21 research outputs found
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Thermophilic Gram-Positive Biocatalysts for Biomass Conversion to Ethanol
Production of energy from renewable sources is receiving increased attention due to the finite nature of fossil fuels and the environmental impact associated with the continued large scale use of fossil energy sources. Biomass, a CO2-neutral abundant resource, is an attractive alternate source of energy. Biomass-derived sugars, such as glucose, xylose, and other minor sugars, can be readily fermented to fuel ethanol and commodity chemicals. Extracellular cellulases produced by fungi are commercially developed for depolymerization of cellulose in biomass to glucose for fermentation by appropriate biocatalysts in a simultaneous saccharification and fermentation (SSF) process. Due to the differences in the optimum conditions for the activity of the fungal cellulases and the growth and fermentation characteristics of the current industrial biocatalysts, SSF of cellulose is envisioned at conditions that are not optimal for the fungal cellulase activity leading to higher than required cost of cellulase in SSF. We have isolated bacterial biocatalysts whose growth and fermentation requirements match the optimum conditions for commercial fungal cellulase activity (pH 5.0 and 50 deg. C). These isolates fermented both glucose and xylose, major components of cellulose and hemicellulose, respectively, to L(+)-lactic acid. Xylose was metabolized through the pentose-phosphate pathway by these organisms as evidenced by the fermentation profile and analysis of the fermentation products of 13C1-xylose by NMR. As expected for the metabolism of xylose by the pentose-phosphate pathway, 13C-lactate accounted for more than 90% of the total 13C-labeled products. All three strains fermented crystalline cellulose to lactic acid with the addition of fungal cellulase (Spezyme CE) (SSF) at an optimum of about 10 FPU/g cellulose. These isolates also fermented cellulose and sugar cane bagasse hemicellulose acid hydrolysate simultaneously. Based on fatty acid profile and 16S rRNA sequence, these isolates cluster with Bacillus coagulans although B. coagulans type strain, ATCC 7050, failed to utilize xylose as a carbon source. For successful production of ethanol from pyruvate, both pyruvate decarboxylase (PDC) and alcohol dehydrogenase (AHD) need to be produced at optimal levels in these biocatalysts. A plasmid containing the S. ventriculi pdc gene and the adh gene from geobacillus stearothermophilus was constructed using plasmid pWH1520 that was successfully used for expression of pdc in B. megaterium. The resulting portable ethanol (PET) plasmid, pJAM423, was transformed into B. megaterium. After xylose induction, a significant fraction of cell cytoplasm was composed of the S. ventriculi PDC and G. stearothermophilus ADH proteins. In preliminary experiments, the amount of ethanol produced by b. megaterium with plasmid pJAM423 was about twice (20 mM) of the bacterium without the plasmid. These results show that the PET operon is functional in B. megaterium but high level ethanol production needs further genetic and metabolic engineering. A genetic transfer system for the second generation biocatalysts needs to be developed for transferring the plasmid pJAM423 and its derivatives for engineering these organisms for ethanol production from biomass derived sugars and cellulose to ethanol. One of the new biocatalysts, strain P4-102B was found to be transformable with plasmids and the method for introducing plasmid pJAM423 into this strain and expression of the encoded DNA is being optimized. These new second generation biocatalysts have the potential to reduce the cost of SSF by minimizing the amount of fungal cellulases, a significant cost component in the use of biomass as a renewable resource for production of fuels and chemicals
A widespread riboswitch candidate that controls bacterial genes involved in molybdenum cofactor and tungsten cofactor metabolism
We have identified a highly conserved RNA motif located upstream of genes encoding molybdate transporters, molybdenum cofactor (Moco) biosynthesis enzymes, and proteins that utilize Moco as a coenzyme. Bioinformatics searches have identified 176 representatives in γ-Proteobacteria, δ-Proteobacteria, Clostridia, Actinobacteria, Deinococcus-Thermus species and DNAs from environmental samples. Using genetic assays, we demonstrate that a Moco RNA in Escherichia coli associated with the Moco biosynthetic operon controls gene expression in response to Moco production. In addition, we provide evidence indicating that this conserved RNA discriminates against closely related analogues of Moco. These results, together with extensive phylogenetic conservation and typical gene control structures near some examples, indicate that representatives of this structured RNA represent a novel class of riboswitches that sense Moco. Furthermore, we identify variants of this RNA that are likely to be triggered by the related tungsten cofactor (Tuco), which carries tungsten in place of molybdenum as the metal constituent
Molecular Factors of Hypochlorite Tolerance in the Hypersaline Archaeon Haloferax volcanii
Halophilic archaea thrive in hypersaline conditions associated with desiccation, ultraviolet (UV) irradiation and redox active compounds, and thus are naturally tolerant to a variety of stresses. Here, we identified mutations that promote enhanced tolerance of halophilic archaea to redox-active compounds using Haloferax volcanii as a model organism. The strains were isolated from a library of random transposon mutants for growth on high doses of sodium hypochlorite (NaOCl), an agent that forms hypochlorous acid (HOCl) and other redox acid compounds common to aqueous environments of high concentrations of chloride. The transposon insertion site in each of twenty isolated clones was mapped using the following: (i) inverse nested two-step PCR (INT-PCR) and (ii) semi-random two-step PCR (ST-PCR). Genes that were found to be disrupted in hypertolerant strains were associated with lysine deacetylation, proteasomes, transporters, polyamine biosynthesis, electron transfer, and other cellular processes. Further analysis revealed a Delta psmA1 (alpha 1) markerless deletion strain that produces only the alpha 2 and beta proteins of 20S proteasomes was hypertolerant to hypochlorite stress compared with wild type, which produces alpha 1, alpha 2, and beta proteins. The results of this study provide new insights into archaeal tolerance of redox active compounds such as hypochlorite
Evaluation of night-time aerosols measurements and lunar irradiance models in the frame of the first multi-instrument nocturnal intercomparison campaign
The first multi-instrument nocturnal aerosol optical depth (AOD) intercomparison campaign was held at the high-mountain Izaña Observatory (Tenerife, Spain) in June 2017, involving 2-min synchronous measurements from two different types of lunar photometers (Cimel CE318-T and Moon Precision Filter Radiometer, LunarPFR) and one stellar photometer. The Robotic Lunar Observatory (ROLO) model developed by the U.S. Geological Survey (USGS) was compared with the open-access ROLO Implementation for Moon photometry Observation (RIMO) model. Results showed rather small differences at Izaña over a 2-month time period covering June and July, 2017 (±0.01 in terms of AOD calculated by means of a day/night/day coherence test analysis and ± 2% in terms of lunar irradiance). The RIMO model has been used in this field campaign to retrieve AOD from lunar
photometric measurements. No evidence of significant differences with the Moon's phase angle was found when comparing raw signals of the six Cimel photometers involved in this field campaign. The raw signal comparison of the participating lunar photometers (Cimel and LunarPFR) performed at coincident wavelengths showed consistent measurements and AOD differences within their combined uncertainties at 870 nm and 675 nm. Slightly larger AOD deviations were observed at 500 nm, pointing to some unexpected instrumental variations during the measurement period. Lunar irradiances retrieved using RIMO for phase angles varying between 0° and 75° (full Moon to near quarter Moon) were compared to the irradiance variations retrieved by Cimel and LunarPFR photometers. Our results showed a relative agreement within ± 3.5% between the RIMO model and the photometer-based lunar irradiances. The AOD retrieved by performing a Langley-plot calibration each night showed a remarkable agreement (better than 0.01) between the lunar photometers. However, when applying the Lunar-Langley calibration using RIMO, AOD differences of up to 0.015 (0.040 for 500 nm) were found, with differences increasing with the Moon's phase angle. These differences are thought to be partly due to the uncertainties in the irradiance models, as well as instrumental deficiencies yet to be fully understood. High AOD variability in stellar measurements was detected during the campaign. Nevertheless, the observed AOD differences in the Cimel/stellar comparison were within the expected combined uncertainties of these two photometric techniques. Our results indicate that lunar photometry is a more reliable technique, especially for low aerosol loading conditions. The uncertainty analysis performed in this paper shows that the combined standard AOD uncertainty in lunar photometry is dependent on the calibration technique (up to 0.014 for Langley-plot with illumination-based correction, 0.012–0.022 for Lunar-Langley calibration, and up to 0.1 for the Sun-Moon Gain Factor method). This analysis also corroborates that the uncertainty of the lunar irradiance model used for AOD calculation is
within the 5–10% expected range. This campaign has allowed us to quantify the important technical difficulties that still exist when routinely monitoring aerosol optical properties at night-time. The small AOD differences observed between the three types of photometers involved in the campaign are only detectable under pristine sky conditions such as those found in this field campaign. Longer campaigns are necessary to understand the observed discrepancies between instruments as well as to provide more conclusive results about the uncertainty involved in the lunar irradiance models.This work has been developed within the framework of the activities of the World Meteorological Organization (WMO) Commission for
Instruments and Methods of Observations (CIMO) Izaña Testbed for Aerosols and Water Vapour Remote Sensing Instruments. AERONET sun
photometers at Izaña have been calibrated within the AERONET Europe TNA, supported by the European Union’s Horizon 2020 research and
innovation program under grant agreement no. 654109 (ACTRIS‒2). CE318-T linearity check has been performed as part of the ESA-funded project “Lunar spectral irradiance measurement and modelling for absolute calibration of EO optical sensors” under ESA contract number:
4000121576/17/NL/AF/hh. LunarPFR has been performing measurements since 2014 in Norway thanks to Svalbard Science Forum funded project, 2014–2016. The authors would like to thank AERONET team for their support and also to NASA’s Navigation and Ancillary Information Facility (NAIF) at the Jet Propulsion Laboratory to help the implementation of the “SPICE” ancillary information system used in this study. We also thank Izaña's ITs for their work to implement the RIMO model in the free-access server. Special thanks should be given to Tom Stone, who has kindly provided us with the USGS/ROLO irradiance values used in the model comparison analysis. This work has also received funding from the European Union’s Horizon 2020 research and innovation programme and from Marie Skłodowska-Curie Individual Fellowships (IF) ACE-GFAT (grant agreement no. 659398). The authors are grateful to Spanish MINECO (CTM2015-66742-R) and Junta de Castilla y León (VA100P17)
Projected WIMP sensitivity of the LUX-ZEPLIN dark matter experiment
LUX-ZEPLIN (LZ) is a next-generation dark matter direct detection experiment that will operate 4850 feet underground at the Sanford Underground Research Facility (SURF) in Lead, South Dakota, USA. Using a two-phase xenon detector with an active mass of 7 tonnes, LZ will search primarily for low-energy interactions with weakly interacting massive particles (WIMPs), which are hypothesized to make up the dark matter in our galactic halo. In this paper, the projected WIMP sensitivity of LZ is presented based on the latest background estimates and simulations of the detector. For a 1000 live day run using a 5.6-tonne fiducial mass, LZ is projected to exclude at 90% confidence level spin-independent WIMP-nucleon cross sections above 1.4 × 10-48cm2 for a 40 GeV/c2 mass WIMP.
Additionally, a 5σ discovery potential is projected, reaching cross sections below the exclusion limits of recent experiments. For spin-dependent WIMP-neutron(-proton) scattering, a sensitivity of 2.3 × 10−43 cm2 (7.1 × 10−42 cm2) for a 40 GeV/c2
mass WIMP is expected. With underground installation well underway, LZ is on track for commissioning at SURF in 2020