libraries in Corynebacterium glutamicum to establish transcription factor based biosensors for key metabolic intermediates in shikimate and ß-ketoadipate pathways.

Presence of aromatic-rich organic matter and its characterization in grout materials: Implications for radionuclide immobilization.

Grout materials are commonly used to immobilize low-level radioactive waste. Organic moieties can be unintentionally present in common ingredients used to make these grout waste forms, which may result in the formation of organo-radionuclide species. These species can positively or negatively affect the immobilization efficiency. However, the presence of organic carbon compounds is rarely considered in models or characterized chemically. Here, we quantify the organic pool of grout formulations with and without slag, as well as the individual dry ingredients used to make the grout samples (ordinary Portland cement (OPC), slag and fly ash), including total organic carbon (TOC) and black carbon, followed by aromaticity evaluation and molecular characterization via Electro Spray Ionization Fourier-Transform Ion Cyclotron Resonance Mass Spectrometry (ESI-FTICRMS). All dry grout ingredients contained significant amounts of organic carbon, ranging from 550 mg/kg to 6250 mg/kg for the TOC pool, with an averaged abundance of 2933 ± 2537 mg/kg, of which 60 ± 29% was composed of black carbon. The significant abundance of a black carbon pool implies the presence of the aromatic-like compounds, which was further identified by both phosphate buffer-assisted aromaticity evaluation (e.g., >1000 mg-C/kg as aromatic-like carbon in the OPC) and dichloromethane (DCM) extraction with ESI-FTICRMS analysis. Besides aromatic-like compounds, other organic moieties were also detected in the OPC, such as carboxyl-containing aliphatic molecules. While the organic compound only consists of minor fractions of the grout materials investigated, our observations of the presence of various radionuclide-binding organic moieties suggests the potential formation of organo-radionuclides, such as radioiodine, which might be present at lower molar concentrations than TOC. Evaluating the role of organic carbon complexation in controlling the disposed radionuclides, especially for those radionuclides with strong association with organic carbon, has important implications for the long-term immobilization of radioactive waste in grout systems.

Iodide uptake by forest soils is principally related to the activity of extracellular oxidases.

129I is a nuclear fission decay product of concern because of its long half-life (16 Ma) and propensity to bioaccumulate. Microorganisms impact iodine mobility in soil systems by promoting iodination (covalent binding) of soil organic matter through processes that are not fully understood. Here, we examined iodide uptake by soils collected at two depths (0-10 and 10-20 cm) from 5 deciduous and coniferous forests in Japan and the United States. Autoclaved soils, and soils amended with an enzyme inhibitor (sodium azide) or an antibacterial agent (bronopol), bound significantly less 125I tracer (93%, 81%, 61% decrease, respectively) than the untreated control soils, confirming a microbial role in soil iodide uptake. Correlation analyses identified the strongest significant correlation between 125I uptake and three explanatory variables, actinobacteria soil biomass (p = 6.04E-04, 1.35E-02 for Kendall-Tau and regression analysis, respectively), soil nitrogen content (p = 4.86E-04, 4.24E-03), and soil oxidase enzyme activity at pH 7.0 using the substrate L-DOPA (p = 2.83E-03, 4.33E-04) and at pH 5.5 using the ABTS (p = 5.09E-03, 3.14E-03). Together, the results suggest that extracellular oxidases, primarily of bacterial origin, are the primary catalyst for soil iodination in aerobic, surface soils of deciduous and coniferous forests, and that soil N content may be indicative of the availability of binding sites for reactive iodine species.

Functional Filaments: Creating and Degrading pH-Indicating PLA Filaments for 3D Printing.

With the rapid pace of advancements in additive manufacturing and techniques such as fused filament fabrication (FFF), the feedstocks used in these techniques should advance as well. While available filaments can be used to print highly customizable parts, the creation of the end part is often the only function of a given feedstock. In this study, novel FFF filaments with inherent environmental sensing functionalities were created by melt-blending poly(lactic acid) (PLA), poly(ethylene glycol) (PEG), and pH indicator powders (bromothymol blue, phenolphthalein, and thymol blue). The new PLA-PEG-indicator filaments were universally more crystalline than the PLA-only filaments (33-41% vs. 19% crystallinity), but changes in thermal stability and mechanical characteristics depended upon the indicator used; filaments containing bromothymol blue and thymol blue were more thermally stable, had higher tensile strength, and were less ductile than PLA-only filaments, while filaments containing phenolphthalein were less thermally stable, had lower tensile strength, and were more ductile. When the indicator-filled filaments were exposed to acidic, neutral, and basic solutions, all filaments functioned as effective pH sensors, though the bromothymol blue-containing filament was only successful as a base indicator. The biodegradability of the new filaments was evaluated by characterizing filament samples after aging in soil and soil slurry mixtures; the amount of physical deterioration and changes in filament crystallinity suggested that the bromothymol blue filament degraded faster than PLA-only filaments, while the phenolphthalein and thymol blue filaments saw decreases in degradation rates.

1H-13C heteronuclear single quantum coherence NMR evidence for iodination of natural organic matter influencing organo-iodine mobility in the environment.

The complex biogeochemical behavior of iodine (I) isotopes and their interaction with natural organic matter (NOM) pose a challenge for transport models. Here, we present results from iodination experiments with humic acid (HA) and fulvic acid (FA) using 1H-13C heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) spectroscopy. Even though not a quantitative approach, 1H-13C HSQC NMR corroborated that iodination of NOM occurs primarily through aromatic electrophilic substitution of proton by I, and also revealed how iodination chemically alters HA and FA in a manner that potentially affects the mobility of iodinated NOM in the environment. Three types of iodination experiments were conducted with HA and FA: a) non-enzymatic iodination by IO3- (pH 3) and I- (pH 4 and 7), b) addition of lactoperoxidase to promote I--iodination in the presence of the co-substrate, H2O2 (pH 7), and c) addition of laccase for facilitating I--iodination in the presence of O2, with or without a mediator (pH 4). When mediators or H2O2 were present, extracellular oxidases and peroxidases enhanced I- incorporation into NOM by between 54% and 3400%. Iodination of HA, which was less than that of FA, enhanced HA's stability (inferred from increases in aliphatic compounds, decreases in carbohydrate moieties, and thus increased molecular hydrophobicity) yet reduced HA's tendency to incorporate more iodine. As such, HA is expected to act more as a sink for iodine in the environment. In contrast, iodination of FA appeared to generate additional iodine binding sites, which resulted in greater iodine uptake capability and enhanced mobility (inferred from decreases in aliphatic compounds, increases in carbohydrates, and thus decreases in molecular hydrophobicity). These results indicate that certain NOM moieties may enhance while others may inhibit radioiodine mobility in the aqueous environment.

Halophyte biorefinery for polyhydroxyalkanoates production from Ulva sp. Hydrolysate with Haloferax mediterranei in pneumatically agitated bioreactors and ultrasound harvesting.

The present study tested the outdoor cultivation of Haloferax mediterranei for PHA production from green macroalgae Ulva sp. in pneumatically agitated bioreactors and applied ultrasonic separation for enhanced settling of archaeal cells. Scaled-up cultivation (40 L) yielded maximum biomass productivity of 50.1 ± 0.11 mg·L-1·h-1 with a PHA productivity of 27 ± 0.01 mg·L-1·h-1 and conversion yield of 0.107 g PHA per gram UlvaDW. The maximum mass fraction of PHA achieved in biomass was calculated to be 56% w/w. Ultrasonic harvesting of Hfx. mediterranei cells approached 30% removal at energy inputs around 7.8 kWh·m-3, and indicated no significant aggregation enhancement by Ca2+ addition. Molecular weight analysis showed an increase in Polydispersity Index (PDI) when the corresponding air velocities were increased suggesting that the polymer was more homogeneous at lower mixing velocities. The current study demonstrated scalable processes for PHA production using Ulva sp. feedstock providing new technologies for halophilic biorefinery.

Large seasonal fluctuations of groundwater radioiodine speciation and concentrations in a riparian wetland in South Carolina.

Recent studies evaluating multiple years of groundwater radioiodine (129I) concentration in a riparian wetland located in South Carolina, USA identified strong seasonal concentration fluctuations, such that summer concentrations were much greater than winter concentrations. These fluctuations were observed only in the wetlands but not in the upland portion of the plume and only with 129I, and not with other contaminants of anthropogenic origin: nitrate/nitrite, strontium-90, technecium-99, tritium, or uranium. This unexplained observation was hypothesized to be the result of strongly coupled processes involving hydrology, water temperature, microbiology, and chemistry. To test this hypothesis, an extensive historical groundwater database was evaluated, and additional measurements of total iodine and iodine speciation were made from recently collected samples. During the summer, the water table decreased by as much as 0.7 m, surface water temperature increased by as much as 15 °C, and total iodine concentrations were consistently greater (up to 680%) than the following winter months. Most of the additional iodine observed in the summer could be attributed to proportional gains in organo-iodine, and not iodide or iodate. Furthermore, 129I concentrations were observed to be two-orders-of-magnitude greater at the bottom of the upland aquifer than at the top. A coupled hydrological and biogeochemical conceptual model is proposed to tie these observations together. First, as the surface water temperature increased during the summer, microbial activity was enhanced, which in turn stimulated the formation of mobile organo-I. Hydrological processes were also likely involved in the observed iodine seasonal changes: (1) as the water table decreased in summer, the remaining upland water entering the wetland was comprised of a greater proportion of water containing elevated iodine concentrations from the low depths, and (2) water flow paths in summer changed such that the wells intercepted more of the contaminant plume and less of the diluting rainwater (due to evapotranspiration) and streamwater (as the lower levels promote a predominantly recharging system). These results underscore the importance of coupled processes influencing contaminant concentrations, and the need to assess seasonal contaminant variations to optimize long-term monitoring programs of wetlands.

An ecophysiological explanation for manganese enrichment in rock varnish.

Desert varnish is a dark rock coating that forms in arid environments worldwide. It is highly and selectively enriched in manganese, the mechanism for which has been a long-standing geological mystery. We collected varnish samples from diverse sites across the western United States, examined them in petrographic thin section using microscale chemical imaging techniques, and investigated the associated microbial communities using 16S amplicon and shotgun metagenomic DNA sequencing. Our analyses described a material governed by sunlight, water, and manganese redox cycling that hosts an unusually aerobic microbial ecosystem characterized by a remarkable abundance of photosynthetic Cyanobacteria in the genus Chroococcidiopsis as the major autotrophic constituent. We then showed that diverse Cyanobacteria, including the relevant Chroococcidiopsis taxon, accumulate extraordinary amounts of intracellular manganese-over two orders of magnitude higher manganese content than other cells. The speciation of this manganese determined by advanced paramagnetic resonance techniques suggested that the Cyanobacteria use it as a catalytic antioxidant-a valuable adaptation for coping with the substantial oxidative stress present in this environment. Taken together, these results indicated that the manganese enrichment in varnish is related to its specific uptake and use by likely founding members of varnish microbial communities.

Development of Antimicrobial PLA Composites for Fused Filament Fabrication.

In addition to possessing the desirable properties of being a biodegradable and biocompatible polymer fabricated from renewable resources, poly (lactic acid) (PLA) has useful mechanical and thermal attributes that has enabled it to be one of the most widely-used plastics for medicine, manufacturing, and agriculture. Yet, PLA composites have not been heavily explored for use in 3D-printing applications, and the range of feasible materials for the technology is limited, which inhibits its potential growth and industry adoption. In this study, tunable, multifunctional antimicrobial PLA composite filaments for 3D-printing have been fabricated and tested via chemical, thermal, mechanical, and antimicrobial experiments. Thermally stable antimicrobial ceramics, ZnO and TiO2, were used as fillers up to 30 wt%, and poly (ethylene glycol) (PEG) was used as a plasticizer to tune the physical material properties. Results demonstrate that the PLA composite filaments exhibit the thermal phase behaviors and thermal stability suitable for 3D-printing. Additionally, PEG can be used to tune the mechanical properties while not affecting the antimicrobial efficacy that ZnO and TiO2 imbue.

Molecular Interaction of Aqueous Iodine Species with Humic Acid Studied by I and C K-Edge X-ray Absorption Spectroscopy.

Iodine-129 is one of three key risk drivers at several US Department of Energy waste management sites. Natural organic matter (NOM) is thought to play important roles in the immobilization of aqueous iodide (I-) and iodate (IO3-) in the environment, but molecular interactions between NOM and iodine species are poorly understood. In this work, we investigated iodine and carbon speciation in three humic acid (HA)-I systems using I K-edge XANES and EXAFS and C K-edge XANES spectroscopy: (1) I- in the presence of laccase (an oxidase enzyme) and a mediator, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) in a pH 4 buffer, (2) I- in the presence of lactoperoxidase (LPO) and H2O2 in a pH 7 buffer, and (3) IO3- in a pH 3 groundwater. Both oxidase and peroxidase systems could oxidize I- to I2 or hypoiodide (HOI) leading to organo-I formation. However, the laccase-ABTS mediator was the most effective and enhanced I- uptake by HA up to 13.5 mg/g, compared to 1.9 mg/g for the LPO-H2O2. IO3- was abiotically reduced to I2 or HOI leading to an organo-I formation. Pathways for HA iodination include covalent modification of aromatic-type rings by I2 / HOI or iodine incorporation into newly formed benzoquinone species arising from the oxidation of phenolic C species. This study improves our molecular-level understanding of NOM-iodine interactions and stresses the important role that mediators may play in the enzymatic reactions between iodine and NOM.

Nagasaki sediments reveal that long-term fate of plutonium is controlled by select organic matter moieties.

Forecasting the long-term fate of plutonium (Pu) is becoming increasingly important as more worldwide military and nuclear-power waste is being generated. Nagasaki sediments containing bomb-derived Pu that was deposited in 1945 provided a unique opportunity to explore the long-term geochemical behavior of Pu. Through a combination of selective extractions and molecular characterization via electrospray ionization Fourier-transform ion cyclotron resonance mass spectrometry (ESI-FTICRMS), we determined that 55 ±â€¯3% of the bomb-derived 239,240Pu was preferentially associated with more persistent organic matter compounds in Nagasaki sediments, particularly those natural organic matter (NOM) stabilized by Fe oxides (NOMFe-oxide). Other organic matter compounds served as a secondary sink of these bomb-derived 239,240Pu (31 ±â€¯2% on average), and <20% of the 239,240Pu was immobilized by inorganic mineral particles. In a narrow, 239,240Pu-enriched layer of only 9-cm depth (total core depth was 600 cm), N-containing carboxyl aliphatic and/or alicyclic molecules (CCAM) in NOMFe-oxide and other NOM fractions immobilized the majority of 239,240Pu. Among the cluster of N-containing CCAM moieties, hydroxamate siderophores, the strongest known Pu chelators in nature, were further detected in these "aged" Nagasaki bomb residue-containing sediments. While present long-term disposal and environmental remediation modeling assume that solubility limits and sorption to mineral surfaces control Pu subsurface mobility, our observations suggest that NOM, which is present in essentially all subsurface systems, undoubtedly plays an important role in sequestrering Pu. Ignoring the role of NOM in controlling Pu fate and transport is not justified in most environmental systems.

Radionuclide uptake by colloidal and particulate humic acids obtained from 14 soils collected worldwide.

Uptake of six particle-reactive and/or redox-sensitive radionuclides (210Pb, 234Th, 7Be, 59Fe, 237Np and 233Pa) by 14 humic acids (HAs) was investigated in artificial groundwater under mildly acidic conditions (pH~5.5). In HA-groundwater slurry, Pb, Be, Fe and Pa bound strongly to particulate HA (>0.45 µm), supporting their application as tracers of soil erosion. Th bound strongly to the colloidal HA (3 kDa-0.45 µm) and as such, would not be a good candidate as a tracer for monitoring soil erosion. HAs likely reduced the oxidized neptunyl form (Np(V)O4+) to Np(IV) based on its enhanced particle-reactivity and Np uptake by particulate HAs, partially retarding the movement of anthropogenic 237Np in field polluted environments. Particulate/colloidal carbonyl/O-aryl (likely through hydroquinone/quinone) functionalities in the HA correlated to Np and Pa uptake, but only particulate O-aryl functionalities was responsible for Fe uptake. The carboxylate- and carbonyl/O-aryl-containing organic functionalities in the HA correlated strongly with Th uptake. In contrast, no significant correlations between organic parameters and Pb or Be uptake implied their predominance of uniform surface adsorption onto particles. This study provides novel insight into the binding of six radionuclides with different organic functionalities of three size fractions, as well as its possible impact on their application in the soil-tracing research.

Microbial Transformation of Iodine: From Radioisotopes to Iodine Deficiency.

Iodine is a biophilic element that is important for human health, both as an essential component of several thyroid hormones and, on the other hand, as a potential carcinogen in the form of radioiodine generated by anthropogenic nuclear activity. Iodine exists in multiple oxidation states (-1, 0, +1, +3, +5, and +7), primarily as molecular iodine (I2), iodide (I-), iodate [Formula: see text] , or organic iodine (org-I). The mobility of iodine in the environment is dependent on its speciation and a series of redox, complexation, sorption, precipitation, and microbial reactions. Over the last 15years, there have been significant advances in iodine biogeochemistry, largely spurred by renewed interest in the fate of radioiodine in the environment. We review the biogeochemistry of iodine, with particular emphasis on the microbial processes responsible for volatilization, accumulation, oxidation, and reduction of iodine, as well as the exciting technological potential of these fascinating microorganisms and enzymes.

Plutonium Partitioning Behavior to Humic Acids from Widely Varying Soils Is Related to Carboxyl-Containing Organic Compounds.

In order to examine the influence of the HA molecular composition on the partitioning of Pu, ten different kinds of humic acids (HAs) of contrasting chemical composition, collected and extracted from different soil types around the world were equilibrated with groundwater at low Pu concentrations (10-14 M). Under mildly acidic conditions (pH ∼ 5.5), 29 ± 24% of the HAs were released as colloidal organic matter (>3 kDa to <0.45 µm), yet this HA fraction accounted for a vast majority of the bound Pu, 76 ± 13% on average. In comparison, the particulate HA fraction bound only 8 ± 4% on average of the added Pu. The truly dissolved Pu fraction was typically <1%. Pu binding was strongly and positively correlated with the concentrations of organic nitrogen in both particulate (>0.45 µm) and colloidal phases in terms of activity percentage and partitioning coefficient values (logKd). Based on molecular characterization of the HAs by solid state 13C nuclear magnetic resonance (NMR) and elemental analysis, Pu binding was correlated to the concentration of carboxylate functionalities and nitrogen groups in the particulate and colloidal phases. The much greater tendency of Pu to bind to colloidal HAs than to particulate HA has implications on whether NOM acts as a Pu source or sink during natural or man-induced episodic flooding.

Polysaccharide Degradation Capability of Actinomycetales Soil Isolates from a Semiarid Grassland of the Colorado Plateau.

Among the bacteria, members of the order Actinomycetales are considered quintessential degraders of complex polysaccharides in soils. However, studies examining complex polysaccharide degradation by Actinomycetales (other than Streptomyces spp.) in soils are limited. Here, we examine the lignocellulolytic and chitinolytic potential of 112 Actinomycetales strains, encompassing 13 families, isolated from a semiarid grassland of the Colorado Plateau in Utah. Members of the Streptomycetaceae, Pseudonocardiaceae, Micromonosporaceae, and Promicromonosporaceae families exhibited robust activity against carboxymethyl cellulose, xylan, chitin, and pectin substrates (except for low/no pectinase activity by the Micromonosporaceae). When incubated in a hydrated mixture of blended Stipa and Hilaria grass biomass over a 5-week period, Streptomyces and Saccharothrix (a member of the Pseudonocardiaceae) isolates produced high levels of extracellular enzyme activity, such as endo- and exocellulase, glucosidase, endo- and exoxylosidase, and arabinofuranosidase. These characteristics make them well suited to degrade the cellulose and hemicellulose components of grass cell walls. On the basis of the polysaccharide degradation profiles of the isolates, relative abundance of Actinomycetales sequences in 16S rRNA gene surveys of Colorado Plateau soils, and analysis of genes coding for polysaccharide-degrading enzymes among 237 Actinomycetales genomes in the CAZy database and 5 genomes from our isolates, we posit that Streptomyces spp. and select members of the Pseudonocardiaceae and Micromonosporaceae likely play an important role in the degradation of hemicellulose, cellulose, and chitin substances in dryland soils.IMPORTANCE Shifts in the relative abundance of Actinomycetales taxa have been observed in soil microbial community surveys during large, manipulated climate change field studies. However, our limited understanding of the ecophysiology of diverse Actinomycetales taxa in soil systems undermines attempts to determine the underlying causes of the population shifts or their impact on carbon cycling in soil. This study combines a systematic analysis of the polysaccharide degradation potential of a diverse collection of Actinomycetales isolates from surface soils of a semiarid grassland with analysis of genomes from five of these isolates and publicly available Actinomycetales genomes for genes encoding polysaccharide-active enzymes. The results address an important gap in knowledge of Actinomycetales ecophysiology-identification of key taxa capable of facilitating lignocellulose degradation in dryland soils. Information from this study will benefit future metagenomic studies related to carbon cycling in dryland soils by providing a baseline linkage of Actinomycetales phylogeny with lignocellulolytic functional potential.

Role of natural organic matter on iodine and (239)(,240)Pu distribution and mobility in environmental samples from the northwestern Fukushima Prefecture, Japan.

In order to assess how environmental factors are affecting the distribution and migration of radioiodine and plutonium that were emitted from the Fukushima Dai-ichi Nuclear Power Plant (FDNPP) accident, we quantified iodine and (239,240)Pu concentration changes in soil samples with different land uses (urban, paddy, deciduous forest and coniferous forest), as well as iodine speciation in surface water and rainwater. Sampling locations were 53-63 km northwest of the FDNPP within a 75-km radius, in close proximity of each other. A ranking of the land uses by their surface soil (<4 cm) stable (127)I concentrations was coniferous forest > deciduous forest > urban > paddy, and (239,240)Pu concentrations ranked as deciduous forest > coniferous forest > paddy ≥ urban. Both were quite distinct from that of (134)Cs and (137)Cs: urban > coniferous forest > deciduous forest > paddy, indicating differences in their sources, deposition phases, and biogeochemical behavior in these soil systems. Although stable (127)I might not have fully equilibrated with Fukushima-derived (129)I, it likely still works as a proxy for the long-term fate of (129)I. Surficial soil (127)I content was well correlated to soil organic matter (SOM) content, regardless of land use type, suggesting that SOM might be an important factor affecting iodine biogeochemistry. Other soil chemical properties, such as Eh and pH, had strong correlations to soil (127)I content, but only within a given land use (e.g., within urban soils). Organic carbon (OC) concentrations and Eh were positively, and pH was negatively correlated to (127)I concentrations in surface water and rain samples. It is also noticeable that (127)I in the wet deposition was concentrated in both the deciduous and coniferous forest throughfall and stemfall water, respectively, comparing to the bulk rainwater. Further, both forest throughfall and stemflow water consisted exclusively of organo-iodine, suggesting all inorganic iodine in the original bulk deposition (∼ 28.6% of total iodine) have been completely converted to organo-iodine. Fukushima-derived (239,240)Pu was detectable at a distance ∼ 61 km away, NW of FDNPP. However, it is confined to the litter layer, even three years after the FDNPP accident-derived emissions. Plutonium-239,240 activities were significantly correlated with soil OC and nitrogen contents, indicating Pu may be associated with nitrogen-containing SOM, similar to what has been observed at other locations in the United States. Together, these finding suggest that natural organic matter (NOM) plays a key role in affecting the fate and transport of I and Pu and may warrant greater consideration for predicting long-term stewardship of contaminated areas and evaluating various remediation options in Japan.

Activity-Based Protein Profiling of Ammonia Monooxygenase in Nitrosomonas europaea.

Nitrosomonas europaea is an aerobic nitrifying bacterium that oxidizes ammonia (NH3) to nitrite (NO2 (-)) through the sequential activities of ammonia monooxygenase (AMO) and hydroxylamine dehydrogenase (HAO). Many alkynes are mechanism-based inactivators of AMO, and here we describe an activity-based protein profiling method for this enzyme using 1,7-octadiyne (17OD) as a probe. Inactivation of NH4 (+)-dependent O2 uptake by N. europaea by 17OD was time- and concentration-dependent. The effects of 17OD were specific for ammonia-oxidizing activity, andde novoprotein synthesis was required to reestablish this activity after cells were exposed to 17OD. Cells were reacted with Alexa Fluor 647 azide using a copper-catalyzed azide-alkyne cycloaddition (CuAAC) (click) reaction, solubilized, and analyzed by SDS-PAGE and infrared (IR) scanning. A fluorescent 28-kDa polypeptide was observed for cells previously exposed to 17OD but not for cells treated with either allylthiourea or acetylene prior to exposure to 17OD or for cells not previously exposed to 17OD. The fluorescent polypeptide was membrane associated and aggregated when heated with ß-mercaptoethanol and SDS. The fluorescent polypeptide was also detected in cells pretreated with other diynes, but not in cells pretreated with structural homologs containing a single ethynyl functional group. The membrane fraction from 17OD-treated cells was conjugated with biotin-azide and solubilized in SDS. Streptavidin affinity-purified polypeptides were on-bead trypsin-digested, and amino acid sequences of the peptide fragments were determined by liquid chromatography-mass spectrometry (LC-MS) analysis. Peptide fragments from AmoA were the predominant peptides detected in 17OD-treated samples. In-gel digestion and matrix-assisted laser desorption ionization-tandem time of flight (MALDI-TOF/TOF) analyses also confirmed that the fluorescent 28-kDa polypeptide was AmoA.

Modulation of FadR binding capacity for acyl-CoA fatty acids through structure-guided mutagenesis.

FadR is a versatile global regulator in Escherichia coli that controls fatty acid metabolism and thereby modulates the ability of this bacterium to grow using fatty acids or acetate as the sole carbon source. FadR regulates fatty acid metabolism in response to intra-cellular concentrations of acyl-CoA lipids. The ability of FadR to bind acyl-CoA fatty acids is thus of significant interest for the engineering of biosynthetic pathways for the production of lipid-based biofuels and commodity chemicals. Based on the available crystal structure of E. coli bound to myristoyl-CoA, we predicted amino acid positions within the effector binding pocket that would alter the ability of FadR to bind acyl-CoA fatty acids without affecting DNA binding. We utilized fluorescence polarization to characterize the in vitro binding properties of wild type and mutant FadR. We found that a Leu102Ala mutant enhanced binding of the effector, likely by increasing the size of the binding pocket for the acyl moiety of the molecule. Conversely, the elimination of the guanidine side chain (Arg213Ala and Arg213Met mutants) of the CoA moiety binding site severely diminished the ability of FadR to bind the acyl-CoA effector. These results demonstrate the ability to fine tune FadR binding capacity. The validation of an efficient method to fully characterize all the binding events involved in the specific activity (effector and DNA operator binding) of FadR has allowed us to increase our understanding of the role of specific amino acids in the binding and recognition of acyl-CoA fatty acids and will greatly facilitate efforts aimed at engineering tunable FadR regulators for synthetic biology.

Radioiodine sorption/desorption and speciation transformation by subsurface sediments from the Hanford Site.

During the last few decades, considerable research efforts have been extended to identify more effective remediation treatment technologies to lower the (129)I concentrations to below federal drinking water standards at the Hanford Site (Richland, USA). Few studies have taken iodate into consideration, though recently iodate, instead of iodide, was identified as the major species in the groundwater of 200-West Area within the Hanford Site. The objective of this study was thus to quantify and understand aqueous radioiodine species transformations and uptake by three sediments collected from the semi-arid, carbonate-rich environment of the Hanford subsurface. All three sediments reduced iodate (IO3(-)) to iodide (I(-)), but the loamy-sand sediment reduced more IO3(-) (100% reduced within 7 days) than the two sand-textured sediments (∼20% reduced after 28 days). No dissolved organo-iodine species were observed in any of these studies. Iodate uptake Kd values ([Isolid]/[Iaq]; 0.8-7.6 L/kg) were consistently and appreciably greater than iodide Kd values (0-5.6 L/kg). Furthermore, desorption Kd values (11.9-29.8 L/kg) for both iodate and iodide were consistently and appreciably greater than uptake Kd values (0-7.6 L/kg). Major fractions of iodine associated with the sediments were unexpectedly strongly bound, such that only 0.4-6.6 % of the total sedimentary iodine could be exchanged from the surface with KCl solution, and 0-1.2% was associated with Fe or Mn oxides (weak NH2HCl/HNO3 extractable fraction). Iodine incorporated into calcite accounted for 2.9-39.4% of the total sedimentary iodine, whereas organic carbon (OC) is likely responsible for the residual iodine (57.1-90.6%) in sediments. The OC, even at low concentrations, appeared to be controlling iodine binding to the sediments, as it was found that the greater the OC concentrations in the sediments, the greater the values of uptake Kd, desorption Kd, and the greater residual iodine concentrations (non-exchangeable, non-calcite-incorporated and non-Mn, Fe-oxide associated). This finding is of particular interest because it suggests that even very low OC concentrations, <0.2%, may have an impact on iodine geochemistry. The findings that these sediments can readily reduce IO3(-), and that IO3(-) sorbs to a greater extent than I(-), sheds light into earlier unexplained Hanford field data that demonstrated increases in groundwater (127)I(-)/(127)IO3(-) ratios and a decrease groundwater (129)IO3(-) concentrations along a transect away from the point sources, where iodine was primarily introduced as IO3(-). While a majority of the radioiodine does not bind to these alkaline sediments, there is likely a second smaller iodine fraction in the Hanford subsurface that is strongly bound, presumably to the sediment OC (and carbonate) phases. This second fraction may have an impact on establishing remediation goals and performance assessment calculations.