Evidence supporting dissimilatory and assimilatory lignin degradation in Enterobacter lignolyticus SCF1

Lignocellulosic biofuels are promising as sustainable alternative fuels, but lignin inhibits access of enzymes to cellulose, and by-products of lignin degradation can be toxic to cells. The fast growth, high efficiency and specificity of enzymes employed in the anaerobic litter deconstruction carried out by tropical soil bacteria make these organisms useful templates for improving biofuel production. The facultative anaerobe Enterobacter lignolyticus SCF1 was initially cultivated from Cloud Forest soils in the Luquillo Experimental Forest in Puerto Rico, based on anaerobic growth on lignin as sole carbon source. The source of the isolate was tropical forest soils that decompose litter rapidly with low and fluctuating redox potentials, where bacteria using oxygen-independent enzymes likely play an important role in decomposition. We have used transcriptomics and proteomics to examine the observed increased growth of SCF1 grown on media amended with lignin compared to unamended growth. Proteomics suggested accelerated xylose uptake and metabolism under lignin-amended growth, with up-regulation of proteins involved in lignin degradation via the 4-hydroxyphenylacetate degradation pathway, catalase/peroxidase enzymes, and the glutathione biosynthesis and glutathione S-transferase (GST) proteins. We also observed increased production of NADH-quinone oxidoreductase, other electron transport chain proteins, and ATP synthase and ATP-binding cassette (ABC) transporters. This suggested the use of lignin as terminal electron acceptor. We detected significant lignin degradation over time by absorbance, and also used metabolomics to demonstrate moderately significant decreased xylose concentrations as well as increased metabolic products acetate and formate in stationary phase in lignin-amended compared to unamended growth conditions. Our data show the advantages of a multi-omics approach toward providing insights as to how lignin may be used in nature by microorganisms coping with poor carbon availability

FY-2007 PNNL Voluntary Protection Program (VPP) Program Evaluation

This document reports the results of the FY-2007 PNNL VPP Program Evaluation, which is a self-assessment of the operational and programmatic performance of the Laboratory related to worker safety and health. The report was compiled by a team of worker representatives and safety professionals who evaluated the Laboratory's worker safety and health programs on the basis of DOE-VPP criteria. The principle elements of DOE's VPP program are: Management Leadership, Employee Involvement, Worksite Analysis, Hazard Prevention and Control, and Safety and Health Training

Le Peuple : organe quotidien du syndicalisme

27 mars 19391939/03/27 (A19,N6637)-1939/03/27

The weak interdomain coupling observed in the 70 kDa subunit of human replication protein A is unaffected by ssDNA binding

Replication protein A (RPA) is a heterotrimeric, multi-functional protein that binds single-stranded DNA (ssDNA) and is essential for eukaryotic DNA metabolism. Using heteronuclear NMR methods we have investigated the domain interactions and ssDNA binding of a fragment from the 70 kDa subunit of human RPA (hRPA70). This fragment contains an N-terminal domain (NTD), which is important for hRPA70–protein interactions, connected to a ssDNA-binding domain (SSB1) by a flexible linker (hRPA70 1–326 ). Correlation analysis of the amide 1 H and 15 N chemical shifts was used to compare the structure of the NTD and SSB1 in hRPA70 1–326 with two smaller fragments that corresponded to the individual domains. High correlation coefficients verified that the NTD and SSB1 maintained their structures in hRPA70 1–326 , indicating weak interdomain coupling. Weak interdomain coupling was also suggested by a comparison of the transverse relaxation rates for hRPA70 1–326 and one of the smaller hRPA70 fragments containing the NTD and the flexible linker (hRPA70 1–168 ). We also examined the structure of hRPA70 1–326 after addition of three different ssDNA substrates. Each of these substrates induced specific amide 1 H and/or 15 N chemical shift changes in both the NTD and SSB1. The NTD and SSB1 have similar topologies, leading to the possibility that ssDNA binding induced the chemical shift changes observed for the NTD. To test this hypothesis we monitored the amide 1 H and 15 N chemical shift changes of hRPA70 1–168 after addition of ssDNA. The same amide 1 H and 15 N chemical shift changes were observed for the NTD in hRPA70 1–168 and hRPA70 1–326 . The NTD residues with the largest amide 1 H and/or 15 N chemical shift changes were localized to a basic cleft that is important for hRPA70–protein interactions. Based on this relationship, and other available data, we propose a model where binding between the NTD and ssDNA interferes with hRPA70–protein interactions

Solubilization and upgrading of high polyethylene terephthalate loadings in a low-costing bifunctional ionic liquid

The solubilization and efficient upgrading of high loadings of polyethylene terephthalate (PET) are important challenges, and most solvents for PET are highly toxic. Herein, a low-cost (ca. $1.2 kg) and biocompatible ionic liquid (IL), cholinium phosphate ([Ch][PO]), is demonstrated for the first time to play bifunctional roles in the solubilization and glycolytic degradation of PET. A high loading of PET (10 wt %) was readily dissolved in [Ch][PO] at relatively low temperatures (120 °C, 3 h) and under water-rich conditions. In-depth analysis of the solution revealed that high PET solubilization in [Ch][PO] could be ascribed to significant PET depolymerization. Acid precipitation yielded terephthalic acid as the dominant depolymerized monomer with a theoretical yield of approximately 95 %. Further exploration showed that in the presence of ethylene glycol (EG), the [Ch][PO]-catalyzed glycolysis of PET could efficiently occur with approximately 100 % conversion of PET and approximately 60.6 % yield of bis(2-hydroxyethyl)terephthalate under metal-free conditions. The IL could be reused at least three times without an apparent decrease in activity. NMR spectroscopy analysis revealed that strong hydrogen-bonding interactions between EG and the IL played an important role in the activation of EG and promotion of the glycolysis reaction. This study opens up avenues for exploring environmentally benign and efficient IL technology for solubilizing and recycling postconsumer polyester plastics

Formaldehyde as a Carbon and Electron Shuttle between Autotroph and Heterotroph Populations in Acidic Hydrothermal Vents of Norris Geyser Basin, Yellowstone National Park

The Norris Geyser Basin in Yellowstone National Park contains a large number of hydrothermal systems, which host microbial populations supported by primary productivity associated with a suite of chemolithotrophic metabolisms. We demonstrate that Metallosphaera yellowstonensis MK1, a facultative autotrophic archaeon isolated from a hyperthermal acidic hydrous ferric oxide (HFO) spring in Norris Geyser Basin, excretes formaldehyde during autotrophic growth. To determine the fate of formaldehyde in this low organic carbon environment, we incubated native microbial mat (containing M. yellowstonensis) from a HFO spring with C-13-formaldehyde. Isotopic analysis of incubation-derived CO2 and biomass showed that formaldehyde was both oxidized and assimilated by members of the community. Autotrophy, formaldehyde oxidation, and formaldehyde assimilation displayed different sensitivities to chemical inhibitors, suggesting that distinct sub-populations in the mat selectively perform these functions. Our results demonstrate that electrons originally resulting from iron oxidation can energetically fuel autotrophic carbon fixation and associated formaldehyde excretion, and that formaldehyde is both oxidized and assimilated by different organisms within the native microbial community. Thus, formaldehyde can effectively act as a carbon and electron shuttle connecting the autotrophic, iron oxidizing members with associated heterotrophic members in the HFO community

Proteome specialization of anaerobic fungi during ruminal degradation of recalcitrant plant fiber

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