Investigation of off-site airborne transport of lead from a superfund removal action site using lead isotope ratios and concentrations

USGS Publications Warehouse

Pribil, Michael J.; Maddaloni, Mark A.; Staiger, Kimberly; Wilson, Eric; Magriples, Nick; Ali, Mustafa; Santella, Dennis

2014-01-01

Lead (Pb) concentration and Pb isotopic composition of surface and subsurface soil samples were used to investigate the potential for off-site air transport of Pb from a former white Pb processing facility to neighboring residential homes in a six block area on Staten Island, NY. Surface and subsurface soil samples collected on the Jewett White Pb site were found to range from 1.122 to 1.138 for 206Pb/207Pb and 2.393 to 2.411 for 208Pb/207Pb. The off-site surface soil samples collected from residential backyards, train trestle, near site grass patches and background areas varied from 1.144 to 1.196 for 206Pb/207Pb and 2.427 to 2.464 for 208Pb/207Pb. Two soil samples collected along Richmond Terrace, where Jewett site soils accumulated after major rain events, varied from 1.136 to 1.147 for 206Pb/207Pb and 2.407 to 2.419 for 208Pb/207Pb. Lead concentration for on-site surface soil samples ranged from 450 to 8000 ug/g, on-site subsurface soil samples ranged from 90,000 to 240,000 ug/g and off-site samples varied from 380 to 3500 ug/g. Lead concentration and isotopic composition for the Staten Island off-site samples were similar to previously published data for other northeastern US cities and reflect re-suspension and re-mobilization of local accumulated Pb. The considerable differences in both the Pb isotopic composition and Pb concentration of on-site and off-site samples resulted in the ability to geochemically trace the transport of particulate Pb. Data in this study indicate minimal off-site surface transport of Pb from the Jewett site into the neighboring residential area.

Channeling by Proximity: The Catalytic Advantages of Active Site Colocalization Using Brownian Dynamics.

PubMed

Bauler, Patricia; Huber, Gary; Leyh, Thomas; McCammon, J Andrew

2010-05-06

Nature often colocalizes successive steps in a metabolic pathway. Such organization is predicted to increase the effective concentration of pathway intermediates near their recipient active sites and to enhance catalytic efficiency. Here, the pathway of a two-step reaction is modeled using a simple spherical approximation for the enzymes and substrate particles. Brownian dynamics are used to simulate the trajectory of a substrate particle as it diffuses between the active site zones of two different enzyme spheres. The results approximate distances for the most effective reaction pathways, indicating that the most effective reaction pathway is one in which the active sites are closely aligned. However, when the active sites are too close, the ability of the substrate to react with the first enzyme was hindered, suggesting that even the most efficient orientations can be improved for a system that is allowed to rotate or change orientation to optimize the likelihood of reaction at both sites.

Characteristics of Zinc Phosphate Coating Activated by Different Concentrations of Nickel Acetate Solution

NASA Astrophysics Data System (ADS)

Abdalla, Khalid; Zuhailawati, H.; Rahmat, Azmi; Azizan, A.

2017-02-01

Activation pretreatment with nickel acetate solution at various concentrations was performed prior to the phosphating step to enhance the corrosion resistance of carbon steel substrates. The activation solution was studied over various concentrations: 10, 50, and 100 g/L. The effects of these concentrations on surface characteristics and microstructural evolution of the coated samples were characterized by scanning electron microscopy and energy-dispersive spectroscopy. The electrochemical behavior was evaluated using potentiodynamic polarization curves, electrochemical impedance spectroscopy, and immersion test in a 3.5 pct NaCl solution. Significant increases in the nucleation sites and surface coverage of zinc phosphate coating were observed as the concentration of activation solution reached 50 g/L. The electrochemical analysis revealed that the activation treatment with 50 g/L nickel acetate solution significantly improved the protection ability of the zinc phosphate coating. The corrosion current density of activated phosphate coating with 50 g/L was reduced by 64.64 and 13.22 pct, compared to the coatings obtained with activation solutions of 10 and 100 g/L, respectively.

Effect of particle surface area on ice active site densities retrieved from droplet freezing spectra

NASA Astrophysics Data System (ADS)

Beydoun, Hassan; Polen, Michael; Sullivan, Ryan C.

2016-10-01

Heterogeneous ice nucleation remains one of the outstanding problems in cloud physics and atmospheric science. Experimental challenges in properly simulating particle-induced freezing processes under atmospherically relevant conditions have largely contributed to the absence of a well-established parameterization of immersion freezing properties. Here, we formulate an ice active, surface-site-based stochastic model of heterogeneous freezing with the unique feature of invoking a continuum assumption on the ice nucleating activity (contact angle) of an aerosol particle's surface that requires no assumptions about the size or number of active sites. The result is a particle-specific property g that defines a distribution of local ice nucleation rates. Upon integration, this yields a full freezing probability function for an ice nucleating particle. Current cold plate droplet freezing measurements provide a valuable and inexpensive resource for studying the freezing properties of many atmospheric aerosol systems. We apply our g framework to explain the observed dependence of the freezing temperature of droplets in a cold plate on the concentration of the particle species investigated. Normalizing to the total particle mass or surface area present to derive the commonly used ice nuclei active surface (INAS) density (ns) often cannot account for the effects of particle concentration, yet concentration is typically varied to span a wider measurable freezing temperature range. A method based on determining what is denoted an ice nucleating species' specific critical surface area is presented and explains the concentration dependence as a result of increasing the variability in ice nucleating active sites between droplets. By applying this method to experimental droplet freezing data from four different systems, we demonstrate its ability to interpret immersion freezing temperature spectra of droplets containing variable particle concentrations. It is shown that general

Hourly elemental concentrations in PM2.5 aerosols sampled simultaneously at urban background and road site

NASA Astrophysics Data System (ADS)

Dall'Osto, M.; Querol, X.; Amato, F.; Karanasiou, A.; Lucarelli, F.; Nava, S.; Calzolai, G.; Chiari, M.

2012-08-01

aerosol plumes associated with shipping oil combustion and smelters activities have a higher impact on the more ventilated UB site; (3) a unique source of Pb-Cl (associated with industrial combustion emissions) is found a to be the major (82%) source of Cl in the urban agglomerate; (4) PM2.5 traffic brake dust (Fe-Cu) is mainly primarily emitted and not resuspended, whereas PM2.5 urban crustal dust (Ca) is found mainly resuspended by both traffic vortex and sea breeze; (5) urban dust (Ca) is found the aerosol source most affected by land wetness, reduced by a factor of eight during rainy days and suggesting that wet roads may be a solution for reducing dust concentrations in road sites, far more effective than street sweeping activities.

Normal Modes Expose Active Sites in Enzymes.

PubMed

Glantz-Gashai, Yitav; Meirson, Tomer; Samson, Abraham O

2016-12-01

Accurate prediction of active sites is an important tool in bioinformatics. Here we present an improved structure based technique to expose active sites that is based on large changes of solvent accessibility accompanying normal mode dynamics. The technique which detects EXPOsure of active SITes through normal modEs is named EXPOSITE. The technique is trained using a small 133 enzyme dataset and tested using a large 845 enzyme dataset, both with known active site residues. EXPOSITE is also tested in a benchmark protein ligand dataset (PLD) comprising 48 proteins with and without bound ligands. EXPOSITE is shown to successfully locate the active site in most instances, and is found to be more accurate than other structure-based techniques. Interestingly, in several instances, the active site does not correspond to the largest pocket. EXPOSITE is advantageous due to its high precision and paves the way for structure based prediction of active site in enzymes.

Normal Modes Expose Active Sites in Enzymes

PubMed Central

Glantz-Gashai, Yitav; Samson, Abraham O.

2016-01-01

Accurate prediction of active sites is an important tool in bioinformatics. Here we present an improved structure based technique to expose active sites that is based on large changes of solvent accessibility accompanying normal mode dynamics. The technique which detects EXPOsure of active SITes through normal modEs is named EXPOSITE. The technique is trained using a small 133 enzyme dataset and tested using a large 845 enzyme dataset, both with known active site residues. EXPOSITE is also tested in a benchmark protein ligand dataset (PLD) comprising 48 proteins with and without bound ligands. EXPOSITE is shown to successfully locate the active site in most instances, and is found to be more accurate than other structure-based techniques. Interestingly, in several instances, the active site does not correspond to the largest pocket. EXPOSITE is advantageous due to its high precision and paves the way for structure based prediction of active site in enzymes. PMID:28002427

Factors influencing the atmospheric concentrations of PCBs at an abandoned e-waste recycling site in South China.

PubMed

Wang, Yan; Wu, Xiaowei; Hou, Minmin; Zhao, Hongxia; Chen, Ruize; Luo, Chunling; Zhang, Gan

2017-02-01

The diurnal atmospheric concentrations of polychlorinated biphenyls (PCBs) were investigated at an abandoned e-waste recycling site in South China during winter and summer. Total PCB concentrations during winter and summer were 27.6-212 and 368-1704pg/m 3 in the particulate phase and 270-697 and 3000-15,500pg/m 3 in the gaseous phase, respectively. Both gaseous and particulate PCB concentrations and compositions exhibited significant difference between winter and summer samples, but no diurnal variations during the measurement period. The correlation analysis between PCB concentrations and meteorological conditions, including atmospheric temperature, humidity, and mixing layer height, suggested that the seasonal variability of atmospheric PCB concentrations was strongly temperature-dependent, while the diurnal variability was probably source-dependent. The temperature-driven variations can also be proved by the significant linear correlation between ln P and 1/T in the Clausius-Clapeyron plot. Although government has implemented controls to reduce e-waste pollution, both the relatively high concentrations of PCBs and the diurnal variation in the air suggested that emissions from occasional e-waste recycling activities may still exist in this recycling area. These results underline the importance of continuing e-waste recycling site management long after abandonment. Copyright © 2016 Elsevier B.V. All rights reserved.

Effects of elevated CO2 concentrations on denitrifying and nitrifying popualtions at terrestrial CO2 leakeage analogous sites

NASA Astrophysics Data System (ADS)

Christine, Dictor Marie; Catherine, Joulian; Valerie, Laperche; Stephanie, Coulon; Dominique, Breeze

2010-05-01

CO2 capture and geological storage (CCS) is recognized to be an important option for carbon abatement in Europe. One of the risks of CCS is the leakage from storage site. A laboratory was conducted on soil samples sampled near-surface from a CO2 leakage analogous site (Latera, Italy) in order to evaluate the impact of an elevated soil CO2 concentration on terrestrial bacterial ecosystems form near surface terrestrial environments and to determine a potential bacterial indicator of CO2 leakage from storage site. Surveys were conducted along a 50m long transect across the vent centre, providing a spectrum of CO2 flux rates, soil gas concentrations and compositions (Beaubien et al., 2007). A bacterial diversity studies, performed by CE-SSCP technique, on a soil profile with increasing CO2 soil concentrations (from 0.3% to 100%) showed that a change on bacterial diversity was noted when CO2 concentration was above 50 % of CO2. From this result, 3 soil samples were taken at 70 cm depth in 3 distinct zones (background soil CO2 content, soil CO2 content of 20% and soil CO2 content of 50%). Then theses soil samples were incubated under closed jars flushed with different air atmospheres (20, 50 and 90 % of CO2) during 18 months. At initial, 3, 6, 12 and 18 months, some soil samples were collected in order to estimate the denitrifying, nitrifying activities as a function of CO2 concentration content and times. Theses enzymatic activities were chosen because one occurs under anaerobic conditions (denitrification) and the other occurs under aerobic conditions (nitrification). Both of them were involved in the nitrogen cycle and are major actors of soil function and groundwater quality preservation. Metabolic diversity using BIOLOG Ecoplates was determined on every soil samples. Physico-chemical parameters (e.g. pH, bulk chemistry, mineralogy) were analyzed to have some information about the evolution of the soil during the incubation with increasing soil CO2 concentrations

Atmospheric concentrations of polybrominated diphenyl ethers at near-source sites.

PubMed

Cahill, Thomas M; Groskova, Danka; Charles, M Judith; Sanborn, James R; Denison, Michael S; Baker, Lynton

2007-09-15

Concentrations of polybrominated diphenyl ethers (PBDEs) were determined in air samples from near suspected sources, namely an indoors computer laboratory, indoors and outdoors at an electronics recycling facility, and outdoors at an automotive shredding and metal recycling facility. The results showed that (1) PBDE concentrations in the computer laboratorywere higherwith computers on compared with the computers off, (2) indoor concentrations at an electronics recycling facility were as high as 650,000 pg/m3 for decabromodiphenyl ether (PBDE 209), and (3) PBDE 209 concentrations were up to 1900 pg/m3 at the downwind fenceline at an automotive shredding/metal recycling facility. The inhalation exposure estimates for all the sites were typically below 110 pg/kg/day with the exception of the indoor air samples adjacent to the electronics shredding equipment, which gave exposure estimates upward of 40,000 pg/kg/day. Although there were elevated inhalation exposures at the three source sites, the exposure was not expected to cause adverse health effects based on the lowest reference dose (RfD) currently in the Integrated Risk Information System (IRIS), although these RfD values are currently being re-evaluated by the U.S. Environmental Protection Agency. More research is needed on the potential health effects of PBDEs.

BOREAS TGB-1 CH4 Concentration and Flux Data from NSA Tower Sites

NASA Technical Reports Server (NTRS)

Hall, Forrest G. (Editor); Conrad, Sara K. (Editor); Crill, Patrick; Varner, Ruth K.

2000-01-01

The BOREAS TGB-1 team made numerous measurements of trace gas concentrations and fluxes at various NSA sites. This data set contains half-hourly averages of ambient methane (CH4) measurements and calculated fluxes for the NSA-Fen in 1996 and the NSA-BP and NSA-OJP tower sites in 1994. The purpose of this study was to determine the CH4 flux from the study area by measuring ambient CH 4 concentrations. This flux can then be compared to the chamber flux measurements taken at the same sites. The data are provided in tabular ASCII files.

Seawater Polluted with Highly Concentrated Polycyclic Aromatic Hydrocarbons Suppresses Osteoblastic Activity in the Scales of Goldfish, Carassius auratus.

PubMed

Suzuki, Nobuo; Sato, Masayuki; Nassar, Hossam F; Abdel-Gawad, Fagr Kh; Bassem, Samah M; Yachiguchi, Koji; Tabuchi, Yoshiaki; Endo, Masato; Sekiguchi, Toshio; Urata, Makoto; Hattori, Atsuhiko; Mishima, Hiroyuki; Shimasaki, Youhei; Oshima, Yuji; Hong, Chun-Sang; Makino, Fumiya; Tang, Ning; Toriba, Akira; Hayakawa, Kazuichi

2016-08-01

We have developed an original in vitro bioassay using teleost scale, that has osteoclasts, osteoblasts, and bone matrix as each marker: alkaline phosphatase (ALP) for osteoblasts and tartrate-resistant acid phosphatase (TRAP) for osteoclasts. Using this scale in vitro bioassay, we examined the effects of seawater polluted with highly concentrated polycyclic aromatic hydrocarbons (PAHs) and nitro-polycyclic aromatic hydrocarbons (NPAHs) on osteoblastic and osteoclastic activities in the present study. Polluted seawater was collected from two sites (the Alexandria site on the Mediterranean Sea and the Suez Canal site on the Red Sea). Total levels of PAHs in the seawater from the Alexandria and Suez Canal sites were 1364.59 and 992.56 ng/l, respectively. We were able to detect NPAHs in both seawater samples. Total levels of NPAHs were detected in the seawater of the Alexandria site (12.749 ng/l) and the Suez Canal site (3.914 ng/l). Each sample of polluted seawater was added to culture medium at dilution rates of 50, 100, and 500, and incubated with the goldfish scales for 6 hrs. Thereafter, ALP and TRAP activities were measured. ALP activity was significantly suppressed by both polluted seawater samples diluted at least 500 times, but TRAP activity did not change. In addition, mRNA expressions of osteoblastic markers (ALP, osteocalcin, and the receptor activator of the NF-κB ligand) decreased significantly, as did the ALP enzyme activity. In fact, ALP activity decreased on treatment with PAHs and NPAHs. We conclude that seawater polluted with highly concentrated PAHs and NPAHs influences bone metabolism in teleosts.

Ambient airborne solids concentrations including volcanic ash at Hanford, Washington sampling sites subsequent to the Mount St. Helens eruption

NASA Technical Reports Server (NTRS)

Sehmel, G. A.

1982-01-01

Airborne solids concentrations were measured on a near daily basis at two Hanford, Washington sites after the eruption of Mount St. Helens on May 18, 1980. These sites are about 211 km east of Mount St. Helens. Collected airborne solids included resuspended volcanic ash plus normal ambient solids. Average airborne solids concentrations were greater at the Hanford meteorological station sampling site which is 24 km northwest of the Horn Rapids dam sampling site. These increased concentrations reflect the sampling site proximity to greater ash fallout depths. Both sites are in low ash fallout areas although the Hanford meteorological station site is closer to the greater ash fallout areas. Airborne solids concentrations were decreased by rain, but airborne solids concentrations rapidly increased as surfaces dried. Airborne concentrations tended to become nearly the same at both sampling sites only for July 12 and 13.

Deep Sequencing of Random Mutant Libraries Reveals the Active Site of the Narrow Specificity CphA Metallo-β-Lactamase is Fragile to Mutations.

PubMed

Sun, Zhizeng; Mehta, Shrenik C; Adamski, Carolyn J; Gibbs, Richard A; Palzkill, Timothy

2016-09-12

CphA is a Zn(2+)-dependent metallo-β-lactamase that efficiently hydrolyzes only carbapenem antibiotics. To understand the sequence requirements for CphA function, single codon random mutant libraries were constructed for residues in and near the active site and mutants were selected for E. coli growth on increasing concentrations of imipenem, a carbapenem antibiotic. At high concentrations of imipenem that select for phenotypically wild-type mutants, the active-site residues exhibit stringent sequence requirements in that nearly all residues in positions that contact zinc, the substrate, or the catalytic water do not tolerate amino acid substitutions. In addition, at high imipenem concentrations a number of residues that do not directly contact zinc or substrate are also essential and do not tolerate substitutions. Biochemical analysis confirmed that amino acid substitutions at essential positions decreased the stability or catalytic activity of the CphA enzyme. Therefore, the CphA active - site is fragile to substitutions, suggesting active-site residues are optimized for imipenem hydrolysis. These results also suggest that resistance to inhibitors targeted to the CphA active site would be slow to develop because of the strong sequence constraints on function.

Identification of the fatty acid activation site on human ClC-2.

PubMed

Cuppoletti, John; Tewari, Kirti P; Chakrabarti, Jayati; Malinowska, Danuta H

2017-06-01

Fatty acids (including lubiprostone and cobiprostone) are human ClC-2 (hClC-2) Cl - channel activators. Molecular and cellular mechanisms underlying this activation were examined. Role of a four-amino acid PKA activation site, RGET 691 , of hClC-2 was investigated using wild-type (WT) and mutant (AGET, RGEA, and AGAA) hClC-2 expressed in 293EBNA cells as well as involvement of PKA, intracellular cAMP concentration ([cAMP] i ), EP 2 , or EP 4 receptor agonist activity. All fatty acids [lubiprostone, cobiprostone, eicosatetraynoic acid (ETYA), oleic acid, and elaidic acid] caused significant rightward shifts in concentration-dependent Cl - current activation (increasing EC 50 s) with mutant compared with WT hClC-2 channels, without changing time and voltage dependence, current-voltage rectification, or methadone inhibition of the channel. As with lubiprostone, cobiprostone activation of hClC-2 occurred with PKA inhibitor (myristoylated protein kinase inhibitor) present or when using double PKA activation site (RRAA 655 /RGEA 691 ) mutant. Cobiprostone did not activate human CFTR. Fatty acids did not increase [cAMP] i in hClC-2/293EBNA or T84 cells. Using T84 CFTR knockdown cells, cobiprostone increased hClC-2 Cl - currents without increasing [cAMP] i, while PGE 2 and forskolin-IBMX increased both. Fatty acids were not agonists of EP 2 or EP 4 receptors. L-161,982, a supposed EP 4 -selective inhibitor, had no effect on lubiprostone-activated hClC-2 Cl - currents but significantly decreased T84 cell barrier function measured by transepithelial resistance and fluorescent dextran transepithelial movement. The present findings show that RGET 691 of hClC-2 (possible binding site) plays an important functional role in fatty acid activation of hClC-2. PKA, [cAMP] i , and EP 2 or EP 4 receptors are not involved. These studies provide the molecular basis for fatty acid regulation of hClC-2. Copyright © 2017 the American Physiological Society.

Ammonia concentrations at a site in Southern Scotland from 2 yr of continuous measurements

NASA Astrophysics Data System (ADS)

Burkhardt, J.; Sutton, M. A.; Milford, C.; Storeton-West, R. L.; Fowler, D.

Atmospheric ammonia (NH 3) concentrations were measured using a continuous-flow annular denuder over a period of 2 yr at a rural site near Edinburgh, Scotland. Meteorological parameters as well as sulphur dioxide (SO 2) concentrations were also recorded. The overall arithmetic mean NH 3 concentration was 1.4 μg m -3. Although an annual cycle with largest NH 3 concentrations in summer was apparent for seasonal geometric mean concentrations, arithmetic mean concentrations were largest in the spring and autumn, indicating the increased importance of occasional high concentration events in these seasons. The NH 3 concentrations were influenced by local sources as well as by background concentrations, dependent on wind direction, whereas SO 2 geometric standard deviations indicated more distant sources. The daily cycle of NH 3 and SO 2 concentrations was dependent on wind speed ( u). At u<1 m s -1, NH 3 concentrations were smallest and SO 2 concentrations were largest around noon, whereas at u>1 m s -1 this cycle was less pronounced for both gases and NH 3 concentrations were largest around 1800 hours. These opposite diurnal cycles may be explained by the interaction of boundary layer mixing with local sources for NH 3 and remote sources for SO 2. Comparing the ammonia data with critical levels and critical loads shows that the critical level is not exceeded at this site over any averaging time. In contrast, the N critical load would probably be exceeded for moorland vegetation near this site, showing that the contribution of atmospheric NH 3 to nitrogen deposition in the long term is a more significant issue than exceedance of critical levels.

Aerosol particle and organic vapor concentrations at industrial work sites in Malaysia.

PubMed

Armstrong, R W; Rood, M J; Sani, S; Mohamed, M; Rashid, M; Jab, A T; Landsberger, S

2001-01-01

The objective of this study was to establish baseline data about air pollutants potentially related to nasopharyngeal carcinoma (NPC) in the Federal Territory and Selangor, Malaysia. During 1991-1993, ambient air quality was monitored at 42 work sites representing ten industrial sectors: adhesive manufacturing, foundries, latex processing, metalworking, plywood/veneer milling, ricemilling, rubber tire manufacturing, sawmilling, shoemaking, and textile related industries. At each work site, aerosol particle size distributions and concentrations of formaldehyde, benzene, toluene, isopropyl alcohol, and furfural were measured. Mean aerosol particle concentrations ranged from 61 micrograms/m3 in foundries to 5,578 micrograms/m3 in ricemills, with five industries (adhesives, metalworking, ricemilling, sawmilling, and shoemaking) exceeding the US EPA 24-hr ambient air standard for PM-10. Formaldehyde concentrations exceeded the threshold limit value (TLV) in adhesives factories. Other vapours and elements measured were well below TLVs.

Bacterial Abundance and Activity across Sites within Two Northern Wisconsin Sphagnum Bogs.

PubMed

Fisher; Graham; Graham

1998-11-01

Abstract Bacterial abundance, temperature, pH, and dissolved organic carbon (DOC) concentration were compared across surface sites within and between two northern Wisconsin Sphagnum peatlands over the summer seasons in 1995 and 1996. Sites of interest were the Sphagnum mat surface, the water-filled moat (lagg) at the bog margin, and the bog lake littoral zone. Significant differences in both bacterial populations and water chemistry were observed between sites. pH was highest in the lake and lowest in the mat at both bogs; the opposite was true for DOC. Large populations of bacteria were present in surface interstitial water from the mat; abundance in this site was consistently higher than in the moat or lake. Bacterial abundance also increased across sites of increasing DOC concentration and declining pH. Bacterial activities (rates of [3H]leucine incorporation) and growth in dilution cultures (with grazers removed) were also assessed in lake, moat, and mat sites. Results using these measures generally supported the trends observed in abundance, although high rates of [3H]leucine incorporation were recorded in the moat at one of the bogs. Our results indicate that bacterial populations in Sphagnum peatlands are not adversely affected by acidity, and that DOC may be more important than pH in determining bacterial abundance in these environments.

Transfer hydrogenation over sodium-modified ceria: Enrichment of redox sites active for alcohol dehydrogenation

DOE PAGES

Nelson, Nicholas C.; Boote, Brett W.; Naik, Pranjali; ...

2017-01-17

Ceria (CeO 2) and sodium-modified ceria (Ce-Na) were prepared through combustion synthesis. Palladium was deposited onto the supports (Pd/CeO 2 and Pd/Ce-Na) and their activity for the aqueous-phase transfer hydrogenation of phenol using 2-propanol under liquid flow conditions was studied. Pd/Ce-Na showed a marked increase (6×) in transfer hydrogenation activity over Pd/CeO 2. Material characterization indicated that water-stable sodium species were not doped into the ceria lattice, but rather existed as subsurface carbonates. Modification of ceria by sodium provided more adsorption and redox active sites (i.e. defects) for 2-propanol dehydrogenation. This effect was an intrinsic property of the Ce-Na supportmore » and independent of Pd. The redox sites active for 2-propanol dehydrogenation were thermodynamically equivalent on both supports/catalysts. At high phenol concentrations, the reaction was limited by 2-propanol adsorption. Furthermore, the difference in catalytic activity was attributed to the different numbers of 2-propanol adsorption and redox active sites on each catalyst.« less

Dissecting the active site of a photoreceptor protein

NASA Astrophysics Data System (ADS)

Hoff, Wouter; Hara, Miwa; Ren, Jie; Moghadam, Farzaneh; Xie, Aihua; Kumauchi, Masato

While enzymes are quite large molecules, functionally important chemical events are often limited to a small region of the protein: the active site. The physical and chemical properties of residues at such active sites are often strongly altered compared to the same groups dissolved in water. Understanding such effects is important for unraveling the mechanisms underlying protein function and for protein engineering, but has proven challenging. Here we report on our ongoing efforts on using photoactive yellow protein (PYP), a bacterial photoreceptor, as a model system for such effects. We will report on the following questions: How many residues affect active site properties? Are these residues in direct physical contact with the active site? Can functionally important residues be recognized in the crystal structure of a protein? What structural resolution is needed to understand active sites? What spectroscopic techniques are most informative? Which weak interactions dominate active site properties?

Spatial variability of dissolved phosphorous concentrations and alkaline phosphatase activity in the East China Sea

NASA Astrophysics Data System (ADS)

Liu, H.; Chang, J.; Ho, T.; Gong, G.

2010-12-01

The concentrations of dissolved inorganic phosphorus (DIP) and alkaline phosphatase activity (APA) have been determined at about 25 sampling stations in the East China Sea since 2003. The stations are mainly distributed from the Changjiang river mouth to northern Taiwan and east to the shelf break. In addition to the Changjiang discharge, we have found a specific nutrient source around a coastal site (122° 2’30’’ E, 28° 40’ N). Elevated DIP and nitrate concentrations have been constantly observed around the sampling station for 8 years, where the surface DIP concentrations are generally around 0.3 µM. The nutrient source may either originate from ground water discharge or coastal upwelling, where lower temperature has been observed in the water column around the station. In general, APA has been negatively correlated with DIP concentrations in the studies sites, with lowest APA around the high DIP station and the Changjiang river mouth.

Active site dynamics of ribonuclease.

PubMed Central

Brünger, A T; Brooks, C L; Karplus, M

1985-01-01

The stochastic boundary molecular dynamics method is used to study the structure, dynamics, and energetics of the solvated active site of bovine pancreatic ribonuclease A. Simulations of the native enzyme and of the enzyme complexed with the dinucleotide substrate CpA and the transition-state analog uridine vanadate are compared. Structural features and dynamical couplings for ribonuclease residues found in the simulation are consistent with experimental data. Water molecules, most of which are not observed in crystallographic studies, are shown to play an important role in the active site. Hydrogen bonding of residues with water molecules in the free enzyme is found to mimic the substrate-enzyme interactions of residues involved in binding. Networks of water stabilize the cluster of positively charged active site residues. Correlated fluctuations between the uridine vanadate complex and the distant lysine residues are mediated through water and may indicate a possible role for these residues in stabilizing the transition state. Images PMID:3866234

Benzodiazepines: rat pinealocyte binding sites and augmentation of norepinephrine-stimulated N-acetyltransferase activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Matthew, E.; Parfitt, A.G.; Sugden, D.

1984-02-01

Studies of (/sup 3/H)diazepam binding to intact rat pineal cells were carried out in tissue culture preparations. The binding was saturable, reversible and proportional to the number of cells used. Scatchard analysis resulted in a linear plot (Kd . 23 nM, maximum binding sites (Bmax) . 1.56 pmol/mg of protein for cells in monolayer culture; Kd . 7 nM, Bmax . 1.3 pmol/mg of protein for cells in suspension culture). Inhibition constants (Ki) for clonazepam (500 nM), flunitrazepam (38 nM) and Ro-5-4864 (5 nM) indicated that the binding sites were probably of the ''peripheral'' type. In addition, the effects ofmore » diazepam on norepinephrine-stimulated N-acetyltransferase (NAT) activity were studied in organ culture and dissociated cell culture. Diazepam (10-50 microM) both prolonged and increased the magnitude of the norepinephrine-induced increase in NAT activity but did not affect the initial rate of rise of enzyme activity. The effect was dose-dependent and was also seen with clonazepam, flunitrazepam and Ro-5-4864, but not with Ro-15-1788. Diazepam, by itself, at these concentrations, had no effect on NAT, but enzyme activity was increased by higher concentrations (0.1-1 mM). Although a relationship between the (/sup 3/H)diazepam binding sites described here and the effect of benzodiazepines on NAT cannot be established from these studies, the data suggest that the benzodiazepines may alter melatonin levels through their action on NAT.« less

Low dielectric response in enzyme active site

PubMed Central

Mertz, Edward L.; Krishtalik, Lev I.

2000-01-01

The kinetics of charge transfer depend crucially on the dielectric reorganization of the medium. In enzymatic reactions that involve charge transfer, atomic dielectric response of the active site and of its surroundings determines the efficiency of the protein as a catalyst. We report direct spectroscopic measurements of the reorganization energy associated with the dielectric response in the active site of α-chymotrypsin. A chromophoric inhibitor of the enzyme is used as a spectroscopic probe. We find that water strongly affects the dielectric reorganization in the active site of the enzyme in solution. The reorganization energy of the protein matrix in the vicinity of the active site is similar to that of low-polarity solvents. Surprisingly, water exhibits an anomalously high dielectric response that cannot be described in terms of the dielectric continuum theory. As a result, sequestering the active site from the aqueous environment inside low-dielectric enzyme body dramatically reduces the dielectric reorganization. This reduction is particularly important for controlling the rate of enzymatic reactions. PMID:10681440

ATP binding at noncatalytic sites of soluble chloroplast F1-ATPase is required for expression of the enzyme activity.

PubMed

Milgrom, Y M; Ehler, L L; Boyer, P D

1990-11-05

The F1-ATPase from chloroplasts (CF1) lacks catalytic capacity for ATP hydrolysis if ATP is not bound at noncatalytic sites. CF1 heat activated in the presence of ADP, with less than one ADP and no ATP at non-catalytic sites, shows a pronounced lag in the onset of ATP hydrolysis after exposure to 5-20 microM ATP. The onset of activity correlates well with the binding of ATP at the last two of the three noncatalytic sites. The dependence of activity on the presence of ATP at non-catalytic sites is shown at relatively low or high free Mg2+ concentrations, with or without bicarbonate as an activating anion, and when the binding of ATP at noncatalytic sites is slowed 3-4-fold by sulfate. The latent CF1 activated by dithiothreitol also requires ATP at noncatalytic sites for ATPase activity. A similar requirement by other F1-ATPases and by ATP synthases seems plausible.

Cyanide does more to inhibit heme enzymes, than merely serving as an active-site ligand

DOE Office of Scientific and Technical Information (OSTI.GOV)

Parashar, Abhinav; Venkatachalam, Avanthika; Gideon, Daniel Andrew

Highlights: • Cyanide (CN) is a well-studied toxic principle, known to inhibit heme-enzymes. • Inhibition is supposed to result from CN binding at the active site as a ligand. • Diverse heme enzymes’ CN inhibition profiles challenge prevailing mechanism. • Poor binding efficiency of CN at low enzyme concentrations and ligand pressures. • CN-based diffusible radicals cause ‘non-productive electron transfers’ (inhibition). - Abstract: The toxicity of cyanide is hitherto attributed to its ability to bind to heme proteins’ active site and thereby inhibit their activity. It is shown herein that the long-held interpretation is inadequate to explain several observations inmore » heme-enzyme reaction systems. Generation of cyanide-based diffusible radicals in heme-enzyme reaction milieu could shunt electron transfers (by non-active site processes), and thus be detrimental to the efficiency of oxidative outcomes.« less

Short-term 222Rn activity concentration changes in underground spaces with limited air exchange with the atmosphere

NASA Astrophysics Data System (ADS)

Fijałkowska-Lichwa, L.; Przylibski, T. A.

2011-04-01

The authors investigated short-time changes in 222Rn activity concentration occurring yearly in two underground tourist facilities with limited air exchange with the atmosphere. One of them is Niedźwiedzia (Bear) Cave in Kletno, Poland - a natural space equipped with locks ensuring isolation from the atmosphere. The other site is Fluorite Adit in Kletno, a section of a disused uranium mine. This adit is equipped with a mechanical ventilation system, operated periodically outside the opening times (at night). Both sites are situated within the same metamorphic rock complex, at similar altitudes, about 2 km apart. The measurements conducted revealed spring and autumn occurrence of convective air movements. In Bear Cave, this process causes a reduction in 222Rn activity concentration in the daytime, i.e. when tourists, guides and other staff are present in the cave. From the point of view of radiation protection, this is the best situation. For the rest of the year, daily concentrations of 222Rn activity in the cave are very stable. In Fluorite Adit, on the other hand, significant variations in daily 222Rn activity concentrations are recorded almost all year round. These changes are determined by the periods of activity and inactivity of mechanical ventilation. Unfortunately this is inactive in the daytime, which results in the highest values of 222Rn activity concentration at the times when tourists and staff are present in the adit. Slightly lower concentrations of radon in Fluorite Adit are recorded in the winter season, when convective air movements carry a substantial amount of radon out into the atmosphere. The incorrect usage of mechanical ventilation in Fluorite Adit results in the most unfavourable conditions in terms of radiation protection. The staff working in that facility are exposed practically throughout the year to the highest 222Rn activity concentrations, both at work (in the adit) and at home (outside their working hours). Therefore, not very well

Habitat influence on antioxidant activity and tannin concentrations of Spondias tuberosa.

PubMed

de Sousa Araújo, Thiago Antônio; de Almeida e Castro, Valerium Thijan Nobre; de Amorim, Elba Lúcia Cavalcanti; de Albuquerque, Ulysses Paulino

2012-06-01

Different habitat conditions can be responsible for the production of secondary metabolites and for the antioxidant properties of plant products. Thus, the aim of this study was to evaluate whether the antioxidant activity and tannin concentrations in the stem bark of Spondias tuberosa Arruda (Anacardiaceae) varied with collection site. The bark was collected from 25 individual trees, distributed in five different landscape units, as follows: agroforestry gardens, areas of pastures, maize cultivation areas, mountain areas and mountain bases, with the former 3 being considered as anthropogenic habitats, and the latter 2 considered as habitats with native coverage. The study was conducted in the rural area of the city of Altinho, Pernambuco State (Northeast Brazil). The DPPH (1,1-diphenyl-2-picrylhydrazyl) method was used to measure the antioxidant activity and tannin concentrations were evaluated by using the radial diffusion method. The results demonstrated that there were no significant differences among the tannin concentrations of the individuals from the native (6.27% ± 1.75) or anthropogenic areas (4.63% ± 2.55), (H = 2.24; p > 0.05). In contrast, there were significant differences (H = 5.1723; p < 0.05) among the CE₅₀ means of the antioxidant activities of the individuals from the native (32.10 µg/ml ± 5.27) and anthropogenic areas (27.07 µg/ml ± 2.29). However, correlations between the tannin concentrations and antioxidant activity of the extracts were not observed in the native (r = 0.39; p > 0.05) or in the anthropogenic areas (r = 0.38; p > 0.05). Because the variation of the antioxidant capacity of S. tuberosa bark was not accompanied by a variation in the tannin concentration, this property may be related to the presence of other metabolite(s).

Methanopyrus kandleri topoisomerase V contains three distinct AP lyase active sites in addition to the topoisomerase active site

PubMed Central

Rajan, Rakhi; Osterman, Amy; Mondragón, Alfonso

2016-01-01

Topoisomerase V (Topo-V) is the only topoisomerase with both topoisomerase and DNA repair activities. The topoisomerase activity is conferred by a small alpha-helical domain, whereas the AP lyase activity is found in a region formed by 12 tandem helix-hairpin-helix ((HhH)2) domains. Although it was known that Topo-V has multiple repair sites, only one had been mapped. Here, we show that Topo-V has three AP lyase sites. The atomic structure and Small Angle X-ray Scattering studies of a 97 kDa fragment spanning the topoisomerase and 10 (HhH)2 domains reveal that the (HhH)2 domains extend away from the topoisomerase domain. A combination of biochemical and structural observations allow the mapping of the second repair site to the junction of the 9th and 10th (HhH)2 domains. The second site is structurally similar to the first one and to the sites found in other AP lyases. The 3rd AP lyase site is located in the 12th (HhH)2 domain. The results show that Topo-V is an unusual protein: it is the only known protein with more than one (HhH)2 domain, the only known topoisomerase with dual activities and is also unique by having three AP lyase repair sites in the same polypeptide. PMID:26908655

Aryl hydrocarbon receptor-mediated activity of atmospheric particulate matter from an urban and a rural site in Switzerland

NASA Astrophysics Data System (ADS)

Wenger, Daniela; Gerecke, Andreas C.; Heeb, Norbert V.; Hueglin, Christoph; Seiler, Cornelia; Haag, Regula; Naegeli, Hanspeter; Zenobi, Renato

Atmospheric particulate matter (PM) is an air-suspended mixture of solid and liquid particles that vary in size, shape, and chemical composition. Long-term exposure to elevated concentrations of fine atmospheric particles is considered to pose a health threat to humans and animals. In this context, it has been hypothesized that toxic chemicals such as polycyclic aromatic hydrocarbons (PAHs) play an important role. Some PAHs are known to be carcinogenic and it has been shown that carcinogenic effects of PAHs are mediated by the aryl hydrocarbon receptor (AhR). In this study, PM1 was collected at a rural and an urban traffic site during an intense winter smog period, in which concentration of PM1 often exceeded 50 μg m -3. We applied an in vitro reporter gene assay (DR-CALUX) to detect and quantify PM1-associated chemicals that induce AhR-mediated gene expression. This activity was expressed as CALUX equivalents of 2,3,7,8-tetrachlorodibenzodioxin (PM-TCDD-CEQs). In addition, concentrations of PAHs in the PM1 extracts were determined using gas chromatography/high-resolution mass spectrometry. Concentrations of PM-TCDD-CEQs ranged from 10 to 85 pg m -3 and from 19 to 87 pg m -3 at the urban and rural site, respectively. By the use of known relative potency factors, the measured concentration of a PAH was converted into a PAH-TCDD-CEQ concentration. ΣPAH-TCDD-CEQ and PM-TCDD-CEQ were highly correlated at both sites ( r2 = 0.90 and 0.69). The calculated ΣPAH-TCDD-CEQs explain between 2% and 20% of the measured PM-TCDD-CEQs. Benzo[ k]fluoranthene was the most important PAH causing approximately 60% of the total ΣPAH-TCDD-CEQ activity. In contrast to NO, CO, PM10, and PM1, the concentration of PM-TCDD-CEQs showed no significant difference between the two sites. No indications were found that road traffic emissions caused elevated concentrations of PM-TCDD-CEQs at the urban traffic site.

Active site structure and catalytic mechanism of phosphodiesterase for degradation of intracellular second messengers

NASA Astrophysics Data System (ADS)

Zhan, Chang-Guo

2002-03-01

Phosphodiesterases are clinical targets for a variety of biological disorders, because this superfamily of enzymes regulate intracellular concentration of cyclic nucleotides that serve as the second messengers playing a critical role in a variety of physiological processes. Understanding structure and mechanism of a phosphodiesterase will provide a solid basis for rational design of the more efficient therapeutics. Although a three-dimensional X-ray crystal structure of the catalytic domain of human phosphodiesterase 4B2B was recently reported, it was uncertain whether a critical bridging ligand in the active site is a water molecule or a hydroxide ion. The identity of this bridging ligand has been determined by performing first-principles quantum chemical calculations on models of the active site. All the results obtained indicate that this critical bridging ligand in the active site of the reported X-ray crystal structure is a hydroxide ion, rather than a water molecule, expected to serve as the nucleophile to initialize the catalytic degradation of the intracellular second messengers.

Per- and polyfluoroalkyl substances in firefighting foam concentrates and water samples collected near sites impacted by the use of these foams.

PubMed

Dauchy, Xavier; Boiteux, Virginie; Bach, Cristina; Rosin, Christophe; Munoz, Jean-François

2017-09-01

To extinguish large-scale fuel fires, fluorosurfactant based foams (FSBFs) were developed in the 1960s and have been used ever since. In this study, 154 per- and polyfluoroalkyl substances (PFASs) including 122 emerging PFASs used as surfactants in FSBFs were sought in nine different foam concentrates. Field investigations were also carried out in the vicinity of four sites where FSBFs are or were intensively used (two airports, a training center for firefighters and an oil storage depot after a large explosion). In the foam concentrates, only three PFASs were quantified with concentrations ranging from 22,500 to 3,188,000 μg/L. Thirteen emerging PFASs were also identified in these samples based on their mass transitions and intensities. Overall, each foam was a mixture of at least two classes of PFASs. In three concentrates, none of the 122 emerging PFASs were identified as the main ingredient. A perfluoroalkyl acid precursor oxidation assay was therefore performed, and revealed the presence of high amounts of unidentified PFASs. In the vicinity of the four investigated sites, several PFASs were systematically quantified in all of the samples collected downstream of the sites. PFAS profiles were heavily influenced by parameters such as route of PFAS transport after use (runoff, seepage, direct discharge), time elapsed since the cessation of firefighting activities, and firefighting foam composition. The PFAS concentrations found around the investigated sites are the highest recorded in France and resulted in the closure of certain drinking water resources. Copyright © 2017 Elsevier Ltd. All rights reserved.

Greenhouse Gas Fluxes at the Tablelands, NL, Canada: A Site of Active Serpentinization

NASA Astrophysics Data System (ADS)

Morrill, P. L.; Morrissey, L. S.; Cumming, E.

2016-12-01

Active sites of serpentinization have been proposed as sites for carbon capture and storage (CCS) projects. However, in addition to their ability to convert carbon dioxide to carbonate rock, sites of serpentinization also have the potential release methane, which is a more power greenhouse gas than carbon dioxide. Very little is known about the natural flux of carbon dioxide sequestered and methane released into the atmosphere from active sites of serpentinization. In this study we measured carbon dioxide, methane, and nitrous oxide gas fluxes at a pool of ultra-basic water discharging from serpentinized rock in Winterhouse Canyon, Gros Morne, Newfoundland. We found that the flux of methane released was 4.6 x 10-7 mol/m2/min and the carbon dioxide sequestered was 1.9 x 10-5 mol/m2/min, while the concentrations of nitrous oxide showed little change. Based on these fluxes we calculated predictive climate change parameters such as net radiative forcing and global warming potential which predicted that despite the methane being released the site still had an overall long-term atmospheric cooling effect based on the natural rate of carbon dioxide sequestration.

Volatile aromatic hydrocarbons and dicarboxylic acid concentrations in air at an urban site in the Southwestern US

NASA Astrophysics Data System (ADS)

Tran, Ngoc K.; Steinberg, Spencer M.; Johnson, Brian J.

Concentrations of benzene, toluene, ethylbenzene, o-xylene, and m- and p-xylene were measured at an urban sampling site in Las Vegas, NV by sorbent sampling followed by thermal desorption and determination by GC-PID. Simultaneously, measurements of oxalic, malonic, succinic, and adipic acids were made at the same site by collection on quartz filters, extraction, esterification, and determination by GC-FID. For the period from April 7, 1997 to June 11, 1997, 201 sets of hydrocarbon measurements and 99 sets of acid measurements were made. Additional measurements of dicarboxylic acids were made on samples that represented potential direct sources, e.g. green plants and road dust. Correlations between the hydrocarbon and CO concentrations (measured by the Clark County Health District at a nearby site) were highly significant and a strong negative correlation of hydrocarbon concentration with ozone concentration (also from the county site) was observed under quiescent atmospheric conditions. In general, dicarboxylic acid concentrations were well correlated with one another (with the exception of adipic acid) but not well correlated with hydrocarbon, CO, and ozone concentrations. Multiple sources and complex formation processes are indicated for the dicarboxylic acids.

Activation of phenylalanine hydroxylase by phenylalanine does not require binding in the active site.

PubMed

Roberts, Kenneth M; Khan, Crystal A; Hinck, Cynthia S; Fitzpatrick, Paul F

2014-12-16

Phenylalanine hydroxylase (PheH), a liver enzyme that catalyzes the hydroxylation of excess phenylalanine in the diet to tyrosine, is activated by phenylalanine. The lack of activity at low levels of phenylalanine has been attributed to the N-terminus of the protein's regulatory domain acting as an inhibitory peptide by blocking substrate access to the active site. The location of the site at which phenylalanine binds to activate the enzyme is unknown, and both the active site in the catalytic domain and a separate site in the N-terminal regulatory domain have been proposed. Binding of catecholamines to the active-site iron was used to probe the accessibility of the active site. Removal of the regulatory domain increases the rate constants for association of several catecholamines with the wild-type enzyme by ∼2-fold. Binding of phenylalanine in the active site is effectively abolished by mutating the active-site residue Arg270 to lysine. The k(cat)/K(phe) value is down 10⁴ for the mutant enzyme, and the K(m) value for phenylalanine for the mutant enzyme is >0.5 M. Incubation of the R270K enzyme with phenylalanine also results in a 2-fold increase in the rate constants for catecholamine binding. The change in the tryptophan fluorescence emission spectrum seen in the wild-type enzyme upon activation by phenylalanine is also seen with the R270K mutant enzyme in the presence of phenylalanine. Both results establish that activation of PheH by phenylalanine does not require binding of the amino acid in the active site. This is consistent with a separate allosteric site, likely in the regulatory domain.

Activation of Phenylalanine Hydroxylase by Phenylalanine Does Not Require Binding in the Active Site

PubMed Central

2015-01-01

Phenylalanine hydroxylase (PheH), a liver enzyme that catalyzes the hydroxylation of excess phenylalanine in the diet to tyrosine, is activated by phenylalanine. The lack of activity at low levels of phenylalanine has been attributed to the N-terminus of the protein’s regulatory domain acting as an inhibitory peptide by blocking substrate access to the active site. The location of the site at which phenylalanine binds to activate the enzyme is unknown, and both the active site in the catalytic domain and a separate site in the N-terminal regulatory domain have been proposed. Binding of catecholamines to the active-site iron was used to probe the accessibility of the active site. Removal of the regulatory domain increases the rate constants for association of several catecholamines with the wild-type enzyme by ∼2-fold. Binding of phenylalanine in the active site is effectively abolished by mutating the active-site residue Arg270 to lysine. The kcat/Kphe value is down 104 for the mutant enzyme, and the Km value for phenylalanine for the mutant enzyme is >0.5 M. Incubation of the R270K enzyme with phenylalanine also results in a 2-fold increase in the rate constants for catecholamine binding. The change in the tryptophan fluorescence emission spectrum seen in the wild-type enzyme upon activation by phenylalanine is also seen with the R270K mutant enzyme in the presence of phenylalanine. Both results establish that activation of PheH by phenylalanine does not require binding of the amino acid in the active site. This is consistent with a separate allosteric site, likely in the regulatory domain. PMID:25453233

Role of volatilization in changing TBA and MTBE concentrations at MTBE-contaminated sites.

PubMed

Eweis, Juana B; Labolle, Eric M; Benson, David A; Fogg, Graham E

2007-10-01

Tertiary butyl alcohol (TBA) is commonly found as an impurity in methyl tertiary butyl ether (MTBE) added to gasoline. Frequent observations of high TBA, and especially rising TBA/MTBE concentration ratios, in groundwater at gasoline spill sites are generally attributed to microbial conversion of MTBE to TBA. Typically overlooked is the role of volatilization in the attenuation of these chemicals especially in the vadose zone, which is a source of contamination to groundwater. Here we show that volatilization, particularly through remediation by vapor extraction, can substantially affect the trends in TBA and MTBE concentrations and the respective mass available to impact groundwater aquifers, through the preferential removal of more volatile compounds, including MTBE, and the apparent enrichment of less volatile compounds like TBA. We demonstrate this phenomenon through numerical simulations of remedial-enhanced volatilization. Results show increases in TBA/MTBE concentration ratios consistent with ratios observed in groundwater at gasoline spill sites. Volatilization is an important, and potentially dominant, process that can result in concentration trends similar to those typically attributed to biodegradation.

Substituting Tyr138 in the active site loop of human phenylalanine hydroxylase affects catalysis and substrate activation.

PubMed

Leandro, João; Stokka, Anne J; Teigen, Knut; Andersen, Ole A; Flatmark, Torgeir

2017-07-01

Mammalian phenylalanine hydroxylase (PAH) is a key enzyme in l-phenylalanine (l-Phe) metabolism and is active as a homotetramer. Biochemical and biophysical work has demonstrated that it cycles between two states with a variably low and a high activity, and that the substrate l-Phe is the key player in this transition. X-ray structures of the catalytic domain have shown mobility of a partially intrinsically disordered Tyr 138 -loop to the active site in the presence of l-Phe. The mechanism by which the loop dynamics are coupled to substrate binding at the active site in tetrameric PAH is not fully understood. We have here conducted functional studies of four Tyr 138 point mutants. A high linear correlation ( r 2 = 0.99) was observed between their effects on the catalytic efficiency of the catalytic domain dimers and the corresponding effect on the catalytic efficiency of substrate-activated full-length tetramers. In the tetramers, a correlation ( r 2 = 0.96) was also observed between the increase in catalytic efficiency (activation) and the global conformational change (surface plasmon resonance signal response) at the same l-Phe concentration. The new data support a similar functional importance of the Tyr 138 -loop in the catalytic domain and the full-length enzyme homotetramer.

Non-competitive inhibition by active site binders.

PubMed

Blat, Yuval

2010-06-01

Classical enzymology has been used for generations to understand the interactions of inhibitors with their enzyme targets. Enzymology tools enabled prediction of the biological impact of inhibitors as well as the development of novel, more potent, ones. Experiments designed to examine the competition between the tested inhibitor and the enzyme substrate(s) are the tool of choice to identify inhibitors that bind in the active site. Competition between an inhibitor and a substrate is considered a strong evidence for binding of the inhibitor in the active site, while the lack of competition suggests binding to an alternative site. Nevertheless, exceptions to this notion do exist. Active site-binding inhibitors can display non-competitive inhibition patterns. This unusual behavior has been observed with enzymes utilizing an exosite for substrate binding, isomechanism enzymes, enzymes with multiple substrates and/or products and two-step binding inhibitors. In many of these cases, the mechanisms underlying the lack of competition between the substrate and the inhibitor are well understood. Tools like alternative substrates, testing the enzyme reaction in the reverse direction and monitoring inhibition time dependence can be applied to enable distinction between 'badly behaving' active site binders and true exosite inhibitors.

Safety Oversight of Decommissioning Activities at DOE Nuclear Sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zull, Lawrence M.; Yeniscavich, William

2008-01-15

The Defense Nuclear Facilities Safety Board (Board) is an independent federal agency established by Congress in 1988 to provide nuclear safety oversight of activities at U.S. Department of Energy (DOE) defense nuclear facilities. The activities under the Board's jurisdiction include the design, construction, startup, operation, and decommissioning of defense nuclear facilities at DOE sites. This paper reviews the Board's safety oversight of decommissioning activities at DOE sites, identifies the safety problems observed, and discusses Board initiatives to improve the safety of decommissioning activities at DOE sites. The decommissioning of former defense nuclear facilities has reduced the risk of radioactive materialmore » contamination and exposure to the public and site workers. In general, efforts to perform decommissioning work at DOE defense nuclear sites have been successful, and contractors performing decommissioning work have a good safety record. Decommissioning activities have recently been completed at sites identified for closure, including the Rocky Flats Environmental Technology Site, the Fernald Closure Project, and the Miamisburg Closure Project (the Mound site). The Rocky Flats and Fernald sites, which produced plutonium parts and uranium materials for defense needs (respectively), have been turned into wildlife refuges. The Mound site, which performed R and D activities on nuclear materials, has been converted into an industrial and technology park called the Mound Advanced Technology Center. The DOE Office of Legacy Management is responsible for the long term stewardship of these former EM sites. The Board has reviewed many decommissioning activities, and noted that there are valuable lessons learned that can benefit both DOE and the contractor. As part of its ongoing safety oversight responsibilities, the Board and its staff will continue to review the safety of DOE and contractor decommissioning activities at DOE defense nuclear sites.« less

Forskolin- and dihydroalprenolol (DHA) binding sites and adenylate cyclase activity in heart of rats fed diets containing different oils

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alam, S.Q.; Ren, Y.F.; Alam, B.S.

1987-05-01

The purpose of the present investigation was to determine if dietary lipids can induce changes in the adenylate cyclase system in rat heart. Three groups of male young Sprague-Dawley rats were fed for 6 weeks diets containing 10% corn oil (I), 8% coconut oil + 2% corn oil (II) or 10% menhaden oil (III). Adenylate cyclase activity (basal, fluoride-, isoproterenol-, and forskolin-stimulated) was higher in heart homogenates of rats in group III than in the other two groups. Concentration of the (/sup 3/H)-forskolin binding sites in the cardiac membranes were significantly higher in rats fed menhaden oil. The values (pmol/mgmore » protein) were 4.8 +/- 0.2 (I), 4.5 +/- 0.7 (II) and 8.4 +/- 0.5 (III). There was no significant difference in the affinity of the forskolin binding sites among the 3 dietary groups. When measured at different concentrations of forskolin, the adenylate cyclase activity in cardiac membranes of rats fed menhaden oil was higher than in the other 2 groups. Concentrations of the (/sup 3/H)DHA binding sites were slightly higher but their affinity was lower in cardiac membranes of rats fed menhaden oil. The results suggest that diets containing fish oil increase the concentration of the forskolin binding sites and may also affect the characteristics of the ..beta..-adrenergic receptor in rat heart.« less

Evaluation of Diazinon and Chlorpyrifos Concentrations and Loads, and other Pesticide Concentrations, at Selected Sites in the San Joaquin Valley, California, April to August, 2001

USGS Publications Warehouse

Domagalski, Joseph L.; Munday, Cathy

2003-01-01

Twelve sites in the San Joaquin Valley of California were monitored weekly during the growing and irrigation season of 2001 for a total of 51 pesticides and pesticide degradation products, with primary interest on the concentration, load, and basin yield of organophosphorus insecticides, especially diazinon and chlorpyrifos. Diazinon was detected frequently, up to 100 percent of the time, at many of the sampling sites, but with generally low concentrations. For all sites, 75 percent of all measured diazinon concentrations were less than 0.02 mg/L, and 90 percent of all measured diazinon concentrations were less than 0.06 mg/L. The highest diazinon concentrations were measured in samples from two west-side tributaries to the San Joaquin River, Orestimba Creek, and Del Puerto Creek. The median concentration of chlorpyrifos was at or less than the laboratory reporting limit (0.005 mg/L) for most sites with the exceptions of two tributaries to the San Joaquin River: Orestimba Creek and the Tuolumne River. For all sites, 75 percent of all measured chlorpyrifos concentrations were less than 0.03 mg/L and 90 percent of all measured chlorpyrifos concentrations were less than 0.07 mg/L. The total load of diazinon out of the basin was just over 7 kilograms, which accounted for about 0.17 percent of the total agricultural applications. The diazinon load from the monitored upstream tributaries accounted for about 50 percent of the load at the mouth of the San Joaquin River. The streamflow from the selected monitored tributaries accounted for about 83 percent of the streamflow at the mouth of the San Joaquin River. The total load of chlorpyrifos out of the basin was 3.75 kilograms, and this accounted for approximately 0.007 percent of the total amount applied. Other pesticides that were frequently detected during this study included herbicides such as metolachlor, simazine, and trifluralin, and insecticides such as carbaryl, carbofuran, and propargite. At Orestimba Creek, DDE

High Concentrations of Organic Contaminants in Air from Ship Breaking Activities in Chittagong, Bangladesh.

PubMed

Nøst, Therese H; Halse, Anne K; Randall, Scott; Borgen, Anders R; Schlabach, Martin; Paul, Alak; Rahman, Atiqur; Breivik, Knut

2015-10-06

The beaches on the coast of Chittagong in Bangladesh are one of the most intense ship breaking areas in the world. The aim of the study was to measure the concentrations of organic contaminants in the air in the city of Chittagong, including the surrounding ship breaking areas using passive air samplers (N = 25). The compounds detected in the highest amounts were the polycyclic aromatic hydrocarbons (PAHs) and short-chain chlorinated paraffins (SCCPs), whereas dichlorodiphenyltrichloroethanes (DDTs), hexachlorobenzene (HCB), and polychlorinated biphenyls (PCBs) were several orders of magnitude lower in comparison. PCBs, PAHs, and HCB were highest at sites near the ship breaking activities, whereas DDTs and SCCPs were higher in the urban areas. Ship breaking activities likely act as atmospheric emission sources of PCBs, PAHs, and HCB, thus adding to the international emphasis on responsible recycling of ships. Concentrations of PAHs, PCBs, DDTs, HCB, and SCCPs in ambient air in Chittagong are high in comparison to those found in similar studies performed in other parts of Asia. Estimated toxic equivalent quotients indicate elevated human health risks caused by inhalation of PAHs at most sites.

Management practices that concentrate visitor activities: Camping impact management at Isle Royale National Park, USA

USGS Publications Warehouse

Marion, J.L.; Farrell, T.A.

2002-01-01

This study assessed campsite conditions and the effectiveness of campsite impact management strategies at Isle Royale National Park, USA. Protocols for assessing indicators of vegetation and soil conditions were developed and applied to 156 campsites and 88 shelters within 36 backcountry campgrounds. The average site was 68 m2 and 83% of sites lost vegetation over areas less than 47 m2. Results reveal that management actions to spatially concentrate camping activities and reduce camping disturbance have been highly successful. Comparisons of disturbed area/overnight stay among other protected areas reinforces this assertion. These reductions in area of camping disturbance are attributed to a designated site camping policy, limitation on site numbers, construction of sites in sloping terrain, use of facilities, and an ongoing program of campsite maintenance. Such actions are most appropriate in higher use backcountry and wilderness settings.

Discriminative structural approaches for enzyme active-site prediction.

PubMed

Kato, Tsuyoshi; Nagano, Nozomi

2011-02-15

Predicting enzyme active-sites in proteins is an important issue not only for protein sciences but also for a variety of practical applications such as drug design. Because enzyme reaction mechanisms are based on the local structures of enzyme active-sites, various template-based methods that compare local structures in proteins have been developed to date. In comparing such local sites, a simple measurement, RMSD, has been used so far. This paper introduces new machine learning algorithms that refine the similarity/deviation for comparison of local structures. The similarity/deviation is applied to two types of applications, single template analysis and multiple template analysis. In the single template analysis, a single template is used as a query to search proteins for active sites, whereas a protein structure is examined as a query to discover the possible active-sites using a set of templates in the multiple template analysis. This paper experimentally illustrates that the machine learning algorithms effectively improve the similarity/deviation measurements for both the analyses.

Ozone concentrations at a selected high-elevation forest site downwind Mexico City

NASA Astrophysics Data System (ADS)

Torres-JArdon, R.

2013-05-01

Torres-Jardón, R.*, Rosas-Pérez, I., Granada-Macías, L. M., Ruiz-Suárez, L. G. Centro de Ciencias de la Atmósfera, UNAM, México D. F. México * rtorres@unam.mx For many years, the vegetation of forest species such as Abies religiosa in natural parks located in the southwest mountains of Mexico City has attracted much attention since these parks have been experiencing a severe decline of unclear etiology. The high ozone levels in the area and the observed naked eye macroscopic, histological and cytological injuries on these species, strongly suggest an important contribution of tropospheric ozone to this deterioration process. Apart of historical short monitoring campaigns for measuring ozone levels in these mountains, it is known just a little is known about the present exposure levels at which the local vegetation is exposed. A continuous ozone analyzer has been in operation since 2011 at a high-elevation forest site (Parque Nacional Miguel Hidalgo, PNMH; 3110 m above mean sea level) located downwind of Mexico City Metropolitan Area (MCMA), in order to characterize the local ozone diel amplitude and its seasonal trend, as well as the influence of MCMA on the local O3 concentrations. Hourly average ozone data in PNMH shows that in general, the diel of ozone concentrations in the forest site has a statistical significant correlation with the pattern of ozone levels observed in several monitoring sites (smog receptor sites) within the MCMA, although the high elevation O3 levels are relatively lower than those in the urban area (around 2200 m above mean sea level). It is possible that a part of the oxidants in the air masses are removed by sink deposition processes during the air mass transport across the hills. The diel amplitude of ozone concentrations is small in the cold season, increasing as the seasons advance to June. As in the city, the highest ozone concentrations occur in April or May and the lowest levels during the rainy season, which extends from

Atmospheric concentrations and temporal variations of C 1C 3 carbonyl compounds at two rural sites in central Ontario

NASA Astrophysics Data System (ADS)

Shepson, P. B.; Hastie, D. R.; Schiff, H. I.; Polizzi, M.; Bottenheim, J. W.; Anlauf, K.; Mackay, G. I.; Karecki, D. R.

Measurements of formaldehyde, acetaldehyde, acetone and propionaldehyde concentrations were made at two rural sites in central Ontario. One site (at Egbert, Ont.) is located ≈60 km northwest of Toronto, while the other site (at Dorset, Ont.) is ≈150 km northeast of the Egbert site. Measurements were made using a modified version of a derivatization technique in which sample air is pumped through Teflon tubes packed with silica gel that is coated with 2,4-dinitrophenylhydrazine (DNPH). The product hydrazones were separated and quantified using HPLC. Quantitative determinations of formaldehyde, acetaldehyde and acetone were made for 49 and 47 samples at the Dorset and Egbert sites, respectively, between 25 July and 30 August 1988. The average concentrations determined at the Dorset site for formaldehyde, acetaldehyde, and acetone were 1.6, 0.46 and 1.8 ppb, respectively, and for the Egbert site the corresponding averages were 1.8, 0.57 and 1.6 ppb. A set of 10 samples from the Egbert site were analysed for propionaldehyde yielding an average concentration of 0.03 ppb. The formaldehyde measurements were compared with measurements made at the same time using Tunable Diode Laser Absorption Spectroscopy. The observed concentrations reported here are compared with previously reported measurements of these species and interpreted in terms of atmospheric variables (e.g. meteorology, concentrations of precursor hydrocarbons) influencing their concentrations.

SITE SPECIFIC REFERENCE PERSON PARAMETERS AND DERIVED CONCENTRATION STANDARDS FOR THE SAVANNAH RIVER SITE

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jannik, T.

2013-03-14

The purpose of this report is twofold. The first is to develop a set of behavioral parameters for a reference person specific for the Savannah River Site (SRS) such that the parameters can be used to determine dose to members of the public in compliance with Department of Energy (DOE) Order 458.1 “Radiation Protection of the Public and the Environment.” A reference person is a hypothetical, gender and age aggregation of human physical and physiological characteristics arrived at by international consensus for the purpose of standardizing radiation dose calculations. DOE O 458.1 states that compliance with the annual dose limitmore » of 100 mrem (1 mSv) to a member of the public may be demonstrated by calculating the dose to the maximally exposed individual (MEI) or to a representative person. Historically, for dose compliance, SRS has used the MEI concept, which uses adult dose coefficients and adult male usage parameters. Beginning with the 2012 annual site environmental report, SRS will be using the representative person concept for dose compliance. The dose to a representative person will be based on 1) the SRS-specific reference person usage parameters at the 95th percentile of appropriate national or regional data, which are documented in this report, 2) the reference person (gender and age averaged) ingestion and inhalation dose coefficients provided in DOE Derived Concentration Technical Standard (DOE-STD-1196-2011), and 3) the external dose coefficients provided in the DC_PAK3 toolbox. The second purpose of this report is to develop SRS-specific derived concentration standards (DCSs) for all applicable food ingestion pathways, ground shine, and water submersion. The DCS is the concentration of a particular radionuclide in water, in air, or on the ground that results in a member of the public receiving 100 mrem (1 mSv) effective dose following continuous exposure for one year. In DOE-STD-1196-2011, DCSs were developed for the ingestion of water

Active-site monovalent cations revealed in a 1.55-Å-resolution hammerhead ribozyme structure.

PubMed

Anderson, Michael; Schultz, Eric P; Martick, Monika; Scott, William G

2013-10-23

We have obtained a 1.55-Å crystal structure of a hammerhead ribozyme derived from Schistosoma mansoni under conditions that permit detailed observations of Na(+) ion binding in the ribozyme's active site. At least two such Na(+) ions are observed. The first Na(+) ion binds to the N7 of G10.1 and the adjacent A9 phosphate in a manner identical with that previously observed for divalent cations. A second Na(+) ion binds to the Hoogsteen face of G12, the general base in the hammerhead cleavage reaction, thereby potentially dissipating the negative charge of the catalytically active enolate form of the nucleotide base. A potential but more ambiguous third site bridges the A9 and scissile phosphates in a manner consistent with that of previous predictions. Hammerhead ribozymes have been observed to be active in the presence of high concentrations of monovalent cations, including Na(+), but the mechanism by which monovalent cations substitute for divalent cations in hammerhead catalysis remains unclear. Our results enable us to suggest that Na(+) directly and specifically substitutes for divalent cations in the hammerhead active site. The detailed geometry of the pre-catalytic active-site complex is also revealed with a new level of precision, thanks to the quality of the electron density maps obtained from what is currently the highest-resolution ribozyme structure in the Protein Data Bank. Copyright © 2013 Elsevier Ltd. All rights reserved.

Polycyclic aromatic hydrocarbons (PAHs) in soils and vegetation near an e-waste recycling site in South China: concentration, distribution, source, and risk assessment.

PubMed

Wang, Yan; Tian, Zhongjing; Zhu, Haolin; Cheng, Zhineng; Kang, Meiling; Luo, Chunling; Li, Jun; Zhang, Gan

2012-11-15

This study determined the concentrations of PAHs generated from e-waste recycling activities and their potential impacts on soil, vegetation, and human health. The total PAH concentrations in soils and plants ranged from 127 to 10,600 and 199 to 2420 ng/g, respectively. Samples from an e-waste burning site had higher PAH concentrations than samples from adjacent locations. The PAHs in plants varied with plant species and tissue, and Lactuca sativa L. contained the highest PAHs of all the vegetable species. Various land use types showed different PAH concentrations in soils, with vegetable fields showing higher concentrations than paddy fields. Low molecular weight PAHs, such as phenanthrene, were the predominant congeners in soils, whereas high molecular weight PAHs, such as fluoranthene, pyrene, and benzo[a]anthracene, were enriched in plants relative to soils. Dissimilar PAH profiles in soil and the corresponding vegetation indicated that the uptake of PAHs by plants was selective. A source analysis showed that the contamination by PAHs originated primarily from the open burning of e-waste. The total daily intakes of PAHs and carcinogenic PAHs through vegetables at the e-waste dismantling site were estimated to be 279 and 108 ng/kg/d, respectively, indicating that the consumption of vegetables grown near e-waste recycling sites is risky and should be completely avoided. Copyright © 2012 Elsevier B.V. All rights reserved.

Endocrine disrupting activities of surface water associated with a West Virginia oil and gas industry wastewater disposal site.

PubMed

Kassotis, Christopher D; Iwanowicz, Luke R; Akob, Denise M; Cozzarelli, Isabelle M; Mumford, Adam C; Orem, William H; Nagel, Susan C

2016-07-01

Currently, >95% of end disposal of hydraulic fracturing wastewater from unconventional oil and gas operations in the US occurs via injection wells. Key data gaps exist in understanding the potential impact of underground injection on surface water quality and environmental health. The goal of this study was to assess endocrine disrupting activity in surface water at a West Virginia injection well disposal site. Water samples were collected from a background site in the area and upstream, on, and downstream of the disposal facility. Samples were solid-phase extracted, and extracts assessed for agonist and antagonist hormonal activities for five hormone receptors in mammalian and yeast reporter gene assays. Compared to reference water extracts upstream and distal to the disposal well, samples collected adjacent and downstream exhibited considerably higher antagonist activity for the estrogen, androgen, progesterone, glucocorticoid and thyroid hormone receptors. In contrast, low levels of agonist activity were measured in upstream/distal sites, and were inhibited or absent at downstream sites with significant antagonism. Concurrent analyses by partner laboratories (published separately) describe the analytical and geochemical profiling of the water; elevated conductivity as well as high sodium, chloride, strontium, and barium concentrations indicate impacts due to handling of unconventional oil and gas wastewater. Notably, antagonist activities in downstream samples were at equivalent authentic standard concentrations known to disrupt reproduction and/or development in aquatic animals. Given the widespread use of injection wells for end-disposal of hydraulic fracturing wastewater, these data raise concerns for human and animal health nearby. Copyright © 2016 Elsevier B.V. All rights reserved.

Endocrine disrupting activities of surface water associated with a West Virginia oil and gas industry wastewater disposal site

USGS Publications Warehouse

Kassotis, Christopher D.; Iwanowicz, Luke R.; Akob, Denise M.; Cozzarelli, Isabelle M.; Mumford, Adam; Orem, William H.; Nagel, Susan C.

2016-01-01

Currently, >95% of end disposal of hydraulic fracturing wastewater from unconventional oil and gas operations in the US occurs via injection wells. Key data gaps exist in understanding the potential impact of underground injection on surface water quality and environmental health. The goal of this study was to assess endocrine disrupting activity in surface water at a West Virginia injection well disposal site. Water samples were collected from a background site in the area and upstream, on, and downstream of the disposal facility. Samples were solid-phase extracted, and extracts assessed for agonist and antagonist hormonal activities for five hormone receptors in mammalian and yeast reporter gene assays. Compared to reference water extracts upstream and distal to the disposal well, samples collected adjacent and downstream exhibited considerably higher antagonist activity for the estrogen, androgen, progesterone, glucocorticoid and thyroid hormone receptors. In contrast, low levels of agonist activity were measured in upstream/distal sites, and were inhibited or absent at downstream sites with significant antagonism. Concurrent analyses by partner laboratories (published separately) describe the analytical and geochemical profiling of the water; elevated conductivity as well as high sodium, chloride, strontium, and barium concentrations indicate impacts due to handling of unconventional oil and gas wastewater. Notably, antagonist activities in downstream samples were at equivalent authentic standard concentrations known to disrupt reproduction and/or development in aquatic animals. Given the widespread use of injection wells for end-disposal of hydraulic fracturing wastewater, these data raise concerns for human and animal health nearby.

Indoor radon activity concentration measurements in the great historical museums of University of Naples, Italy.

PubMed

Quarto, Maria; Pugliese, Mariagabriella; Loffredo, Filomena; La Verde, Giuseppe; Roca, Vincenzo

2016-01-01

Indoor radon activity concentrations were measured in seven Museums of University of Naples, very old buildings of great historical value. The measurements were performed using a time-integrated technique based on LR-115 solid-state nuclear track detectors. The annual average concentrations were found to range from 40 up to 1935 Bq m(-3) and in 26 % of measurement sites, the values were higher than 500 Bq m(-3) which is the limit value of Italian legislation for workplace. Moreover, we analysed the seasonal variations of radon concentrations observing the highest average in cold weather than in warm. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Metal active site elasticity linked to activation of homocysteine in methionine synthases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Koutmos, Markos; Pejchal, Robert; Bomer, Theresa M.

2008-04-02

Enzymes possessing catalytic zinc centers perform a variety of fundamental processes in nature, including methyl transfer to thiols. Cobalamin-independent (MetE) and cobalamin-dependent (MetH) methionine synthases are two such enzyme families. Although they perform the same net reaction, transfer of a methyl group from methyltetrahydrofolate to homocysteine (Hcy) to form methionine, they display markedly different catalytic strategies, modular organization, and active site zinc centers. Here we report crystal structures of zinc-replete MetE and MetH, both in the presence and absence of Hcy. Structural investigation of the catalytic zinc sites of these two methyltransferases reveals an unexpected inversion of zinc geometry uponmore » binding of Hcy and displacement of an endogenous ligand in both enzymes. In both cases a significant movement of the zinc relative to the protein scaffold accompanies inversion. These structures provide new information on the activation of thiols by zinc-containing enzymes and have led us to propose a paradigm for the mechanism of action of the catalytic zinc sites in these and related methyltransferases. Specifically, zinc is mobile in the active sites of MetE and MetH, and its dynamic nature helps facilitate the active site conformational changes necessary for thiol activation and methyl transfer.« less

Tracking Dissolved Methane Concentrations near Active Seeps and Gas Hydrates: Sea of Japan.

NASA Astrophysics Data System (ADS)

Snyder, G. T.; Aoki, S.; Matsumoto, R.; Tomaru, H.; Owari, S.; Nakajima, R.; Doolittle, D. F.; Brant, B.

2015-12-01

A number of regions in the Sea of Japan are known for active gas venting and for gas hydrate exposures on the sea floor. In this investigation we employed several gas sensors mounted on a ROV in order to determine the concentrations of dissolved methane in the water near these sites. Methane concentrations were determined during two-second intervals throughout each ROV deployment during the cruise. The methane sensor deployments were coupled with seawater sampling using Niskin bottles. Dissolved gas concentrations were later measured using gas chromatography in order to compare with the sensor results taken at the same time. The observed maximum dissolved methane concentrations were much lower than saturation values, even when the ROV manipulators were in contact with gas hydrate. Nonetheless, dissolved concentrations did reach several thousands of nmol/L near gas hydrate exposures and gas bubbles, more than two orders of magnitude over the instrumental detection limits. Most of the sensors tested were able to detect dissolved methane concentrations as low as 10 nmol/L which permitted detection when the ROV approached methane plume sites, even from several tens of meters above the sea floor. Despite the low detection limits, the methane sensors showed variable response times when returning to low-background seawater (~5nM). For some of the sensors, the response time necessary to return to background values occurred in a matter of minutes, while for others it took several hours. Response time, as well as detection limit, should be an important consideration when selecting methane sensors for ROV or AUV investigations. This research was made possible, in part, through funding provided by the Japanese Ministry of Economy, Trade and Industry (METI).

Concentration of Actinides in Plant Mounds at Safety Test Nuclear Sites in Nevada

DOE Office of Scientific and Technical Information (OSTI.GOV)

David S. Shafer; Jenna Gommes

Plant mounds or blow-sand mounds are accumulations of soil particles and plant debris around large shrubs and are common features in deserts in the southwestern United States. Believed to be an important factor in their formation, the shrubs create surface roughness that causes wind-suspended particles to be deposited and resist further suspension. Shrub mounds occur in some plant communities on the Nevada Test Site, the Nevada Test and Training Range (NTTR), and Tonopah Test Range (TTR), including areas of surface soil contamination from past nuclear testing. In the 1970s as part of early studies to understand properties of actinides inmore » the environment, the Nevada Applied Ecology Group (NAEG) examined the accumulation of isotopes of Pu, {sup 241}Am, and U in plant mounds at safety test sites. The NAEG studies found concentrations of these contaminants to be greater in shrub mounds than in the surrounding areas of desert pavement. For example, at Project 57 on the NTTR, it was estimated that 15 percent of the radionuclide inventory of the site was associated with shrub mounds, which accounted for 17 percent of the surface area of the site, a ratio of inventory to area of 0.85. At Clean Slate III at the TTR, 29 percent of the inventory was associated with approximately 32 percent of the site covered by shrub mounds, a ratio of 0.91. While the total inventory of radionuclides in intershrub areas was greater, the ratio of radionuclide inventory to area was 0.40 and 0.38, respectively, at the two sites. The comparison between the shrub mounds and adjacent desert pavement areas was made for only the top 5 cm since radionuclides at safety test sites are concentrated in the top 5 cm of intershrub areas. Not accounting for radionuclides associated with the shrub mounds would cause the inventory of contaminants and potential exposure to be underestimated. As part of its Environmental Restoration Soils Subproject, the U.S. Department of Energy (DOE), National Nuclear

Effects of decontamination work on riverine radiocaesium activity concentrations in Fukushima affected area

NASA Astrophysics Data System (ADS)

Taniguchi, K.; Onda, Y.; Yoshimura, K.; Smith, H.; Brake, W.; Kubo, T.; Kuramoto, T.; Sato, T.; Onuma, S.

2016-12-01

Radionuclides such as Cs-134 and Cs-137 were widely distributed in the area affected by the accident at Fukushima Daiichi nuclear power plant. The radionuclides were deposited on the surface, absorbed by soil particles, and transported via river systems to Pacific Ocean due to rainfall events. In order to reduce air dose rate surrounding residential area, decontamination works have been conducted between 2013 and 2016 Fiscal Years. In paddy field and farmland contaminated by the fallout, 5 cm of surface soil was stripped, and then clean sands put on the surface. This work could reduce radiocaesium inventory, while the coverage of vegetation was significantly decreased. Therefore, runoff characteristics in the decontaminated area were different before and after the decontamination. Activity concentrations of particulate Cs-137 were measured in Abukuma river system and 8 small catchments located in coastal zone of Fukushima affected area. In all monitoring sites, Cs-137 concentrations have decreased over an entire monitoring period. Kuchibuto river, which is a tributary of Abukuma river showed significant effect of decontamination. In Yamakiya district, in the watershed of the tributary, the decontamination work had conducted from 2013 FY to December 2015. Particulate Cs-137 concentration at two monitoring sites located in the district showed around 30% of decline in the beginning of 2014 FY whereas the decline was not so significant at sites in lower reach of the tributary. Decontaminated paddy field and farmland can be judged as the important source of suspended sediments in the tributary.

Risks to children from exposure to lead in air during remedial or removal activities at Superfund sites: a case study of the RSR lead smelter Superfund site.

PubMed

Khoury, Ghassan A; Diamond, Gary L

2003-01-01

Superfund sites that are contaminated with lead and undergoing remedial action generate lead-enriched dust that can be released into the air. Activities that can emit lead-enriched dust include demolition of lead smelter buildings, stacks, and baghouses; on-site traffic of heavy construction vehicles; and excavation of soil. Typically, air monitoring stations are placed around the perimeter of a site of an ongoing remediation to monitor air lead concentrations that might result from site emissions. The National Ambient Air Quality (NAAQ) standard, established in 1978 to be a quarterly average of 1.5 microg/m(3), is often used as a trigger level for corrective action to reduce emissions. This study explored modeling approaches for assessing potential risks to children from air lead emissions from the RSR Superfund site in West Dallas, TX, during demolition and removal of a smelter facility. The EPA Integrated Exposure Uptake Biokinetic (IEUBK) model and the International Commission of Radiologic Protection (ICRP) lead model were used to simulate blood lead concentrations in children, based on monitored air lead concentrations. Although air lead concentrations at monitoring stations located in the downwind community intermittently exceeded the NAAQ standard, both models indicated that exposures to children in the community areas did not pose a significant long-term or acute risk. Long-term risk was defined as greater than 5% probability of a child having a long-term blood lead concentration that exceeded 10 microg/dl, which is the CDC and the EPA blood lead concern level. Short-term or acute risk was defined as greater than 5% probability of a child having a blood lead concentration on any given day that exceeded 20 microg/dl, which is the CDC trigger level for medical evaluation (this is not intended to imply that 20 microg/dl is a threshold for health effects in children exposed acutely to airborne lead). The estimated potential long-term and short-term exposures

Neural network model for the prediction of PM10 daily concentrations in two sites in the Western Mediterranean.

PubMed

de Gennaro, Gianluigi; Trizio, Livia; Di Gilio, Alessia; Pey, Jorge; Pérez, Noemi; Cusack, Michael; Alastuey, Andrés; Querol, Xavier

2013-10-01

An artificial neural network (ANN) was developed and tested to forecast PM10 daily concentration in two contrasted environments in NE Spain, a regional background site (Montseny), and an urban background site (Barcelona-CSIC), which was highly influenced by vehicular emissions. In order to predict 24-h average PM10 concentrations, the artificial neural network previously developed by Caselli et al. (2009) was improved by using hourly PM concentrations and deterministic factors such as a Saharan dust alert. In particular, the model input data for prediction were the hourly PM10 concentrations 1-day in advance, local meteorological data and information about air masses origin. The forecasted performance indexes for both sites were calculated and they showed better results for the regional background site in Montseny (R(2)=0.86, SI=0.75) than for urban site in Barcelona (R(2)=0.73, SI=0.58), influenced by local and sometimes unexpected sources. Moreover, a sensitivity analysis conducted to understand the importance of the different variables included among the input data, showed that local meteorology and air masses origin are key factors in the model forecasts. This result explains the reason for the improvement of ANN's forecasting performance at the Montseny site with respect to the Barcelona site. Moreover, the artificial neural network developed in this work could prove useful to predict PM10 concentrations, especially, at regional background sites such as those on the Mediterranean Basin which are primarily affected by long-range transports. Hence, the artificial neural network presented here could be a powerful tool for obtaining real time information on air quality status and could aid stakeholders in their development of cost-effective control strategies. © 2013 Elsevier B.V. All rights reserved.

Nicotinamide Cofactors Suppress Active-Site Labeling of Aldehyde Dehydrogenases.

PubMed

Stiti, Naim; Chandrasekar, Balakumaran; Strubl, Laura; Mohammed, Shabaz; Bartels, Dorothea; van der Hoorn, Renier A L

2016-06-17

Active site labeling by (re)activity-based probes is a powerful chemical proteomic tool to globally map active sites in native proteomes without using substrates. Active site labeling is usually taken as a readout for the active state of the enzyme because labeling reflects the availability and reactivity of active sites, which are hallmarks for enzyme activities. Here, we show that this relationship holds tightly, but we also reveal an important exception to this rule. Labeling of Arabidopsis ALDH3H1 with a chloroacetamide probe occurs at the catalytic Cys, and labeling is suppressed upon nitrosylation and oxidation, and upon treatment with other Cys modifiers. These experiments display a consistent and strong correlation between active site labeling and enzymatic activity. Surprisingly, however, labeling is suppressed by the cofactor NAD(+), and this property is shared with other members of the ALDH superfamily and also detected for unrelated GAPDH enzymes with an unrelated hydantoin-based probe in crude extracts of plant cell cultures. Suppression requires cofactor binding to its binding pocket. Labeling is also suppressed by ALDH modulators that bind at the substrate entrance tunnel, confirming that labeling occurs through the substrate-binding cavity. Our data indicate that cofactor binding adjusts the catalytic Cys into a conformation that reduces the reactivity toward chloroacetamide probes.

[Effect of propofol at different effect-site concentrations on approximate entropy of transient evoked otoacoustic emission signals in adults].

PubMed

Zhang, Xin-jian; Gu, Miao-ning; Xiao, Jin-fang

2009-01-01

To study the effect of propofol at different effect-site concentrations on approximate entropy (ApEn) of transient evoked otoacoustic emission (TEOAE) signals in adults and investigate the possibility of using ApEn for monitoring anesthesia depth. Fifteen ASA class I or II patients (aged 18-49 years with normal hearing) undergoing elective surgery under general anesthesia were enrolled in this study. Anesthesia was maintained with target-controlled infusion of propofol. With the effect-site concentrations of 1, 2, 3 and 4 microg/ml, TEOAE signals were monitored and recorded before and after anesthesia. ApEn of TEOAE in 4 frequency ranges (0-2, 1-3, 2.5-4.5, and 4-6 kHz) were calculated using MATLAB software. The ApEn of TEOAE in different frequency ranges showed no significant differences at the same effect-site concentration of propofol, or at different effect-site concentrations in the same frequency range (P>0.05). Anesthesia with propofol at different effect-site concentrations does not obviously affect ApEn of TEOAE signals in adults, and ApEn can not be used as the indicator for evaluating the depth of anesthesia.

A temporal and spatial assessment of TBT concentrations at dredged material disposal sites around the coast of England and Wales.

PubMed

Bolam, Thi; Barry, Jon; Law, Robin J; James, David; Thomas, Boby; Bolam, Stefan G

2014-02-15

Despite legislative interventions since the 1980s, contemporary concentrations of organotin compounds in marine sediments still impose restrictions on the disposal of dredged material in the UK. Here, we analyse temporal and spatial data to assess the effectiveness of the ban on the use of TBT paints in reducing concentrations at disposal sites. At a national scale, there was a statistically significant increase in the proportion of samples in which the concentration was below the limit of detection (LOD) from 1998 to 2010. This was observed for sediments both inside and outside the disposal sites. However, this temporal decline in organotin concentration is disposal site-specific. Of the four sites studied in detail, two displayed significant increases in proportion of samples below LOD over time. We argue that site-specificity in the effectiveness of the TBT ban results from variations in historical practices at source and unique environmental characteristics of each site. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

Quantum mechanical design of enzyme active sites.

PubMed

Zhang, Xiyun; DeChancie, Jason; Gunaydin, Hakan; Chowdry, Arnab B; Clemente, Fernando R; Smith, Adam J T; Handel, T M; Houk, K N

2008-02-01

The design of active sites has been carried out using quantum mechanical calculations to predict the rate-determining transition state of a desired reaction in presence of the optimal arrangement of catalytic functional groups (theozyme). Eleven versatile reaction targets were chosen, including hydrolysis, dehydration, isomerization, aldol, and Diels-Alder reactions. For each of the targets, the predicted mechanism and the rate-determining transition state (TS) of the uncatalyzed reaction in water is presented. For the rate-determining TS, a catalytic site was designed using naturalistic catalytic units followed by an estimation of the rate acceleration provided by a reoptimization of the catalytic site. Finally, the geometries of the sites were compared to the X-ray structures of related natural enzymes. Recent advances in computational algorithms and power, coupled with successes in computational protein design, have provided a powerful context for undertaking such an endeavor. We propose that theozymes are excellent candidates to serve as the active site models for design processes.

Electroencephalographic response following midazolam-induced general anesthesia: relationship to plasma and effect-site midazolam concentrations.

PubMed

Miyake, Wakako; Oda, Yutaka; Ikeda, Yuko; Hagihira, Satoshi; Iwaki, Hiroyoshi; Asada, Akira

2010-06-01

To examine the relationships between effect-site concentrations and electroencephalographic parameters after the induction of general anesthesia with midazolam. Twenty-four patients with American Society of Anesthesiologists status I or II were randomly allocated to receive either an intravenous (i.v.) bolus of midazolam 0.2 mg kg(-1) (small-dose group, n = 12) or 0.3 mg kg(-1) (large-dose group, n = 12) for induction of general anesthesia in a double-blind experimental design. The bispectral index (BIS), 95% spectral edge frequency (SEF95), spectral power density, and plasma concentrations of midazolam were measured for 60 min following the induction of general anesthesia. Plasma and simulated effect-site concentrations of midazolam were significantly higher in the large-dose group than in the small-dose group (P = 0.005 and <0.001, respectively). There was a correlation between the relative beta ratio and BIS (r (2) = 0.30, P < 0.001; n = 168); however, effect-site concentrations of midazolam showed no association with BIS, relative beta ratio, or SEF95 (r (2) = 0.07, 0.11 and 0.01, respectively; n = 168). The electroencephalographic spectral power density in the beta-band (>/=13 and <30 Hz) was significantly increased after induction and was significantly larger in the large-dose group than in the small-dose group (P = 0.009). Following the induction of general anesthesia with i.v. midazolam 0.2 or 0.3 mg kg(-1), the BIS was positively correlated with the relative beta ratio. Despite a rapid decrease in the plasma and effect-site concentrations of midazolam, the average BIS remained >60 for 60 min after induction, reflecting an increased power of the electroencephalographic high-frequency band.

Microstructure-scaled active sites imaging of a solid oxide fuel cell composite cathode

NASA Astrophysics Data System (ADS)

Nagasawa, Tsuyoshi; Hanamura, Katsunori

2017-11-01

Active sites for oxygen reduction reaction in strontium-doped lanthanum manganite (LSM)/scandia-stabilized zirconia (ScSZ) composite cathode of solid oxide fuel cell (SOFC) is visualized in microstructure scale by oxygen isotope labeling. In order to quench a reaction, a SOFC power generation equipment with a nozzle for direct helium gas impinging jet to the cell is prepared. A typical electrolyte-supported cell is operated by supplying 18O2 at 1073 K and abruptly quenched to room temperature. During the quench, the temperature of the cell is decreased from 1073 K to 673 K in 1 s. The 18O concentration distribution in the cross section of the quenched cathode is obtained by secondary ion mass spectrometry (SIMS) with a spatial resolution of 50 nm. The obtained 18O mapping gives the first visualization of highly distributed active sites in the composite cathode both in macroscopic and particle scales.

Site-Specific Reference Person Parameters and Derived Concentration Standards for the Savannah River Site

DOE PAGES

Stone, Daniel K.; Higley, Kathryn A.; Jannik, G. Timothy

2014-05-01

The U.S. Department of Energy Order 458.1 states that the compliance with the 1 mSv annual dose constraint to a member of the public may be demonstrated by calculating dose to the maximally exposed individual (MEI) or to a representative person. Historically, the MEI concept was used for dose compliance at the Savannah River Site (SRS) using adult dose coefficients and adult male usage parameters. For future compliance, SRS plans to use the representative person concept for dose estimates to members of the public. The representative person dose will be based on the reference person dose coefficients from the U.S.more » DOE Derived Concentration Technical Standard and on usage parameters specific to SRS for the reference and typical person. Usage parameters and dose coefficients were determined for inhalation, ingestion and external exposure pathways. The parameters for the representative person were used to calculate and tabulate SRS-specific derived concentration standards (DCSs) for the pathways not included in DOE-STD-1196-2011.« less

Relationships between airborne fungal spore concentration of Cladosporium and the summer climate at two sites in Britain

NASA Astrophysics Data System (ADS)

Hollins, P. D.; Kettlewell, P. S.; Atkinson, M. D.; Stephenson, D. B.; Corden, J. M.; Millington, W. M.; Mullins, J.

Cladosporium conidia have been shown to be important aeroallergens in many regions throughout the world, but annual spore concentrations vary considerably between years. Understanding these annual fluctuations may be of value in the clinical management of allergies. This study investigates the number of days in summer when spore concentration exceeds the allergenic threshold in relation to regional temperature and precipitation at two sites in England and Wales over 27 years. Results indicate that number of days in summer when the Cladosporium spores are above the allergenic concentration is positively correlated with regional temperature and negatively correlated with precipitation for both sites over the study period. Further analysis used a winter North Atlantic Oscillation index to explore the potential for long-range forecasting of the aeroallergen. For both spore measurement sites, a positive correlation exists between the winter North Atlantic Oscillation index and the number of days in summer above the allergenic threshold for Cladosporium spore concentration.

Mercury concentrations in water, and mercury and selenium concentrations in fish from Brownlee Reservoir and selected sites in Boise and Snake Rivers, Idaho and Oregon, 2013

USGS Publications Warehouse

MacCoy, Dorene E.

2014-01-01

Mercury (Hg) analyses were conducted on samples of sport fish and water collected from six sampling sites in the Boise and Snake Rivers, and Brownlee Reservoir to meet National Pollution Discharge and Elimination System (NPDES) permit requirements for the City of Boise, Idaho. A water sample was collected from each site during October and November 2013 by the City of Boise personnel and was analyzed by the Boise City Public Works Water Quality Laboratory. Total Hg concentrations in unfiltered water samples ranged from 0.73 to 1.21 nanograms per liter (ng/L) at five river sites; total Hg concentration was highest (8.78 ng/L) in a water sample from Brownlee Reservoir. All Hg concentrations in water samples were less than the EPA Hg chronic aquatic life criterion in Idaho (12 ng/L). The EPA recommended a water-quality criterion of 0.30 milligrams per kilogram (mg/kg) methylmercury (MeHg) expressed as a fish-tissue residue value (wet-weight MeHg in fish tissue). MeHg residue in fish tissue is considered to be equivalent to total Hg in fish muscle tissue and is referred to as Hg in this report. The Idaho Department of Environmental Quality adopted the EPA’s fish-tissue criterion and a reasonable potential to exceed (RPTE) threshold 20 percent lower than the criterion or greater than 0.24 mg/kg based on an average concentration of 10 fish from a receiving waterbody. NPDES permitted discharge to waters with fish having Hg concentrations exceeding 0.24 mg/kg are said to have a reasonable potential to exceed the water-quality criterion and thus are subject to additional permit obligations, such as requirements for increased monitoring and the development of a Hg minimization plan. The Idaho Fish Consumption Advisory Program (IFCAP) issues fish advisories to protect general and sensitive populations of fish consumers and has developed an action level of 0.22 mg/kg wet weight Hg in fish tissue. Fish consumption advisories are water body- and species-specific and are used to

Activated Prothrombin Complex Concentrate Versus 4-Factor Prothrombin Complex Concentrate for Vitamin K-Antagonist Reversal.

PubMed

Rowe, A Shaun; Dietrich, Scott K; Phillips, John W; Foster, Kaci E; Canter, Joshua R

2018-06-01

To compare the international normalized ratio normalization efficacy of activated prothrombin complex concentrates and 4-factor prothrombin complex concentrates and to evaluate the thrombotic complications in patients treated with these products for warfarin-associated hemorrhage. Retrospective, Multicenter Cohort. Large, Community, Teaching Hospital. Patients greater than 18 years old and received either activated prothrombin complex concentrate or 4-factor prothrombin complex concentrate for the treatment of warfarin-associated hemorrhage. We excluded those patients who received either agent for an indication other than warfarin-associated hemorrhage, pregnant, had a baseline international normalized ratio of less than 2, received a massive transfusion as defined by hospital protocol, received plasma for treatment of warfarin-associated hemorrhage, or were treated for an acute warfarin ingestion. Patients in the activated prothrombin complex concentrate group (enrolled from one hospital) with an international normalized ratio of less than 5 received 500 IU and those with an international normalized ratio greater than 5 received 1,000 IU. Patients in the 4-factor prothrombin complex concentrate (enrolled from a separate hospital) group received the Food and Drug Administration approved dosing algorithm. A total of 158 patients were included in the final analysis (activated prothrombin complex concentrate = 118; 4-factor prothrombin complex concentrate = 40). Those in the 4-factor prothrombin complex concentrate group had a higher pretreatment international normalized ratio (2.7 ± 1.8 vs 3.5 ± 2.9; p = 0.0164). However, the posttreatment international normalized ratio was similar between the groups. In addition, even when controlling for differences in the pretreatment international normalized ratio, there was no difference in the ability to achieve a posttreatment international normalized ratio of less than 1.4 (odds ratio, 0.753 [95% CI, 0.637-0.890]; p

Alteration of natural (37)Ar activity concentration in the subsurface by gas transport and water infiltration.

PubMed

Guillon, Sophie; Sun, Yunwei; Purtschert, Roland; Raghoo, Lauren; Pili, Eric; Carrigan, Charles R

2016-05-01

High (37)Ar activity concentration in soil gas is proposed as a key evidence for the detection of underground nuclear explosion by the Comprehensive Nuclear Test-Ban Treaty. However, such a detection is challenged by the natural background of (37)Ar in the subsurface, mainly due to Ca activation by cosmic rays. A better understanding and improved capability to predict (37)Ar activity concentration in the subsurface and its spatial and temporal variability is thus required. A numerical model integrating (37)Ar production and transport in the subsurface is developed, including variable soil water content and water infiltration at the surface. A parameterized equation for (37)Ar production in the first 15 m below the surface is studied, taking into account the major production reactions and the moderation effect of soil water content. Using sensitivity analysis and uncertainty quantification, a realistic and comprehensive probability distribution of natural (37)Ar activity concentrations in soil gas is proposed, including the effects of water infiltration. Site location and soil composition are identified as the parameters allowing for a most effective reduction of the possible range of (37)Ar activity concentrations. The influence of soil water content on (37)Ar production is shown to be negligible to first order, while (37)Ar activity concentration in soil gas and its temporal variability appear to be strongly influenced by transient water infiltration events. These results will be used as a basis for practical CTBTO concepts of operation during an OSI. Copyright © 2016 Elsevier Ltd. All rights reserved.

Altered binding of thioflavin t to the peripheral anionic site of acetylcholinesterase after phosphorylation of the active site by chlorpyrifos oxon or dichlorvos

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sultatos, L.G.; Kaushik, R.

2008-08-01

The peripheral anionic site of acetylcholinesterase, when occupied by a ligand, is known to modulate reaction rates at the active site of this important enzyme. The current report utilized the peripheral anionic site specific fluorogenic probe thioflavin t to determine if the organophosphates chlorpyrifos oxon and dichlorvos bind to the peripheral anionic site of human recombinant acetylcholinesterase, since certain organophosphates display concentration-dependent kinetics when inhibiting this enzyme. Incubation of 3 nM acetylcholinesterase active sites with 50 nM or 2000 nM inhibitor altered both the B{sub max} and K{sub d} for thioflavin t binding to the peripheral anionic site. However, thesemore » changes resulted from phosphorylation of Ser203 since increasing either inhibitor from 50 nM to 2000 nM did not alter further thioflavin t binding kinetics. Moreover, the organophosphate-induced decrease in B{sub max} did not represent an actual reduction in binding sites, but instead likely resulted from conformational interactions between the acylation and peripheral anionic sites that led to a decrease in the rigidity of bound thioflavin t. A drop in fluorescence quantum yield, leading to an apparent decrease in B{sub max}, would accompany the decreased rigidity of bound thioflavin t molecules. The organophosphate-induced alterations in K{sub d} represented changes in binding affinity of thioflavin t, with diethylphosphorylation of Ser203 increasing K{sub d}, and dimethylphosphorylation of Ser203 decreasing K{sub d}. These results indicate that chlorpyrifos oxon and dichlorvos do not bind directly to the peripheral anionic site of acetylcholinesterase, but can affect binding to that site through phosphorylation of Ser203.« less

Time and size resolved Measurement of Mass Concentration at an Urban Site

NASA Astrophysics Data System (ADS)

Karg, E.; Ferron, G. A.; Heyder, J.

2003-04-01

Time- and size-resolved measurements of ambient particles are necessary for modelling of atmospheric particle transport, the interpretation of particulate pollution events and the estimation of particle deposition in the human lungs. In the size range 0.01 - 2 µm time- and size-resolved data are obtained from differential mobility and optical particle counter measurements and from gravimetric filter analyses on a daily basis (PM2.5). By comparison of the time averaged and size integrated particle volume concentration with PM2.5 data, an average density of ambient particles can be estimated. Using this density, the number concentration data can be converted in time- and size-resolved mass concentration. Such measurements were carried out at a Munich downtown crossroads. The spectra were integrated in the size ranges 10 - 100 nm, 100 - 500 nm and 500 - 2000 nm. Particles in these ranges are named ultrafine, fine and coarse particles. These ranges roughly represent freshly emitted particles, aged/accumulated particles and particles entrained by erosive processes. An average number concentration of 80000 1/cm3 (s.d. 67%), a particle volume concentration of 53 µm3/cm3 (s.d. 76%) and a PM2.5 mass concentration of 27 µg/m3 was found. These particle volume- and PM2.5 data imply an average density of 0.51 g/cm3. Average number concentration showed 95.3%, 4.7% and 0.006% of the total particle concentration in the size ranges mentioned above. Mass concentration was 14.7%, 80.2% and 5.1% of the total, assuming the average density to be valid for all particles. The variability in mass concentration was 94%, 75% and 33% for the three size ranges. Nearly all ambient particles were in the ultrafine size range, whereas most of the mass concentration was in the fine size range. However, a considerable mass fraction of nearly 15% was found in the ultrafine size range. As the sampling site was close to the road and traffic emissions were the major source of the particles, 1) the

Trace metals from historical mining sites and past metallurgical activity remain bioavailable to wildlife today.

PubMed

Camizuli, Estelle; Scheifler, Renaud; Garnier, Stéphane; Monna, Fabrice; Losno, Rémi; Gourault, Claude; Hamm, Gilles; Lachiche, Caroline; Delivet, Guillaume; Chateau, Carmela; Alibert, Paul

2018-02-21

Throughout history, ancient human societies exploited mineral resources all over the world, even in areas that are now protected and considered to be relatively pristine. Here, we show that past mining still has an impact on wildlife in some French protected areas. We measured cadmium, copper, lead, and zinc concentrations in topsoils and wood mouse kidneys from sites located in the Cévennes and the Morvan. The maximum levels of metals in these topsoils are one or two orders of magnitude greater than their commonly reported mean values in European topsoils. The transfer to biota was effective, as the lead concentration (and to a lesser extent, cadmium) in wood mouse kidneys increased with soil concentration, unlike copper and zinc, providing direct evidence that lead emitted in the environment several centuries ago is still bioavailable to free-ranging mammals. The negative correlation between kidney lead concentration and animal body condition suggests that historical mining activity may continue to play a role in the complex relationships between trace metal pollution and body indices. Ancient mining sites could therefore be used to assess the long-term fate of trace metals in soils and the subsequent risks to human health and the environment.

Environmental controls on the activity of aquifer microbial communities in the 300 area of the Hanford site.

PubMed

Konopka, Allan; Plymale, Andrew E; Carvajal, Denny A; Lin, Xueju; McKinley, James P

2013-11-01

Aquifer microbes in the 300 Area of the Hanford Site in southeastern Washington State, USA, are located in an oligotrophic environment and are periodically exposed to U(VI) concentrations that can range up to 10 μM in small sediment fractures. Assays of (3)H-leucine incorporation indicated that both sediment-associated and planktonic microbes were metabolically active, and that organic C was growth-limiting in the sediments. Although bacteria suspended in native groundwater retained high activity when exposed to 100 μM U(VI), they were inhibited by U(VI) <1 μM in synthetic groundwater that lacked added bicarbonate. Chemical speciation modeling suggested that positively charged species and particularly (UO2)3(OH)5 (+) rose in concentration as more U(VI) was added to synthetic groundwater, but that carbonate complexes dominated U(VI) speciation in natural groundwater. U toxicity was relieved when increasing amounts of bicarbonate were added to synthetic groundwater containing 4.5 μM U(VI). Pertechnetate, an oxyanion that is another contaminant of concern at the Hanford Site, was not toxic to groundwater microbes at concentrations up to 125 μM.

Optimal effect-site concentration of remifentanil when combined with dexmedetomidine in patients undergoing cystoscopy

PubMed Central

Heo, Bongha; Kim, Minsun; Lee, Hyunjung; Park, Sanghee

2014-01-01

Background Cystoscopic procedure is a very common practice in the field of urology due to its ability to survey the bladder for a variety of indications. However, patients who undergo cystoscopy feel intense pain and discomfort. This study investigated the half maximal effective concentration (EC50) of remifentanil in preventing cystoscope insertion pain under sedation using dexmedetomidine. Methods The study was prospectively conducted on 18 male patients, aged 18 to 65. Remifentail infusion was initiated together with dexmedetomidine, and started at a dose of 2.4 ng/ml on the first patient. The effect-site concentration (Ce) of remifentanil for each subsequent patient was determined by the previous patient's response using Dixon's up-and-down method with an interval of 0.3 ng/ml. Patients received a loading dose of 1.0 µg/kg dexmedetomidine over 10 minutes, followed by a maintenance dose of 0.6 µg/kg/hr. After the patient's OAA/S score (Observer's Assessment of Alertness/Sedation scale) reached 3-4, and the Ce of remifentanil reached target concentration, the urologist was allowed to insert the cystoscope and the pain responses were observed. Results The effect-site concentration of remifentanil required to prevent cystoscope insertion pain in 50% of patients under sedation using dexmedetomidine was 1.30 ± 0.12 ng/ml by Dixon's up-and-down method. The logistic regression curve of the probability of response showed that the EC50 and EC95 values (95% confidence limits) of remifentanil were 1.33 ng/ml (1.12-1.52 ng/ml) and 1.58 ng/ml (1.44-2.48 ng/ml), respectively. Conclusions Cystoscopic procedure can be carried out successfully without any pain or adverse effects by optimal remifentanil effect-site concentration (EC50, 1.33 ng/ml; EC95, 1.58 ng/ ml) combined with sedation using dexmedetomidine. PMID:24567812

Active and regulatory sites of cytosolic 5'-nucleotidase.

PubMed

Pesi, Rossana; Allegrini, Simone; Careddu, Maria Giovanna; Filoni, Daniela Nicole; Camici, Marcella; Tozzi, Maria Grazia

2010-12-01

Cytosolic 5'-nucleotidase (cN-II), which acts preferentially on 6-hydroxypurine nucleotides, is essential for the survival of several cell types. cN-II catalyses both the hydrolysis of nucleotides and transfer of their phosphate moiety to a nucleoside acceptor through formation of a covalent phospho-intermediate. Both activities are regulated by a number of phosphorylated compounds, such as diadenosine tetraphosphate (Ap₄A), ADP, ATP, 2,3-bisphosphoglycerate (BPG) and phosphate. On the basis of a partial crystal structure of cN-II, we mutated two residues located in the active site, Y55 and T56. We ascertained that the ability to catalyse the transfer of phosphate depends on the presence of a bulky residue in the active site very close to the aspartate residue that forms the covalent phospho-intermediate. The molecular model indicates two possible sites at which adenylic compounds may interact. We mutated three residues that mediate interaction in the first activation site (R144, N154, I152) and three in the second (F127, M436 and H428), and found that Ap₄A and ADP interact with the same site, but the sites for ATP and BPG remain uncertain. The structural model indicates that cN-II is a homotetrameric protein that results from interaction through a specific interface B of two identical dimers that have arisen from interaction of two identical subunits through interface A. Point mutations in the two interfaces and gel-filtration experiments indicated that the dimer is the smallest active oligomerization state. Finally, gel-filtration and light-scattering experiments demonstrated that the native enzyme exists as a tetramer, and no further oligomerization is required for enzyme activation. © 2010 The Authors Journal compilation © 2010 FEBS.

Water in the Active Site of Ketosteroid Isomerase

PubMed Central

Hanoian, Philip; Hammes-Schiffer, Sharon

2011-01-01

Classical molecular dynamics simulations were utilized to investigate the structural and dynamical properties of water in the active site of ketosteroid isomerase (KSI) to provide insight into the role of these water molecules in the enzyme-catalyzed reaction. This reaction is thought to proceed via a dienolate intermediate that is stabilized by hydrogen bonding with residues Tyr16 and Asp103. A comparative study was performed for the wild-type (WT) KSI and the Y16F, Y16S, and Y16F/Y32F/Y57F (FFF) mutants. These systems were studied with three different bound ligands: equilenin, which is an intermediate analog, and the intermediate states of two steroid substrates. Several distinct water occupation sites were identified in the active site of KSI for the WT and mutant systems. Three additional sites were identified in the Y16S mutant that were not occupied in WT KSI or the other mutants studied. The number of water molecules directly hydrogen bonded to the ligand oxygen was approximately two waters in the Y16S mutant, one water in the Y16F and FFF mutants, and intermittent hydrogen bonding of one water molecule in WT KSI. The molecular dynamics trajectories of the Y16F and FFF mutants reproduced the small conformational changes of residue 16 observed in the crystal structures of these two mutants. Quantum mechanical/molecular mechanical calculations of 1H NMR chemical shifts of the protons in the active site hydrogen-bonding network suggest that the presence of water in the active site does not prevent the formation of short hydrogen bonds with far-downfield chemical shifts. The molecular dynamics simulations indicate that the active site water molecules exchange much more frequently for WT KSI and the FFF mutant than for the Y16F and Y16S mutants. This difference is most likely due to the hydrogen-bonding interaction between Tyr57 and an active site water molecule that is persistent in the Y16F and Y16S mutants but absent in the FFF mutant and significantly less

Water in the active site of ketosteroid isomerase.

PubMed

Hanoian, Philip; Hammes-Schiffer, Sharon

2011-08-09

Classical molecular dynamics simulations were utilized to investigate the structural and dynamical properties of water in the active site of ketosteroid isomerase (KSI) to provide insight into the role of these water molecules in the enzyme-catalyzed reaction. This reaction is thought to proceed via a dienolate intermediate that is stabilized by hydrogen bonding with residues Tyr16 and Asp103. A comparative study was performed for the wild-type (WT) KSI and the Y16F, Y16S, and Y16F/Y32F/Y57F (FFF) mutants. These systems were studied with three different bound ligands: equilenin, which is an intermediate analog, and the intermediate states of two steroid substrates. Several distinct water occupation sites were identified in the active site of KSI for the WT and mutant systems. Three additional sites were identified in the Y16S mutant that were not occupied in WT KSI or the other mutants studied. The number of water molecules directly hydrogen bonded to the ligand oxygen was approximately two in the Y16S mutant and one in the Y16F and FFF mutants, with intermittent hydrogen bonding of one water molecule in WT KSI. The molecular dynamics trajectories of the Y16F and FFF mutants reproduced the small conformational changes of residue 16 observed in the crystal structures of these two mutants. Quantum mechanical/molecular mechanical calculations of (1)H NMR chemical shifts of the protons in the active site hydrogen-bonding network suggest that the presence of water in the active site does not prevent the formation of short hydrogen bonds with far-downfield chemical shifts. The molecular dynamics simulations indicate that the active site water molecules exchange much more frequently for WT KSI and the FFF mutant than for the Y16F and Y16S mutants. This difference is most likely due to the hydrogen-bonding interaction between Tyr57 and an active site water molecule that is persistent in the Y16F and Y16S mutants but absent in the FFF mutant and significantly less probable

Active Site Characterization of Proteases Sequences from Different Species of Aspergillus.

PubMed

Morya, V K; Yadav, Virendra K; Yadav, Sangeeta; Yadav, Dinesh

2016-09-01

A total of 129 proteases sequences comprising 43 serine proteases, 36 aspartic proteases, 24 cysteine protease, 21 metalloproteases, and 05 neutral proteases from different Aspergillus species were analyzed for the catalytically active site residues using MEROPS database and various bioinformatics tools. Different proteases have predominance of variable active site residues. In case of 24 cysteine proteases of Aspergilli, the predominant active site residues observed were Gln193, Cys199, His364, Asn384 while for 43 serine proteases, the active site residues namely Asp164, His193, Asn284, Ser349 and Asp325, His357, Asn454, Ser519 were frequently observed. The analysis of 21 metalloproteases of Aspergilli revealed Glu298 and Glu388, Tyr476 as predominant active site residues. In general, Aspergilli species-specific active site residues were observed for different types of protease sequences analyzed. The phylogenetic analysis of these 129 proteases sequences revealed 14 different clans representing different types of proteases with diverse active site residues.

Half-of-the-sites reactivity of outer-membrane phospholipase A against an active-site-directed inhibitor.

PubMed

Ubarretxena-Belandia, I; Cox, R C; Dijkman, R; Egmond, M R; Verheij, H M; Dekker, N

1999-03-01

The reaction of a novel active-site-directed phospholipase A1 inhibitor with the outer-membrane phospholipase A (OMPLA) was investigated. The inhibitor 1-p-nitrophenyl-octylphosphonate-2-tridecylcarbamoyl-3-et hanesulfonyl -amino-3-deoxy-sn-glycerol irreversibly inactivated OMPLA. The inhibition reaction did not require the cofactor calcium or an unprotonated active-site His142. The inhibition of the enzyme solubilized in hexadecylphosphocholine micelles was characterized by a rapid (t1/2 = 20 min) and complete loss of enzymatic activity, concurrent with the covalent modification of 50% of the active-site serines, as judged from the amount of p-nitrophenolate (PNP) released. Modification of the remaining 50% occurred at a much lower rate, indicative of half-of-the-sites reactivity against the inhibitor of this dimeric enzyme. Inhibition of monomeric OMPLA solubilized in hexadecyl-N,N-dimethyl-1-ammonio-3-propanesulfonate resulted in an equimolar monophasic release of PNP, concurrent with the loss of enzymatic activity (t1/2 = 14 min). The half-of-the-sites reactivity is discussed in view of the dimeric nature of this enzyme.

Physical activity and cancer risk: dose-response and cancer, all sites and site-specific.

PubMed

Thune, I; Furberg, A S

2001-06-01

The association between physical activity and overall and site-specific cancer risk is elaborated in relation to whether any observed dose-response association between physical activity and cancer can be interpreted in terms of how much physical activity (type, intensity, duration, frequency) is needed to influence site- and gender-specific cancer risk. Observational studies were reviewed that have examined the independent effect of the volume of occupational physical activity (OPA) and/or leisure time physical activity (LPA) on overall and site-specific cancer risk. The evidence of cohort and case-control studies suggests that both leisure time and occupational physical activity protect against overall cancer risk, with a graded dose-response association suggested in both sexes. Confounding effects such as diet, body weight, and parity are often included as a covariate in the analyses, with little influence on the observed associations. A crude graded inverse dose-response association was observed between physical activity and colon cancer in 48 studies including 40,674 colon/colorectal cancer cases for both sexes. A dose-response effect of physical activity on colon cancer risk was especially observed, when participation in activities of at least moderate activity (>4.5 MET) and demonstrated by activities expressed as MET-hours per week. An observed inverse association with a dose-response relationship between physical activity and breast cancer was also identified in the majority of the 41 studies including 108,031 breast cancer cases. The dose-response relationship was in particular observed in case-control studies and supported by observations in cohort studies when participation in activities of at least moderate activity (>4.5 MET) and demonstrated by activities expressed by MET-hours per week. This association between physical activity and breast cancer risk is possibly dependent on age at exposure, age at diagnosis, menopausal status and other effect

The significance of low substrate concentration measurements for mechanistic interpretation in cholinesterases.

PubMed

Stojan, Jure

2013-03-25

Cholinesterases do not follow the Michaelis-Menten kinetics. In the past, many reaction schemes were suggested to explain their complex interactions during the substrate turnover. Covalent catalysis was recognized very early and therefore, double intermediate traditional reaction scheme for the hydrolysis of good substrates at low concentrations was postulated. However, at intermediate and high substrate concentrations homotropic pseudocooperative effects take place in all cholinesterases, due to the nature of their buried active center. In this study, the significance and usefulness of experimental data obtained at low substrate concentrations, where only one substrate molecule accesses the active site at a time, are to be specified for the overall mechanistic evaluations. Indeed, different interpretations are expected when data are processed with equations derived from different reaction schemes. Consequently, the scheme with two substrate binding sites which comprises the structurally evidenced fully occupied active site as ultimate cause for substantially decreased cholinesterase activity at extremely high substrate concentrations is considered here. A special emphasis is put on butyrylcholinesterase, the enzyme with the largest active site among cholinesterases, where the pseudocooperative effects appear at much higher concentrations than in acetylcholinesterases. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Active Site Metal Occupancy and Cyclic Di-GMP Phosphodiesterase Activity of Thermotoga maritima HD-GYP.

PubMed

Miner, Kyle D; Kurtz, Donald M

2016-02-16

HD-GYPs make up a subclass of the metal-dependent HD phosphohydrolase superfamily and catalyze conversion of cyclic di(3',5')-guanosine monophosphate (c-di-GMP) to 5'-phosphoguanylyl-(3'→5')-guanosine (pGpG) and GMP. Until now, the only reported crystal structure of an HD-GYP that also exhibits c-di-GMP phosphodiesterase activity contains a His/carboxylate ligated triiron active site. However, other structural and phylogenetic correlations indicate that some HD-GYPs contain dimetal active sites. Here we provide evidence that an HD-GYP c-di-GMP phosphodiesterase, TM0186, from Thermotoga maritima can accommodate both di- and trimetal active sites. We show that an as-isolated iron-containing TM0186 has an oxo/carboxylato-bridged diferric site, and that the reduced (diferrous) form is necessary and sufficient to catalyze conversion of c-di-GMP to pGpG, but that conversion of pGpG to GMP requires more than two metals per active site. Similar c-di-GMP phosphodiesterase activities were obtained with divalent iron or manganese. On the basis of activity correlations with several putative metal ligand residue variants and molecular dynamics simulations, we propose that TM0186 can accommodate both di- and trimetal active sites. Our results also suggest that a Glu residue conserved in a subset of HD-GYPs is required for formation of the trimetal site and can also serve as a labile ligand to the dimetal site. Given the anaerobic growth requirement of T. maritima, we suggest that this HD-GYP can function in vivo with either divalent iron or manganese occupying di- and trimetal sites.

Diffusional correlations among multiple active sites in a single enzyme.

PubMed

Echeverria, Carlos; Kapral, Raymond

2014-04-07

Simulations of the enzymatic dynamics of a model enzyme containing multiple substrate binding sites indicate the existence of diffusional correlations in the chemical reactivity of the active sites. A coarse-grain, particle-based, mesoscopic description of the system, comprising the enzyme, the substrate, the product and solvent, is constructed to study these effects. The reactive and non-reactive dynamics is followed using a hybrid scheme that combines molecular dynamics for the enzyme, substrate and product molecules with multiparticle collision dynamics for the solvent. It is found that the reactivity of an individual active site in the multiple-active-site enzyme is reduced substantially, and this effect is analyzed and attributed to diffusive competition for the substrate among the different active sites in the enzyme.

Fog chemistry at three sites in Norway

NASA Astrophysics Data System (ADS)

Wang, Youliang; Zhang, Jinwei; Marcotte, Aurelie R.; Karl, Matthias; Dye, Christian; Herckes, Pierre

2015-01-01

Fog composition was investigated at three sites in Norway, one in suburban Oslo and two coastal sites in the area of the Mongstad refinery. Overall fog frequency during the study periods was low. Fog pH was around 5 with slightly lower values at Hakadal, the suburban site, compared to the coastal sites, which were slightly above 5. Major ions at the coastal sites were sodium and chloride consistent with the marine environment. The ion chemistry at the suburban site was dominated by ammonium, sulfate and nitrate, consistent with fogs in anthropogenically impacted environments. Overall concentrations of major ions were very low, orders of magnitude lower than those in polluted urban fogs. Organic matter concentrations were also low (< 3 mgC/L) consistent with limited anthropogenic impact and little biogenic activity in the winter months. Selected amine concentrations were determined and ranged from nanomolar concentrations for ethylamines to several hundred nanomolar concentrations for dimethylamine, the most abundant amine investigated. While N-nitrosodimehylamine was detected in fog, the concentrations were very low in the fogs.

Testing the applicability of rapid on-site enzymatic activity detection for surface water monitoring

NASA Astrophysics Data System (ADS)

Stadler, Philipp; Vogl, Wolfgang; Juri, Koschelnik; Markus, Epp; Maximilian, Lackner; Markus, Oismüller; Monika, Kumpan; Peter, Strauss; Regina, Sommer; Gabriela, Ryzinska-Paier; Farnleitner Andreas, H.; Matthias, Zessner

2015-04-01

bacteria. Therefore the applicability of on-site enzymatic activity determination as a direct surrogate or proxy parameter for microbiological standard assays and quantification of fecal indicator bacteria (FIB) concentration could not be approved and further research in this field is necessary. Presently we conclude that rapid on-site detection of enzymatic activity is applicable for surface water monitoring and that it constitutes a complementary on-site monitoring parameter with high potential. Selection of the type of measured enzymatic activities has to be done on a catchment-specific basis and further work is needed to learn more about its detailed information characteristics in different habitats. The accomplishment of this method detecting continuous data of enzymatic activity in high temporal resolution caused by a target bacterial member is on the way of becoming a powerful tool for water quality monitoring, health related water quality- and early warning requirements.

Radionuclide activities and metal concentrations in sediments of the Sebou Estuary, NW Morocco, following a flooding event.

PubMed

Laissaoui, A; Mas, J L; Hurtado, S; Ziad, N; Villa, M; Benmansour, M

2013-06-01

This study presents metal concentrations (Fe, Mg, Mn, Co, Cu, Zn, Pb, As, Sr and V) and radionuclide activities ((40)K, (137)Cs, (210)Pb, (226)Ra, (228)Ac, (234)Th and (212)Pb) in surface deposits and a sediment core from the Sebou Estuary, Northwest Morocco. Samples were collected in April 2009, about 2 months after a flooding event, and analysed using a well-type coaxial gamma-ray detector and inductively coupled plasma-quadrupole mass spectrometry. Activities of radionuclides and concentrations of almost all elements in surface samples displayed only moderate spatial variation, suggesting homogenous deposition of eroded local soil in response to intense precipitation. Excess (210)Pb displayed relatively constant activity throughout the sediment core, preventing dating and precluding determination of the historical accumulation rates of pollutants at the core site. Some elements showed non-systematic trends with depth and displayed local maxima and minima. Other elements presented relatively systematic concentration trends or relatively constant levels with discrete maxima and/or minima. Except for Mn, Sr and Cr, all metal concentrations in sediment were below levels typical of polluted systems, suggesting little human impact or losses of metals from sediment particles.

Direct instrumental identification of catalytically active surface sites

NASA Astrophysics Data System (ADS)

Pfisterer, Jonas H. K.; Liang, Yunchang; Schneider, Oliver; Bandarenka, Aliaksandr S.

2017-09-01

The activity of heterogeneous catalysts—which are involved in some 80 per cent of processes in the chemical and energy industries—is determined by the electronic structure of specific surface sites that offer optimal binding of reaction intermediates. Directly identifying and monitoring these sites during a reaction should therefore provide insight that might aid the targeted development of heterogeneous catalysts and electrocatalysts (those that participate in electrochemical reactions) for practical applications. The invention of the scanning tunnelling microscope (STM) and the electrochemical STM promised to deliver such imaging capabilities, and both have indeed contributed greatly to our atomistic understanding of heterogeneous catalysis. But although the STM has been used to probe and initiate surface reactions, and has even enabled local measurements of reactivity in some systems, it is not generally thought to be suited to the direct identification of catalytically active surface sites under reaction conditions. Here we demonstrate, however, that common STMs can readily map the catalytic activity of surfaces with high spatial resolution: we show that by monitoring relative changes in the tunnelling current noise, active sites can be distinguished in an almost quantitative fashion according to their ability to catalyse the hydrogen-evolution reaction or the oxygen-reduction reaction. These data allow us to evaluate directly the importance and relative contribution to overall catalyst activity of different defects and sites at the boundaries between two materials. With its ability to deliver such information and its ready applicability to different systems, we anticipate that our method will aid the rational design of heterogeneous catalysts.

The identity of the active site of oxalate decarboxylase and the importance of the stability of active-site lid conformations1

PubMed Central

Just, Victoria J.; Burrell, Matthew R.; Bowater, Laura; McRobbie, Iain; Stevenson, Clare E. M.; Lawson, David M.; Bornemann, Stephen

2007-01-01

Oxalate decarboxylase (EC 4.1.1.2) catalyses the conversion of oxalate into carbon dioxide and formate. It requires manganese and, uniquely, dioxygen for catalysis. It forms a homohexamer and each subunit contains two similar, but distinct, manganese sites termed sites 1 and 2. There is kinetic evidence that only site 1 is catalytically active and that site 2 is purely structural. However, the kinetics of enzymes with mutations in site 2 are often ambiguous and all mutant kinetics have been interpreted without structural information. Nine new site-directed mutants have been generated and four mutant crystal structures have now been solved. Most mutants targeted (i) the flexibility (T165P), (ii) favoured conformation (S161A, S164A, D297A or H299A) or (iii) presence (Δ162–163 or Δ162–164) of a lid associated with site 1. The kinetics of these mutants were consistent with only site 1 being catalytically active. This was particularly striking with D297A and H299A because they disrupted hydrogen bonds between the lid and a neighbouring subunit only when in the open conformation and were distant from site 2. These observations also provided the first evidence that the flexibility and stability of lid conformations are important in catalysis. The deletion of the lid to mimic the plant oxalate oxidase led to a loss of decarboxylase activity, but only a slight elevation in the oxalate oxidase side reaction, implying other changes are required to afford a reaction specificity switch. The four mutant crystal structures (R92A, E162A, Δ162–163 and S161A) strongly support the hypothesis that site 2 is purely structural. PMID:17680775

Role of active site rigidity in activity: MD simulation and fluorescence study on a lipase mutant.

PubMed

Kamal, Md Zahid; Mohammad, Tabrez Anwar Shamim; Krishnamoorthy, G; Rao, Nalam Madhusudhana

2012-01-01

Relationship between stability and activity of enzymes is maintained by underlying conformational flexibility. In thermophilic enzymes, a decrease in flexibility causes low enzyme activity while in less stable proteins such as mesophiles and psychrophiles, an increase in flexibility is associated with enhanced enzyme activity. Recently, we identified a mutant of a lipase whose stability and activity were enhanced simultaneously. In this work, we probed the conformational dynamics of the mutant and the wild type lipase, particularly flexibility of their active site using molecular dynamic simulations and time-resolved fluorescence techniques. In contrast to the earlier observations, our data show that active site of the mutant is more rigid than wild type enzyme. Further investigation suggests that this lipase needs minimal reorganization/flexibility of active site residues during its catalytic cycle. Molecular dynamic simulations suggest that catalytically competent active site geometry of the mutant is relatively more preserved than wild type lipase, which might have led to its higher enzyme activity. Our study implies that widely accepted positive correlation between conformation flexibility and enzyme activity need not be stringent and draws attention to the possibility that high enzyme activity can still be accomplished in a rigid active site and stable protein structures. This finding has a significant implication towards better understanding of involvement of dynamic motions in enzyme catalysis and enzyme engineering through mutations in active site.

40 CFR 61.154 - Standard for active waste disposal sites.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 8 2011-07-01 2011-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an active... visible emissions to the outside air from any active waste disposal site where asbestos-containing waste...

40 CFR 61.154 - Standard for active waste disposal sites.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 40 Protection of Environment 9 2013-07-01 2013-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an active... visible emissions to the outside air from any active waste disposal site where asbestos-containing waste...

40 CFR 61.154 - Standard for active waste disposal sites.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 40 Protection of Environment 9 2012-07-01 2012-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an active... visible emissions to the outside air from any active waste disposal site where asbestos-containing waste...

40 CFR 61.154 - Standard for active waste disposal sites.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 8 2010-07-01 2010-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an active... visible emissions to the outside air from any active waste disposal site where asbestos-containing waste...

40 CFR 61.154 - Standard for active waste disposal sites.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 40 Protection of Environment 9 2014-07-01 2014-07-01 false Standard for active waste disposal... for Asbestos § 61.154 Standard for active waste disposal sites. Each owner or operator of an active... visible emissions to the outside air from any active waste disposal site where asbestos-containing waste...

Microbial activity of trench leachates from shallow-land, low-level radioactive waste disposal sites.

PubMed Central

Francis, A J; Dobbs, S; Nine, B J

1980-01-01

Trench leachate samples collected anoxically from shallow-land, low-level radioactive waste disposal sites were analyzed for total aerobic and anaerobic populations, sulfate reducers, denitrifiers, and methanogens. Among the several aerobic and anaerobic bacteria isolated, only Bacillus sp., Pseudomonas sp., Citrobacter sp., and Clostridium sp. were identified. Mixed bacterial cultures isolated from the trench leachates were able to grow anaerobically in trench leachates, which indicates that the radionuclides and organic chemicals present were not toxic to these bacteria. Changes in concentrations of several of the organic constituents of the waste leachate samples were observed due to anaerobic microbial activity. Growth of a mixed culture of trench-water bacteria in media containing a mixture of radionuclides, 60Co, 85Sr, and 134,137Cs, was not affected at total activity concentrations of 2.6 X 10(2) and 2.7 X 10(3) pCi/ml. PMID:7406490

Concentration and trend of 1,4-dioxane in wells sampled during 2002–2017 in the vicinity of the Tucson International Airport Area Superfund Site, Arizona

USGS Publications Warehouse

Tillman, Fred D.

2017-09-25

Industrial activities causing extensive groundwater contamination led to the listing of the Tucson International Airport Area (TIAA) as a Superfund Site in 1983. Early groundwater investigations identified volatile organic compounds (VOCs), including the chlorinated solvents trichloroethylene (TCE) and perchloroethylene (PCE), in wells in the area. Several responsible parties were identified and cleanup activities began in the late 1980s. In 2002, the compound 1,4-dioxane was discovered in wells in the area and has since been detected in measurable concentrations throughout the site. The U.S. Environmental Protection Agency (USEPA) classifies 1,4-dioxane as a likely human carcinogen.The purpose of this map is to present 1,4-dioxane concentrations in wells sampled from 2002 through mid-2017 in the TIAA Superfund Site area to indicate both the current status and trends in 1,4-dioxane groundwater contamination. This map includes data from wells in the commercial and residential community in the TIAA and does not include data from wells in suspected or confirmed source areas, such as Air Force Plant 44 and Tucson International Airport, or from wells within treatment facilities.

Flexibility Matters: Cooperative Active Sites in Covalent Organic Framework and Threaded Ionic Polymer.

PubMed

Sun, Qi; Aguila, Briana; Perman, Jason; Nguyen, Nicholas; Ma, Shengqian

2016-12-07

The combination of two or more reactive centers working in concert on a substrate to facilitate the reaction is now considered state of the art in catalysis, yet there still remains a tremendous challenge. Few heterogeneous systems of this sort have been exploited, as the active sites spatially separated within the rigid framework are usually difficult to cooperate. It is now shown that this roadblock can be surpassed. The underlying principle of the strategy presented here is the integration of catalytic components with excellent flexibility and porous heterogeneous catalysts, as demonstrated by the placement of linear ionic polymers in close proximity to surface Lewis acid active sites anchored on the walls of a covalent organic framework (COF). Using the cycloaddition of the epoxides and CO 2 as a model reaction, dramatic activity improvements have been achieved for the composite catalysts in relation to the individual catalytic component. Furthermore, they also clearly outperform the benchmark catalytic systems formed by the combination of the molecular organocatalysts and heterogeneous Lewis acid catalysts, while affording additional recyclability. The extraordinary flexibility and enriched concentration of the catalytically active moieties on linear polymers facilitate the concerted catalysis, thus leading to superior catalytic performance. This work therefore uncovers an entirely new strategy for designing bifunctional catalysts with double-activation behavior and opens a new avenue in the design of multicapable systems that mimic biocatalysis.

Relationships between environmental organochlorine contaminant residues, plasma corticosterone concentrations, and intermediary metabolic enzyme activities in Great Lakes herring gull embryos.

PubMed Central

Lorenzen, A; Moon, T W; Kennedy, S W; Glen, G A

1999-01-01

Experiments were conducted to survey and detect differences in plasma corticosterone concentrations and intermediary metabolic enzyme activities in herring gull (Larus argentatus) embryos environmentally exposed to organochlorine contaminants in ovo. Unincubated fertile herring gull eggs were collected from an Atlantic coast control site and various Great Lakes sites in 1997 and artificially incubated in the laboratory. Liver and/or kidney tissues from approximately half of the late-stage embryos were analyzed for the activities of various intermediary metabolic enzymes known to be regulated, at least in part, by corticosteroids. Basal plasma corticosterone concentrations were determined for the remaining embryos. Yolk sacs were collected from each embryo and a subset was analyzed for organochlorine contaminants. Regression analysis of individual yolk sac organochlorine residue concentrations, or 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (TEQs), with individual basal plasma corticosterone concentrations indicated statistically significant inverse relationships for polychlorinated dibenzo-p-dioxins/polychlorinated dibenzofurans (PCDDs/PCDFs), total polychlorinated biphenyls (PCBs), non-ortho PCBs, and TEQs. Similarly, inverse relationships were observed for the activities of two intermediary metabolic enzymes (phosphoenolpyruvate carboxykinase and malic enzyme) when regressed against PCDDs/PCDFs. Overall, these data suggest that current levels of organochlorine contamination may be affecting the hypothalamo-pituitary-adrenal axis and associated intermediary metabolic pathways in environmentally exposed herring gull embryos in the Great Lakes. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:10064546

10 CFR 63.16 - Review of site characterization activities. 2

Code of Federal Regulations, 2012 CFR

2012-01-01

... which such activities are carried out and to observe excavations, borings, and in situ tests, as they... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of site characterization activities. 2 2 In addition to the review of site characterization activities...

10 CFR 63.16 - Review of site characterization activities. 2

Code of Federal Regulations, 2011 CFR

2011-01-01

... which such activities are carried out and to observe excavations, borings, and in situ tests, as they... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of site characterization activities. 2 2 In addition to the review of site characterization activities...

10 CFR 63.16 - Review of site characterization activities. 2

Code of Federal Regulations, 2013 CFR

2013-01-01

... which such activities are carried out and to observe excavations, borings, and in situ tests, as they... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of site characterization activities. 2 2 In addition to the review of site characterization activities...

10 CFR 63.16 - Review of site characterization activities. 2

Code of Federal Regulations, 2010 CFR

2010-01-01

... which such activities are carried out and to observe excavations, borings, and in situ tests, as they... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of site characterization activities. 2 2 In addition to the review of site characterization activities...

10 CFR 63.16 - Review of site characterization activities. 2

Code of Federal Regulations, 2014 CFR

2014-01-01

... which such activities are carried out and to observe excavations, borings, and in situ tests, as they... IN A GEOLOGIC REPOSITORY AT YUCCA MOUNTAIN, NEVADA Licenses Preapplication Review § 63.16 Review of site characterization activities. 2 2 In addition to the review of site characterization activities...

Physiological epidermal growth factor concentrations activate high affinity receptors to elicit calcium oscillations.

PubMed

Marquèze-Pouey, Béatrice; Mailfert, Sébastien; Rouger, Vincent; Goaillard, Jean-Marc; Marguet, Didier

2014-01-01

Signaling mediated by the epidermal growth factor (EGF) is crucial in tissue development, homeostasis and tumorigenesis. EGF is mitogenic at picomolar concentrations and is known to bind its receptor on high affinity binding sites depending of the oligomerization state of the receptor (monomer or dimer). In spite of these observations, the cellular response induced by EGF has been mainly characterized for nanomolar concentrations of the growth factor, and a clear definition of the cellular response to circulating (picomolar) concentrations is still lacking. We investigated Ca2+ signaling, an early event in EGF responses, in response to picomolar doses in COS-7 cells where the monomer/dimer equilibrium is unaltered by the synthesis of exogenous EGFR. Using the fluo5F Ca2+ indicator, we found that picomolar concentrations of EGF induced in 50% of the cells a robust oscillatory Ca2+ signal quantitatively similar to the Ca2+ signal induced by nanomolar concentrations. However, responses to nanomolar and picomolar concentrations differed in their underlying mechanisms as the picomolar EGF response involved essentially plasma membrane Ca2+ channels that are not activated by internal Ca2+ store depletion, while the nanomolar EGF response involved internal Ca2+ release. Moreover, while the picomolar EGF response was modulated by charybdotoxin-sensitive K+ channels, the nanomolar response was insensitive to the blockade of these ion channels.

Antiandrogenic activity of phthalate mixtures: Validity of concentration addition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Christen, Verena; Crettaz, Pierre; Oberli-Schrämmli, Aurelia

2012-03-01

Phthalates and bisphenol A have very widespread use leading to significant exposure of humans. They are suspected to interfere with the endocrine system, including the androgen, estrogen and the thyroid hormone system. Here we analyzed the antiandrogenic activity of six binary, and one ternary mixture of phthalates exhibiting complete antiandrogenic dose–response curves, and binary mixtures of phthalates and bisphenol A at equi-effective concentrations of EC{sub 10}, EC{sub 25} and EC{sub 50} in MDA-kb2 cells. Mixture activity followed the concentration addition (CA) model with a tendency to synergism at high and antagonism at low concentrations. Isoboles and the toxic unit approachmore » (TUA) confirmed the additive to synergistic activity of the binary mixtures BBP + DBP, DBP + DEP and DEP + BPA at high concentrations. Both methods indicate a tendency to antagonism for the EC{sub 10} mixtures BBP + DBP, BBP + DEP and DBP + DEP, and the EC{sub 25} mixture of DBP + BPA. A ternary mixture revealed synergism at the EC{sub 50}, and weak antagonistic activity at the EC{sub 25} level by the TUA. A mixture of five phthalates representing a human urine composition and reflecting exposure to corresponding parent compounds showed no antiandrogenic activity. Our study demonstrates that CA is an appropriate concept to account for mixture effects of antiandrogenic phthalates and bisphenol A. The interaction indicates a departure from additivity to antagonism at low concentrations, probably due to interaction with the androgen receptor and/or cofactors. This study emphasizes that a risk assessment of phthalates should account for mixture effects by applying the CA concept. -- Highlights: ► Antiandrogenic activity of mixtures of 2 and 3 phthalates are assessed in MDA-kb2 cells. ► Mixture activities followed the concentration addition model. ► A tendency to synergism at high and antagonism at low levels occurred.« less

Diurnal variations of airborne pollen concentration and the effect of ambient temperature in three sites of Mexico City

NASA Astrophysics Data System (ADS)

Ríos, B.; Torres-Jardón, R.; Ramírez-Arriaga, E.; Martínez-Bernal, A.; Rosas, I.

2016-05-01

Pollen is an important cause of allergic respiratory ailments in the Mexico City Metropolitan Area (MCMA). However, very little is known if ambient air temperature correlates with the early blooming of plants observed in other urban areas around the world. A research study was conducted during the dry season of 2012-2013 at three representative sites of the MCMA with different urban characteristics with the aim to understand the relationships between the profusion and diversity of pollen against temperature and other meteorological variables and degree of urbanization. Pollen samples were collected using a Hirst-type trap sampler in the sites: Merced (highly urbanized), Iztapalapa (medium-high urbanized) and Coyoacan (moderately urbanized). Urbanization levels were determined using a composite index based on population density, proportion of surface covered by construction and asphalt, and urban heat island intensity. A set of representative pollen sampling tapes were assayed under a light microscope at magnification of ×1,000 and converted to grains per cubic meter. The most representative pollen types found in the three sites were, regardless of urbanization levels were: Fraxinus, Cupressaceae/Taxodiaceae, Casuarina, Alnus, Myrtaceae, and Pinus. Total pollen concentration was greatest in the moderately urbanized area, although earlier blooming took place at the highly urbanized zone. Total pollen concentration in the medium-high urbanized site has the lowest because the green areas in this zone of MCMA are few. In a diurnal basis, the most abundant pollen types peaked near midday or in the afternoon evening at the three sites. A Spearman test showed a positive correlation among bihourly pollen concentrations, temperature and relative humidity in all sites, but wind speed just correlated in Iztapalapa and Coyoacan. The results obtained suggest that Urban Heat Island Intensity can disturb flowering periods and pollen concentrations, largely in the highly urbanized

Sublethal Effects of Imidacloprid on Honey Bee Colony Growth and Activity at Three Sites in the U.S.

PubMed

Meikle, William G; Adamczyk, John J; Weiss, Milagra; Gregorc, Ales; Johnson, Don R; Stewart, Scott D; Zawislak, Jon; Carroll, Mark J; Lorenz, Gus M

2016-01-01

Imidacloprid is a neonicotinoid pesticide heavily used by the agricultural industry and shown to have negative impacts on honey bees above certain concentrations. We evaluated the effects of different imidacloprid concentrations in sugar syrup using cage and field studies, and across different environments. Honey bee colonies fed sublethal concentrations of imidicloprid (0, 5, 20 and 100 ppb) over 6 weeks in field trials at a desert site (Arizona), a site near intensive agriculture (Arkansas) and a site with little nearby agriculture but abundant natural forage (Mississippi) were monitored with respect to colony metrics, such as adult bee and brood population sizes, as well as pesticide residues. Hive weight and internal hive temperature were monitored continuously over two trials in Arizona. Colonies fed 100 ppb imidacloprid in Arizona had significantly lower adult bee populations, brood surface areas and average frame weights, and reduced temperature control, compared to colonies in one or more of the other treatment groups, and consumption rates of those colonies were lower compared to other colonies in Arizona and Arkansas, although no differences in capped brood or average frame weight were observed among treatments in Arkansas. At the Mississippi site, also rich in alternative forage, colonies fed 5 ppb imidacloprid had less capped brood than control colonies, but contamination of control colonies was detected. In contrast, significantly higher daily hive weight variability among colonies fed 5 ppb imidacloprid in Arizona suggested greater foraging activity during a nectar flow post treatment, than any other treatment group. Imidacloprid concentrations in stored honey corresponded well with the respective syrup concentrations fed to the colonies and remained stable within the hive for at least 7 months after the end of treatment.

Computer simulation of the active site of human serum cholinesterase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kefang Jiao; Song Li; Zhengzheng Lu

1996-12-31

The first 3D-structure of acetylchelinesterase from Torpedo California electric organ (T.AChE) was published by JL. Sussman in 1991. We have simulated 3D-structure of human serum cholinesterase (H.BuChE) and the active site of H.BuChE. It is discovered by experiment that the residue of H.BuChE is still active site after a part of H.BuChE is cut. For example, the part of 21KD + 20KD is active site of H.BuChE. The 20KD as it is. Studies on these peptides by Hemelogy indicate that two active peptides have same negative electrostatic potential maps diagram. These negative electrostatic areas attached by acetyl choline with positivemore » electrostatic potency. We predict that 147...236 peptide of AChE could be active site because it was as 20KD as with negative electrostatic potential maps. We look forward to proving from other ones.« less

The potential for portable X-ray fluorescence determination of soil copper at ancient metallurgy sites, and considerations beyond measurements of total concentrations.

PubMed

Tighe, M; Rogan, G; Wilson, S C; Grave, P; Kealhofer, L; Yukongdi, P

2018-01-15

Copper (Cu) at ancient metallurgy sites represents the earliest instance of anthropogenically generated metal pollution. Such sites are spread across a wide range of environments from Eurasia to South America, and provide a unique opportunity to investigate the past and present extent and impact of metalworking contamination. Establishing the concentration and extent of soil Cu at archaeometallurgy sites can enhance archaeological interpretations of site use but can also, more fundamentally, provide an initial indication of contamination risk from such sites. Systematic evaluations of total soil Cu concentrations at ancient metalworking sites have not been conducted, due in part to the limitations of conventional laboratory-based protocols. In this paper, we first review what is known about Cu soil concentrations at ancient metallurgy sites. We then assess the benefits and challenges of portable X-ray fluorescence spectrometry (pXRF) as an alternative, rapid technique for the assessment of background and contaminant levels of Cu in soils. We conclude that pXRF is an effective tool for identifying potential contamination. Finally, we provide an overview of some major considerations beyond total Cu concentrations, such as bioavailability assessments, that will need to be considered at such sites to move toward a complete assessment of environmental and human risk. Copyright © 2017 Elsevier Ltd. All rights reserved.

Selecting optimal monitoring site locations for peak ambient particulate material concentrations using the MM5-CAMx4 numerical modelling system.

PubMed

Sturman, Andrew; Titov, Mikhail; Zawar-Reza, Peyman

2011-01-15

Installation of temporary or long term monitoring sites is expensive, so it is important to rationally identify potential locations that will achieve the requirements of regional air quality management strategies. A simple, but effective, numerical approach to selecting ambient particulate matter (PM) monitoring site locations has therefore been developed using the MM5-CAMx4 air pollution dispersion modelling system. A new method, 'site efficiency,' was developed to assess the ability of any monitoring site to provide peak ambient air pollution concentrations that are representative of the urban area. 'Site efficiency' varies from 0 to 100%, with the latter representing the most representative site location for monitoring peak PM concentrations. Four heavy pollution episodes in Christchurch (New Zealand) during winter 2005, representing 4 different aerosol dispersion patterns, were used to develop and test this site assessment technique. Evaluation of the efficiency of monitoring sites was undertaken for night and morning aerosol peaks for 4 different particulate material (PM) spatial patterns. The results demonstrate that the existing long term monitoring site at Coles Place is quite well located, with a site efficiency value of 57.8%. A temporary ambient PM monitoring site (operating during winter 2006) showed a lower ability to capture night and morning peak aerosol concentrations. Evaluation of multiple site locations used during an extensive field campaign in Christchurch (New Zealand) in 2000 indicated that the maximum efficiency achieved by any site in the city would be 60-65%, while the efficiency of a virtual background site is calculated to be about 7%. This method of assessing the appropriateness of any potential monitoring site can be used to optimize monitoring site locations for any air pollution measurement programme. Copyright © 2010 Elsevier B.V. All rights reserved.

De novo active sites for resurrected Precambrian enzymes

NASA Astrophysics Data System (ADS)

Risso, Valeria A.; Martinez-Rodriguez, Sergio; Candel, Adela M.; Krüger, Dennis M.; Pantoja-Uceda, David; Ortega-Muñoz, Mariano; Santoyo-Gonzalez, Francisco; Gaucher, Eric A.; Kamerlin, Shina C. L.; Bruix, Marta; Gavira, Jose A.; Sanchez-Ruiz, Jose M.

2017-07-01

Protein engineering studies often suggest the emergence of completely new enzyme functionalities to be highly improbable. However, enzymes likely catalysed many different reactions already in the last universal common ancestor. Mechanisms for the emergence of completely new active sites must therefore either plausibly exist or at least have existed at the primordial protein stage. Here, we use resurrected Precambrian proteins as scaffolds for protein engineering and demonstrate that a new active site can be generated through a single hydrophobic-to-ionizable amino acid replacement that generates a partially buried group with perturbed physico-chemical properties. We provide experimental and computational evidence that conformational flexibility can assist the emergence and subsequent evolution of new active sites by improving substrate and transition-state binding, through the sampling of many potentially productive conformations. Our results suggest a mechanism for the emergence of primordial enzymes and highlight the potential of ancestral reconstruction as a tool for protein engineering.

Natural and anthropogenic radionuclide activity concentrations in the New Zealand diet.

PubMed

Pearson, Andrew J; Gaw, Sally; Hermanspahn, Nikolaus; Glover, Chris N

2016-01-01

To support New Zealand's food safety monitoring regime, a survey was undertaken to establish radionuclide activity concentrations across the New Zealand diet. This survey was undertaken to better understand the radioactivity content of the modern diet and also to assess the suitability of the current use of milk as a sentinel for dietary radionuclide trends. Thirteen radionuclides were analysed in 40 common food commodities, including animal products, fruits, vegetables, cereal grains and seafood. Activity was detected for (137)Caesium, (90)Strontium and (131)Iodine. No other anthropogenic radionuclides were detected. Activity concentrations of the three natural radionuclides of Uranium and the daughter radionuclide (210)Polonium were detected in the majority of food sampled, with a large variation in magnitude. The maximum activity concentrations were detected in shellfish for all these radionuclides. Based on the established activity concentrations and ranges, the New Zealand diet contains activity concentrations of anthropogenic radionuclides far below the Codex Alimentarius guideline levels. Activity concentrations obtained for milk support its continued use as a sentinel for monitoring fallout radionuclides in terrestrial agriculture. The significant levels of natural and anthropogenic radionuclide activity concentrations detected in finfish and molluscs support undertaking further research to identify a suitable sentinel for New Zealand seafood monitoring. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Molecular dynamics explorations of active site structure in designed and evolved enzymes.

PubMed

Osuna, Sílvia; Jiménez-Osés, Gonzalo; Noey, Elizabeth L; Houk, K N

2015-04-21

This Account describes the use of molecular dynamics (MD) simulations to reveal how mutations alter the structure and organization of enzyme active sites. As proposed by Pauling about 70 years ago and elaborated by many others since then, biocatalysis is efficient when functional groups in the active site of an enzyme are in optimal positions for transition state stabilization. Changes in mechanism and covalent interactions are often critical parts of enzyme catalysis. We describe our explorations of the dynamical preorganization of active sites using MD, studying the fluctuations between active and inactive conformations normally concealed to static crystallography. MD shows how the various arrangements of active site residues influence the free energy of the transition state and relates the populations of the catalytic conformational ensemble to the enzyme activity. This Account is organized around three case studies from our laboratory. We first describe the importance of dynamics in evaluating a series of computationally designed and experimentally evolved enzymes for the Kemp elimination, a popular subject in the enzyme design field. We find that the dynamics of the active site is influenced not only by the original sequence design and subsequent mutations but also by the nature of the ligand present in the active site. In the second example, we show how microsecond MD has been used to uncover the role of remote mutations in the active site dynamics and catalysis of a transesterase, LovD. This enzyme was evolved by Tang at UCLA and Codexis, Inc., and is a useful commercial catalyst for the production of the drug simvastatin. X-ray analysis of inactive and active mutants did not reveal differences in the active sites, but relatively long time scale MD in solution showed that the active site of the wild-type enzyme preorganizes only upon binding of the acyl carrier protein (ACP) that delivers the natural acyl group to the active site. In the absence of bound ACP

Potential anti-cholinesterase and β-site amyloid precursor protein cleaving enzyme 1 inhibitory activities of cornuside and gallotannins from Cornus officinalis fruits.

PubMed

Bhakta, Himanshu Kumar; Park, Chan Hum; Yokozawa, Takako; Tanaka, Takashi; Jung, Hyun Ah; Choi, Jae Sue

2017-07-01

Cholinesterase (ChE) and β-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitors are promising agents for the treatment of Alzheimer's disease (AD). In the present study, we examined the inhibitory activity of seven compounds isolated from the fruits of Cornus officinalis, cornuside, polymeric proanthocyanidins, 1,2,3-tri-O-galloyl-β-D-glucose, 1,2,3,6-tetra-O-galloyl-β-D-glucose, tellimagrandin I, tellimagrandin II, and isoterchebin, against acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and BACE1. All of the compounds displayed concentration-dependent in vitro inhibitory activity toward the ChEs and BACE1. Among them, tellimagrandin II exhibited the best inhibitory activity toward ChEs, whereas the best BACE1 inhibitor was 1,2,3,6-tetra-O-galloyl-β-D-glucose. Isoterchebin and polymeric proanthocyanidins were also significant ChE inhibitors. The kinetic and docking studies demonstrated that all compounds interacted with both the catalytic active sites and the peripheral anionic sites of the ChEs and BACE1. Tellimagrandin II, isoterchebin, and the polymeric proanthocyanidins exhibited concentration-dependent inhibition of peroxynitrite-mediated protein tyrosine nitration. In conclusion, we identified significant ChE and BACE1 inhibitors from Corni Fructus that could have value as new multi-targeted compounds for anti-AD agents.

Increased concentrations of polycyclic aromatic hydrocarbons in Alpine streams during annual snowmelt: investigating effects of sampling method, site characteristics, and meteorology.

PubMed

Shahpoury, Pourya; Hageman, Kimberly J; Matthaei, Christoph D; Alumbaugh, Robert E; Cook, Michelle E

2014-10-07

Silicone passive samplers and macroinvertebrates were used to measure time-integrated concentrations of polycyclic aromatic hydrocarbons (PAHs) in alpine streams during annual snowmelt. The three sampling sites were located near a main highway in Arthur's Pass National Park in the Southern Alps of New Zealand. A similar set of PAH congeners, composed of 2-4 rings, were found in silicone passive samplers and macroinvertebrates. The background PAH concentrations were similar at all sites, implying that proximity to the highway did not affect concentrations. In passive samplers, an increase of PAH concentrations by up to seven times was observed during snowmelt. In macroinvertebrates, the concentration changes were moderate; however, macroinvertebrate sampling did not occur during the main pulse observed in the passive samplers. The extent of vegetation in the catchment appeared to affect the concentration patterns seen at the different stream sites. A strong correlation was found between PAH concentrations in passive samplers and the amount of rainfall in the study area, indicating that the washout of contaminants from snowpack by rainfall was an important process.

Regression models for estimating salinity and selenium concentrations at selected sites in the Upper Colorado River Basin, Colorado, 2009-2012

USGS Publications Warehouse

Linard, Joshua I.; Schaffrath, Keelin R.

2014-01-01

Elevated concentrations of salinity and selenium in the tributaries and main-stem reaches of the Colorado River are a water-quality concern and have been the focus of remediation efforts for many years. Land-management practices with the objective of limiting the amount of salt and selenium that reaches the stream have focused on improving the methods by which irrigation water is conveyed and distributed. Federal land managers implement improvements in accordance with the Colorado River Basin Salinity Control Act of 1974, which directs Federal land managers to enhance and protect the quality of water available in the Colorado River. In an effort to assist in evaluating and mitigating the detrimental effects of salinity and selenium, the U.S. Geological Survey, in cooperation with the Bureau of Reclamation, the Colorado River Water Resources District, and the Bureau of Land Management, analyzed salinity and selenium data collected at sites to develop regression models. The study area and sites are on the Colorado River or in one of three small basins in Western Colorado: the White River Basin, the Lower Gunnison River Basin, and the Dolores River Basin. By using data collected from water years 2009 through 2011, regression models able to estimate concentrations were developed for salinity at six sites and selenium at six sites. At a minimum, data from discrete measurement of salinity or selenium concentration, streamflow, and specific conductance at each of the sites were needed for model development. Comparison of the Adjusted R2 and standard error statistics of the two salinity models developed at each site indicated the models using specific conductance as the explanatory variable performed better than those using streamflow. The addition of multiple explanatory variables improved the ability to estimate selenium concentration at several sites compared with use of solely streamflow or specific conductance. The error associated with the log-transformed salinity

Relocating the Active-Site Lysine in Rhodopsin: 2. Evolutionary Intermediates.

PubMed

Devine, Erin L; Theobald, Douglas L; Oprian, Daniel D

2016-08-30

The visual pigment rhodopsin is a G protein-coupled receptor that covalently binds its retinal chromophore via a Schiff base linkage to an active-site Lys residue in the seventh transmembrane helix. Although this residue is strictly conserved among all type II retinylidene proteins, we found previously that the active-site Lys in bovine rhodopsin (Lys296) can be moved to three other locations (G90K, T94K, S186K) while retaining the ability to form a pigment with retinal and to activate transducin in a light-dependent manner [ Devine et al. ( 2013 ) Proc. Natl. Acad. Sci. USA 110 , 13351 - 13355 ]. Because the active-site Lys is not functionally constrained to be in helix seven, it is possible that it could relocate within the protein, most likely via an evolutionary intermediate with two active-site Lys. Therefore, in this study we characterized potential evolutionary intermediates with two Lys in the active site. Four mutant rhodopsins were prepared in which the original Lys296 was left untouched and a second Lys residue was substituted for G90K, T94K, S186K, or F293K. All four constructs covalently bind 11-cis-retinal, form a pigment, and activate transducin in a light-dependent manner. These results demonstrate that rhodopsin can tolerate a second Lys in the retinal binding pocket and suggest that an evolutionary intermediate with two Lys could allow migration of the Schiff base Lys to a position other than the observed, highly conserved location in the seventh TM helix. From sequence-based searches, we identified two groups of natural opsins, insect UV cones and neuropsins, that contain Lys residues at two positions in their active sites and also have intriguing spectral similarities to the mutant rhodopsins studied here.

Allosteric site-mediated active site inhibition of PBP2a using Quercetin 3-O-rutinoside and its combination.

PubMed

Rani, Nidhi; Vijayakumar, Saravanan; P T V, Lakshmi; Arunachalam, Annamalai

2016-08-01

Recent crystallographic study revealed the involvement of allosteric site in active site inhibition of penicillin binding protein (PBP2a), where one molecule of Ceftaroline (Cef) binds to the allosteric site of PBP2a and paved way for the other molecule (Cef) to bind at the active site. Though Cef has the potency to inhibit the PBP2a, its adverse side effects are of major concern. Previous studies have reported the antibacterial property of Quercetin derivatives, a group of natural compounds. Hence, the present study aims to evaluate the effect of Quercetin 3-o-rutinoside (Rut) in allosteric site-mediated active site inhibition of PBP2a. The molecular docking studies between allosteric site and ligands (Rut, Que, and Cef) revealed a better binding efficiency (G-score) of Rut (-7.790318) and Cef (-6.194946) with respect to Que (-5.079284). Molecular dynamic (MD) simulation studies showed significant changes at the active site in the presence of ligands (Rut and Cef) at allosteric site. Four different combinations of Rut and Cef were docked and their G-scores ranged between -6.320 and -8.623. MD studies revealed the stability of the key residue (Ser403) with Rut being at both sites, compared to other complexes. Morphological analysis through electron microscopy confirmed that combination of Rut and Cefixime was able to disturb the bacterial cell membrane in a similar fashion to that of Rut and Cefixime alone. The results of this study indicate that the affinity of Rut at both sites were equally good, with further validations Rut could be considered as an alternative for inhibiting MRSA growth.

Influence of active site location on catalytic activity in de novo-designed zinc metalloenzymes.

PubMed

Zastrow, Melissa L; Pecoraro, Vincent L

2013-04-17

While metalloprotein design has now yielded a number of successful metal-bound and even catalytically active constructs, the question of where to put a metal site along a linear, repetitive sequence has not been thoroughly addressed. Often several possibilities in a given sequence may exist that would appear equivalent but may in fact differ for metal affinity, substrate access, or protein dynamics. We present a systematic variation of active site location for a hydrolytically active ZnHis3O site contained within a de novo-designed three-stranded coiled coil. We find that the maximal rate, substrate access, and metal-binding affinity are dependent on the selected position, while catalytic efficiency for p-nitrophenyl acetate hydrolysis can be retained regardless of the location of the active site. This achievement demonstrates how efficient, tailor-made enzymes which control rate, pKa, substrate and solvent access (and selectivity), and metal-binding affinity may be realized. These findings may be applied to the more advanced de novo design of constructs containing secondary interactions, such as hydrogen-bonding channels. We are now confident that changes to location for accommodating such channels can be achieved without location-dependent loss of catalytic efficiency. These findings bring us closer to our ultimate goal of incorporating the secondary interactions we believe will be necessary in order to improve both active site properties and the catalytic efficiency to be competitive with the native enzyme, carbonic anhydrase.

Measurements of radon activity concentration in mouse tissues and organs.

PubMed

Ishimori, Yuu; Tanaka, Hiroshi; Sakoda, Akihiro; Kataoka, Takahiro; Yamaoka, Kiyonori; Mitsunobu, Fumihiro

2017-05-01

The purpose of this study is to investigate the biokinetics of inhaled radon, radon activity concentrations in mouse tissues and organs were determined after mice had been exposed to about 1 MBq/m 3 of radon in air. Radon activity concentrations in mouse blood and in other tissues and organs were measured with a liquid scintillation counter and with a well-type HP Ge detector, respectively. Radon activity concentration in mouse blood was 0.410 ± 0.016 Bq/g when saturated with 1 MBq/m 3 of radon activity concentration in air. In addition, average partition coefficients obtained were 0.74 ± 0.19 for liver, 0.46 ± 0.13 for muscle, 9.09 ± 0.49 for adipose tissue, and 0.22 ± 0.04 for other organs. With these results, a value of 0.414 for the blood-to-air partition coefficient was calculated by means of our physiologically based pharmacokinetic model. The time variation of radon activity concentration in mouse blood during exposure to radon was also calculated. All results are compared in detail with those found in the literature.

Cadmium, Chromium, and Copper Concentration plus Semen-Quality in Environmental Pollution Site, China

PubMed Central

LI, Yan; GAO, Qiaoyan; LI, Mingcai; LI, Mengyang; GAO, Xueming

2014-01-01

Abstract Background The environmental pollution is one of the factors contributing to the decrease of sperm quality for human beings. The aim of this study was to assess cadmium (Cd), chromium (Cr), and copper (Cu) concentration of man in environmental pollution site, and explore relationships between men exposure to Cd, Cr, and Cu and semen-quality parameters in environmental pollution site. Methods Ninety five men were recruited through pollution area and controls in 2011. We measured semen quality using Computer-aided Semen Quality Analysis, and Cd, Cr, and Cu levels in seminal plasma using Graphite Gurnace Atomic Absorption Spectroscopy. Spearman rank correlation analysis was used to evaluate the correlation between Cd, Cr and Cu concentration in seminal plasma and semen quality. Results The mean of seminal plasma Cd, Cr, and Cu values in pollution area was higher than the controls. Seminal plasma Cr values displayed a significant negative correlation with total motility and normomorph sperm rate. Seminal plasma Cu values also displayed a negative correlation with normomorph sperm rate. Conclusions Male reproductive health may be threatened by environmental pollution, and it may be influence local population diathesis. PMID:26060677

Active Sites Environmental Monitoring Program: Mid-FY 1991 report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ashwood, T.L.; Wickliff, D.S.; Morrissey, C.M.

1991-10-01

This report summarizes the activities of the Active Sites Environmental Monitoring Program (ASEMP) from October 1990 through March 1991. The ASEMP was established in 1989 by Solid Waste Operations and the Environmental Sciences Division to provide early detection and performance monitoring at active low-level radioactive waste (LLW) disposal sites in Solid Waste Storage Area (SWSA) 6 and transuranic (TRU) waste storage sites in SWSA 5 as required by chapters II and III of US Department of Energy Order 5820.2A. Monitoring results continue to demonstrate the no LLW is being leached from the storage vaults on the tumulus pads. Loading ofmore » vaults on Tumulus II began during this reporting period and 115 vaults had been loaded by the end of March 1991.« less

Sublethal Effects of Imidacloprid on Honey Bee Colony Growth and Activity at Three Sites in the U.S.

PubMed Central

Adamczyk, John J.; Weiss, Milagra; Gregorc, Ales; Johnson, Don R.; Stewart, Scott D.; Zawislak, Jon; Carroll, Mark J.; Lorenz, Gus M.

2016-01-01

Imidacloprid is a neonicotinoid pesticide heavily used by the agricultural industry and shown to have negative impacts on honey bees above certain concentrations. We evaluated the effects of different imidacloprid concentrations in sugar syrup using cage and field studies, and across different environments. Honey bee colonies fed sublethal concentrations of imidicloprid (0, 5, 20 and 100 ppb) over 6 weeks in field trials at a desert site (Arizona), a site near intensive agriculture (Arkansas) and a site with little nearby agriculture but abundant natural forage (Mississippi) were monitored with respect to colony metrics, such as adult bee and brood population sizes, as well as pesticide residues. Hive weight and internal hive temperature were monitored continuously over two trials in Arizona. Colonies fed 100 ppb imidacloprid in Arizona had significantly lower adult bee populations, brood surface areas and average frame weights, and reduced temperature control, compared to colonies in one or more of the other treatment groups, and consumption rates of those colonies were lower compared to other colonies in Arizona and Arkansas, although no differences in capped brood or average frame weight were observed among treatments in Arkansas. At the Mississippi site, also rich in alternative forage, colonies fed 5 ppb imidacloprid had less capped brood than control colonies, but contamination of control colonies was detected. In contrast, significantly higher daily hive weight variability among colonies fed 5 ppb imidacloprid in Arizona suggested greater foraging activity during a nectar flow post treatment, than any other treatment group. Imidacloprid concentrations in stored honey corresponded well with the respective syrup concentrations fed to the colonies and remained stable within the hive for at least 7 months after the end of treatment. PMID:28030617

Temporal changes in nitrogen and phosphorus concentrations with comparisons to conservation practices and agricultural activities in the Lower Grand River, Missouri and Iowa, and selected watersheds, 1969–2015

USGS Publications Warehouse

Krempa, Heather M.; Flickinger, Allison K.

2017-08-01

This report presents the results of a cooperative study by the U.S. Geological Survey and Missouri Department of Natural Resources to estimate total nitrogen (TN) and total phosphorus (TP) concentrations at monitoring sites within and near the Lower Grand River hydrological unit. The primary objectives of the study were to quantify temporal changes in TN and TP concentrations and compare those concentrations to conservation practices and agricultural activities. Despite increases in funding during 2011–15 for conservation practices in the Lower Grand River from the Mississippi River Basin Healthy Watersheds Initiative, decreases in flow-normalized TN and TP concentrations during this time at the long-term Grand River site were less than at other long-term sites, which did not receive funding from the Mississippi River Basin Healthy Watersheds Initiative. The relative differences in the magnitude of flow-normalized TN and TP concentrations among long-term sites are directly related to the amount of agricultural land use within the watershed. Significant relations were determined between nitrogen from cattle manure and flow-normalized TN concentrations at selected long-term sites, indicating livestock manure may be a substantial source of nitrogen within the selected long-term site watersheds. Relations between flow-normalized TN and TP concentrations with Conservation Reserve Program acres and with nitrogen and phosphorus from commercial fertilizer indicate that changes in these factors alone did not have a substantial effect on stream TN and TP concentrations; other landscape activities, runoff, within-bank nutrients that are suspended during higher streamflows, or a combination of these have had a greater effect on stream TN and TP concentrations; or there is a lag time that is obscuring relations. Temporal changes in flow-adjusted TN and TP concentrations were not substantial at Lower Grand River Mississippi River Basin Healthy Watersheds Initiative sites

Requirement of histidine 217 for ubiquinone reductase activity (Qi site) in the cytochrome bc1 complex.

PubMed

Gray, K A; Dutton, P L; Daldal, F

1994-01-25

Folding models suggest that the highly conserved histidine 217 of the cytochrome b subunit from the cytochrome bc1 complex is close to the quinone reductase (Qi) site. This histidine (bH217) in the cytochrome b polypeptide of the photosynthetic bacterium Rhodobacter capsulatus has been replaced with three other residues, aspartate (D), arginine (R), and leucine (L). bH217D and bH217R are able to grow photoheterotrophically and contain active cytochrome bc1 complexes (60% of wild-type activity), whereas the bH217L mutant is photosynthetically incompetent and contains a cytochrome bc1 complex that has only 10% of the wild-type activity. Single-turnover flash-activated electron transfer experiments show that cytochrome bH is reduced via the Qo site with near native rates in the mutant strains but that electron transfer between cytochrome bH and quinone bound at the Qi site is greatly slowed. These results are consistent with redox midpoint potential (Em) measurements of the cytochrome b subunit hemes and the Qi site quinone. The Em values of cyt bL and bH are approximately the same in the mutants and wild type, although the mutant strains have a larger relative concentration of what may be the high-potential form of cytochrome bH, called cytochrome b150. However, the redox properties of the semiquinone at the Qi site are altered significantly. The Qi site semiquinone stability constant of bH217R is 10 times higher than in the wild type, while in the other two strains (bH217D and bH217L) the stability constant is much lower than in the wild type. Thus H217 appears to have major effects on the redox properties of the quinone bound at the Qi site. These data are incorporated into a suggestion that H217 forms part of the binding pocket of the Qi site in a manner reminiscent of the interaction between quinone bound at the Qb site and H190 of the L subunit of the bacterial photosynthetic reaction center.

Biotic, temporal and spatial variability of tritium concentrations in transpirate samples collected in the vicinity of a near-surface low-level nuclear waste disposal site and nearby research reactor.

PubMed

Twining, J R; Hughes, C E; Harrison, J J; Hankin, S; Crawford, J; Johansen, M; Dyer, L

2011-06-01

The results of a 21 month sampling program measuring tritium in tree transpirate with respect to local sources are reported. The aim was to assess the potential of tree transpirate to indicate the presence of sub-surface seepage plumes. Transpirate gathered from trees near low-level nuclear waste disposal trenches contained activity concentrations of (3)H that were significantly higher (up to ∼700 Bq L(-1)) than local background levels (0-10 Bq L(-1)). The effects of the waste source declined rapidly with distance to be at background levels within 10s of metres. A research reactor 1.6 km south of the site contributed significant (p < 0.01) local fallout (3)H but its influence did not reach as far as the disposal trenches. The elevated (3)H levels in transpirate were, however, substantially lower than groundwater concentrations measured across the site (ranging from 0 to 91% with a median of 2%). Temporal patterns of tree transpirate (3)H, together with local meteorological observations, indicate that soil water within the active root zones comprised a mixture of seepage and rainfall infiltration. The degree of mixing was variable given that the soil water activity concentrations were heterogeneous at a scale equivalent to the effective rooting volume of the trees. In addition, water taken up by roots was not well mixed within the trees. Based on correlation modelling, net rainfall less evaporation (a surrogate for infiltration) over a period of from 2 to 3 weeks prior to sampling seems to be the optimum predictor of transpirate (3)H variability for any sampled tree at this site. The results demonstrate successful use of (3)H in transpirate from trees to indicate the presence and general extent of sub-surface contamination at a low-level nuclear waste site. Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.

Variability in Saponin Content, Cancer Antiproliferative Activity and Physicochemical Properties of Concentrated Agave Sap.

PubMed

Santos-Zea, Liliana; Rosas-Pérez, Aratza Mireya; Leal-Díaz, Ana María; Gutiérrez-Uribe, Janet A

2016-08-01

Concentrated agave sap (CAS) has gained popularity as an unrefined sweetener. It is obtained by boiling "aguamiel" that contains phytochemicals with diverse bioactivities. Saponins have been the most widely studied agave phytochemicals due to their cancer antiproliferative effect but their concentration may vary due to maturity of the agave plant and collection site. In this study, 18 CAS samples produced in different states of Mexico were analyzed using multivariate methods to determine which physicochemical or phytochemical parameters were responsible for variation. Additionally, extracts with different saponin profiles were tested to determine possible correlations with antiproliferative activity. Total soluble solids, pH, and water activity were similar to those reported for other agave sweeteners. Antioxidant capacity of samples was correlated to browning index. Eleven steroidal saponins were found in CAS samples and they were the main source of variability. Magueyoside B, a kammogenin tetraglycoside, was the most abundant saponin in all samples. With respect to bioactivity, multivariate analysis indicated that magueyoside B and a gentrogenin tetraglycoside were compounds strongly related with bioactivity. CAS from Hidalgo, Puebla, and Veracruz had higher concentration of magueyoside B than from the other kamogenin tetraglycoside found in the samples from other Mexican states. These results could be used as a first approach to characterize and standardize CAS to validate the potential health benefits derived from its consumption. © 2016 Institute of Food Technologists®

Reduction of Urease Activity by Interaction with the Flap Covering the Active Site

PubMed Central

Macomber, Lee; Minkara, Mona S.; Hausinger, Robert P.; Merz, Kenneth M.

2015-01-01

With the increasing appreciation for the human microbiome coupled with the global rise of antibiotic resistant organisms, it is imperative that new methods be developed to specifically target pathogens. To that end, a novel computational approach was devised to identify compounds that reduce the activity of urease, a medically important enzyme of Helicobacter pylori, Proteus mirabilis, and many other microorganisms. Urease contains a flexible loop that covers its active site; Glide was used to identify small molecules predicted to lock this loop in an open conformation. These compounds were screened against the model urease from Klebsiella aerogenes and the natural products epigallocatechin and quercetin were shown to inhibit at low and high micromolar concentrations, respectively. These molecules exhibit a strong time-dependent inactivation of urease that was not due to their oxygen sensitivity. Rather, these compounds appear to inactivate urease by reacting with a specific Cys residue located on the flexible loop. Substitution of this cysteine by alanine in the C319A variant increased the urease resistance to both epigallocatechin and quercetin, as predicted by the computational studies. Protein dynamics are integral to the function of many enzymes; thus, identification of compounds that lock an enzyme into a single conformation presents a useful approach to define potential inhibitors. PMID:25594724

Antioxidant Activities of Functional Beverage Concentrates Containing Herbal Medicine Extracts.

PubMed

Park, Seon-Joo; Kim, Mi-Ok; Kim, Jung Hoan; Jeong, Sehyun; Kim, Min Hee; Yang, Su-Jin; Lee, Jongsung; Lee, Hae-Jeung

2017-03-01

This study investigated the antioxidant activity of functional beverage concentrates containing herbal medicine extracts (FBCH) using various antioxidant assays, such as 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radical scavenging activity, and reducing power assay. The total polyphenolic content of FBCH (81.45 mg/100 g) was higher than Ssanghwa tea (SHT, 37.56 mg/100 g). The antioxidant activities of FBCH showed 52.92% DPPH and 55.18% ABTS radical scavenging activities at 100 mg/mL, respectively. FBCH showed significantly higher antioxidant activities compared to the SHT (DPPH, 23.43%; ABTS, 22.21%; reducing power optical density; 0.23, P <0.05). In addition, intracellular reactive oxygen species generation significantly decreased in a concentration-dependent manner following FBCH treatment. These results suggest that the addition of herbal medicine extract contributes to the improved functionality of beverage concentrates.

Factors affecting 210Po and 210Pb activity concentrations in mussels and implications for environmental bio-monitoring programmes.

PubMed

Carvalho, Fernando P; Oliveira, João M; Alberto, G

2011-02-01

The activity of (210)Po and (210)Pb was determined in mussels of the same size (3.5-4.0 cm shell length) sampled monthly over a 17-month period at the Atlantic coast of Portugal. Average radionuclide concentration values in mussels were 759±277 Bq kg(-1) for (210)Po (range 460-1470 Bq kg(-1) dry weight), and 45±19 Bq kg(-1) for (210)Pb (range 23-96 Bq kg(-1) dry weight). Environmental parameters and mussel biometric parameters were monitored during the same period. Although there was no seasonal variation of radionuclide concentrations in sea water during the study period, the concentration of radionuclide activity in mussels varied seasonally displaying peaks of high concentrations in winter and low concentrations in summer. Analysis of radionuclide data in relation to the physiological Condition Index of mussels revealed that (210)Po and (210)Pb activities in the mussel (average activity per individual) remained nearly constant during the investigation period, while mussel body weight fluctuated due to fat storage/expenditure in the soft tissues. Similar variation of radionuclide concentrations was observed in mussels transplanted from the sea coast into the Tejo Estuary. However, under estuarine environmental conditions and with higher food availability throughout the year, transplanted mussel Condition Index was higher than in coastal mussels and average radionuclide concentrations were 210±75 Bq kg(-1) (dry weight) for (210)Po and 10±4 Bq kg(-1) (dry weight) for (210)Pb, therefore lower than in coastal mussels with similar shell length. It is concluded that the apparent seasonal fluctuation and inter-site difference of radionuclide concentrations were mostly caused by mussel body weight fluctuation and not by radionuclide body burden fluctuation. This interpretation can be extended to the apparent seasonal fluctuation in concentrations of lipophilic and lipophobic contaminants in mussels, and provides an explanation for occasional high concentrations of

All the catalytic active sites of MoS 2 for hydrogen evolution

DOE PAGES

Li, Guoqing; Zhang, Du; Qiao, Qiao; ...

2016-11-29

MoS 2 presents a promising low-cost catalyst for the hydrogen evolution reaction (HER), but the understanding about its active sites has remained limited. Here we present an unambiguous study of the catalytic activities of all possible reaction sites of MoS 2, including edge sites, sulfur vacancies, and grain boundaries. We demonstrate that, in addition to the well-known catalytically active edge sites, sulfur vacancies provide another major active site for the HER, while the catalytic activity of grain boundaries is much weaker. Here, the intrinsic turnover frequencies (Tafel slopes) of the edge sites, sulfur vacancies, and grain boundaries are estimated tomore » be 7.5 s –1 (65–75 mV/dec), 3.2 s –1 (65–85 mV/dec), and 0.1 s –1 (120–160 mV/dec), respectively. We also demonstrate that the catalytic activity of sulfur vacancies strongly depends on the density of the vacancies and the local crystalline structure in proximity to the vacancies. Unlike edge sites, whose catalytic activity linearly depends on the length, sulfur vacancies show optimal catalytic activities when the vacancy density is in the range of 7–10%, and the number of sulfur vacancies in high crystalline quality MoS 2 is higher than that in low crystalline quality MoS 2, which may be related with the proximity of different local crystalline structures to the vacancies.« less

Machine-Learning Methods Enable Exhaustive Searches for Active Bimetallic Facets and Reveal Active Site Motifs for CO 2 Reduction

DOE PAGES

Ulissi, Zachary W.; Tang, Michael T.; Xiao, Jianping; ...

2017-07-27

Bimetallic catalysts are promising for the most difficult thermal and electrochemical reactions, but modeling the many diverse active sites on polycrystalline samples is an open challenge. Here, we present a general framework for addressing this complexity in a systematic and predictive fashion. Active sites for every stable low-index facet of a bimetallic crystal are enumerated and cataloged, yielding hundreds of possible active sites. The activity of these sites is explored in parallel using a neural-network-based surrogate model to share information between the many density functional theory (DFT) relaxations, resulting in activity estimates with an order of magnitude fewer explicit DFTmore » calculations. Sites with interesting activity were found and provide targets for follow-up calculations. This process was applied to the electrochemical reduction of CO 2 on nickel gallium bimetallics and indicated that most facets had similar activity to Ni surfaces, but a few exposed Ni sites with a very favorable on-top CO configuration. This motif emerged naturally from the predictive modeling and represents a class of intermetallic CO 2 reduction catalysts. These sites rationalize recent experimental reports of nickel gallium activity and why previous materials screens missed this exciting material. Most importantly these methods suggest that bimetallic catalysts will be discovered by studying facet reactivity and diversity of active sites more systematically.« less

Machine-Learning Methods Enable Exhaustive Searches for Active Bimetallic Facets and Reveal Active Site Motifs for CO 2 Reduction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ulissi, Zachary W.; Tang, Michael T.; Xiao, Jianping

Bimetallic catalysts are promising for the most difficult thermal and electrochemical reactions, but modeling the many diverse active sites on polycrystalline samples is an open challenge. Here, we present a general framework for addressing this complexity in a systematic and predictive fashion. Active sites for every stable low-index facet of a bimetallic crystal are enumerated and cataloged, yielding hundreds of possible active sites. The activity of these sites is explored in parallel using a neural-network-based surrogate model to share information between the many density functional theory (DFT) relaxations, resulting in activity estimates with an order of magnitude fewer explicit DFTmore » calculations. Sites with interesting activity were found and provide targets for follow-up calculations. This process was applied to the electrochemical reduction of CO 2 on nickel gallium bimetallics and indicated that most facets had similar activity to Ni surfaces, but a few exposed Ni sites with a very favorable on-top CO configuration. This motif emerged naturally from the predictive modeling and represents a class of intermetallic CO 2 reduction catalysts. These sites rationalize recent experimental reports of nickel gallium activity and why previous materials screens missed this exciting material. Most importantly these methods suggest that bimetallic catalysts will be discovered by studying facet reactivity and diversity of active sites more systematically.« less

Mutation at a strictly conserved, active site tyrosine in the copper amine oxidase leads to uncontrolled oxygenase activity.

PubMed

Chen, Zhi-Wei; Datta, Saumen; Dubois, Jennifer L; Klinman, Judith P; Mathews, F Scott

2010-08-31

The copper amine oxidases carry out two copper-dependent processes: production of their own redox-active cofactor (2,4,5-trihydroxyphenylalanine quinone, TPQ) and the subsequent oxidative deamination of substrate amines. Because the same active site pocket must facilitate both reactions, individual active site residues may serve multiple roles. We have examined the roles of a strictly conserved active site tyrosine Y305 in the copper amine oxidase from Hansenula polymorpha kinetically, spetroscopically (Dubois and Klinman (2006) Biochemistry 45, 3178), and, in the present work, structurally. While the Y305A enzyme is almost identical to the wild type, a novel, highly oxygenated species replaces TPQ in the Y305F active sites. This new structure not only provides the first direct detection of peroxy intermediates in cofactor biogenesis but also indicates the critical control of oxidation chemistry that can be conferred by a single active site residue.

137Cs activity concentration in mosses in the Calabria region, south of Italy

NASA Astrophysics Data System (ADS)

Caridi, F.; Belvedere, A.; D'Agostino, M.; Marguccio, S.

2017-05-01

Many studies carried out after the Chernobyl nuclear accident in 1986 showed that mosses are suitable bioindicators of the radioactive fall-out, given their long life expectancy. 137Cs activity concentration was measured through HPGe gamma spectrometry in different mosses (Hypnum Cupressiforme, Hypnum Andoi, Homalothecium Sericeum, Philonotis Fontana), collected, in 2008 and 2015, from four sampling sites in the Calabria region, south of Italy. Experimental specific activities allowed us to determine the effective and the biological half-time of 137Cs for the investigated plants. The effective half-time was found in the range from (4.2 ± 1.1) to (7.4 ± 1.8) years; the biological half-time varied from (4.9 ± 1.3) to (9.9 ± 2.4) years. Data reported in this article provide useful information on the environmental risk of the studied area and can be further used for a radiological mapping.

Concentrations of polycyclic aromatic hydrocarbons: Their potential health risks and sources at three non-urban sites in Japan.

PubMed

Jadoon, Waqar A; Sakugawa, Hiroshi

2016-09-18

This investigation was undertaken to monitor particulate polycyclic aromatic hydrocarbons (PAHs) concentrations in order to determine their emission sources and potential human health risks in remote and rural areas of Japan. Seventeen PAHs in aerosol samples collected in remote (Kamihaya), coastal (Hiki) and inland (Higashi-Hiroshima) areas of Japan during 2013-2014 were analyzed using gas chromatography/mass spectrometry. Total PAH (Σ17PAH) concentrations in aerosol samples were in the range of 0.08-6.51 ng m(-3), 0.09-4.74 ng m(-3), and 0.21-6.53 ng m(-3) at Kamihaya, Hiki, and Higashi-Hiroshima sites, with mean concentrations of 1.63, 1.18, and 2.43 ng m(-3), respectively. Significant seasonal variation in concentrations occurred at Hiki and Higashi-Hiroshima, while no significant variation occurred at Kamihaya. Ambient air temperature greatly affected PAH concentrations in Higashi-Hiroshima, but had only moderate effects in Kamihaya and Hiki. Wind direction also influenced the concentrations of PAHs. Vehicle exhaust, industrial emissions, biomass combustion, and domestic heating and cooking were identified as the main PAH emission sources using principal component analysis. Backward trajectory calculations showed that domestically generated PAHs were significant in Kamihaya and Hiki, while in Higashi-Hiroshima concentrations were mainly influenced by long-range transport. The incremental lifetime lung cancer risk had values of 3.38 × 10(-5) and 1.84 × 10(-5) at Higashi-Hiroshima and Hiki, which are greater than the US EPA acceptable level (10(-6)). Typically, 5-6-ring PAHs contributed 95% to this overall health risk, of which benzo(a)pyrene was the largest contributor, followed by dibenz(a,h)anthracene at both residential sites. Clearly, stricter guidelines for PAHs need to be implemented at these sites to protect the population.

Functional Evolution of PLP-dependent Enzymes based on Active-Site Structural Similarities

PubMed Central

Catazaro, Jonathan; Caprez, Adam; Guru, Ashu; Swanson, David; Powers, Robert

2014-01-01

Families of distantly related proteins typically have very low sequence identity, which hinders evolutionary analysis and functional annotation. Slowly evolving features of proteins, such as an active site, are therefore valuable for annotating putative and distantly related proteins. To date, a complete evolutionary analysis of the functional relationship of an entire enzyme family based on active-site structural similarities has not yet been undertaken. Pyridoxal-5’-phosphate (PLP) dependent enzymes are primordial enzymes that diversified in the last universal ancestor. Using the Comparison of Protein Active Site Structures (CPASS) software and database, we show that the active site structures of PLP-dependent enzymes can be used to infer evolutionary relationships based on functional similarity. The enzymes successfully clustered together based on substrate specificity, function, and three-dimensional fold. This study demonstrates the value of using active site structures for functional evolutionary analysis and the effectiveness of CPASS. PMID:24920327

Chapter 12: Daily Patterns of Marbled Murrelet Activity at Inland Sites

Treesearch

Nancy L. Naslund; Brian P. O’Donnell

1995-01-01

Patterns in the daily activity of Marbled Murrelets (Brachyramphus marmoratus) at inland sites has been studied throughout their range from California to Alaska. Murrelets are most active at inland sites around dawn, and to a lesser degree, at dusk. Throughout their range, peak levels of activity (detections) occur in the hour around dawn, but...

An Active Site Water Network in the Plasminogen Activator Pla from Yersinia pestis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eren, Elif; Murphy, Megan; Goguen, Jon

2010-08-13

The plasminogen activator Pla from Yersinia pestis is an outer membrane protease (omptin) that is important for the virulence of plague. Here, we present the high-resolution crystal structure of wild-type, enzymatically active Pla at 1.9 {angstrom}. The structure shows a water molecule located between active site residues D84 and H208, which likely corresponds to the nucleophilic water. A number of other water molecules are present in the active site, linking residues important for enzymatic activity. The R211 sidechain in loop L4 is close to the nucleophilic water and possibly involved in the stabilization of the oxyanion intermediate. Subtle conformational changesmore » of H208 result from the binding of lipopolysaccharide to the outside of the barrel, explaining the unusual dependence of omptins on lipopolysaccharide for activity. The Pla structure suggests a model for the interaction with plasminogen substrate and provides a more detailed understanding of the catalytic mechanism of omptin proteases.« less

Ligand-binding specificity and promiscuity of the main lignocellulolytic enzyme families as revealed by active-site architecture analysis.

PubMed

Tian, Li; Liu, Shijia; Wang, Shuai; Wang, Lushan

2016-03-24

Biomass can be converted into sugars by a series of lignocellulolytic enzymes, which belong to the glycoside hydrolase (GH) families summarized in CAZy databases. Here, using a structural bioinformatics method, we analyzed the active site architecture of the main lignocellulolytic enzyme families. The aromatic amino acids Trp/Tyr and polar amino acids Glu/Asp/Asn/Gln/Arg occurred at higher frequencies in the active site architecture than in the whole enzyme structure. And the number of potential subsites was significantly different among different families. In the cellulase and xylanase families, the conserved amino acids in the active site architecture were mostly found at the -2 to +1 subsites, while in β-glucosidase they were mainly concentrated at the -1 subsite. Families with more conserved binding amino acid residues displayed strong selectivity for their ligands, while those with fewer conserved binding amino acid residues often exhibited promiscuity when recognizing ligands. Enzymes with different activities also tended to bind different hydroxyl oxygen atoms on the ligand. These results may help us to better understand the common and unique structural bases of enzyme-ligand recognition from different families and provide a theoretical basis for the functional evolution and rational design of major lignocellulolytic enzymes.

Anisotropic Covalency Contributions to Superexchange Pathways in Type One Copper Active Sites

PubMed Central

2015-01-01

Type one (T1) Cu sites deliver electrons to catalytic Cu active sites: the mononuclear type two (T2) Cu site in nitrite reductases (NiRs) and the trinuclear Cu cluster in the multicopper oxidases (MCOs). The T1 Cu and the remote catalytic sites are connected via a Cys-His intramolecular electron-transfer (ET) bridge, which contains two potential ET pathways: P1 through the protein backbone and P2 through the H-bond between the Cys and the His. The high covalency of the T1 Cu–S(Cys) bond is shown here to activate the T1 Cu site for hole superexchange via occupied valence orbitals of the bridge. This covalency-activated electronic coupling (HDA) facilitates long-range ET through both pathways. These pathways can be selectively activated depending on the geometric and electronic structure of the T1 Cu site and thus the anisotropic covalency of the T1 Cu–S(Cys) bond. In NiRs, blue (π-type) T1 sites utilize P1 and green (σ-type) T1 sites utilize P2, with P2 being more efficient. Comparing the MCOs to NiRs, the second-sphere environment changes the conformation of the Cys-His pathway, which selectively activates HDA for superexchange by blue π sites for efficient turnover in catalysis. These studies show that a given protein bridge, here Cys-His, provides different superexchange pathways and electronic couplings depending on the anisotropic covalencies of the donor and acceptor metal sites. PMID:25310460

Functional evolution of PLP-dependent enzymes based on active-site structural similarities.

PubMed

Catazaro, Jonathan; Caprez, Adam; Guru, Ashu; Swanson, David; Powers, Robert

2014-10-01

Families of distantly related proteins typically have very low sequence identity, which hinders evolutionary analysis and functional annotation. Slowly evolving features of proteins, such as an active site, are therefore valuable for annotating putative and distantly related proteins. To date, a complete evolutionary analysis of the functional relationship of an entire enzyme family based on active-site structural similarities has not yet been undertaken. Pyridoxal-5'-phosphate (PLP) dependent enzymes are primordial enzymes that diversified in the last universal ancestor. Using the comparison of protein active site structures (CPASS) software and database, we show that the active site structures of PLP-dependent enzymes can be used to infer evolutionary relationships based on functional similarity. The enzymes successfully clustered together based on substrate specificity, function, and three-dimensional-fold. This study demonstrates the value of using active site structures for functional evolutionary analysis and the effectiveness of CPASS. © 2014 Wiley Periodicals, Inc.

Sensitivity to experimental data of pollutant site mean concentration in stormwater runoff.

PubMed

Mourad, M; Bertrand-Krajewski, J L; Chebbo, G

2005-01-01

Urban wet weather discharges are known to be a great source of pollutants for receiving waters, which protection requires the estimation of long-term discharged pollutant loads. Pollutant loads can be estimated by multiplying a site mean concentration (SMC) by the total runoff volume during a given period of time. The estimation of the SMC value as a weighted mean value with event runoff volumes as weights is affected by uncertainties due to the variability of event mean concentrations and to the number of events used. This study carried out on 13 catchments gives orders of magnitude of these uncertainties and shows the limitations of usual practices using few measured events. The results obtained show that it is not possible to propose a standard minimal number of events to be measured on any catchment in order to evaluate the SMC value with a given uncertainty.

Athletic Activity and Hormone Concentrations in High School Female Athletes

PubMed Central

Wojtys, Edward M.; Jannausch, Mary L.; Kreinbrink, Jennifer L.; Harlow, Siobán D.; Sowers, MaryFran R.

2015-01-01

Context: Physical activity may affect the concentrations of circulating endogenous hormones in female athletes. Understanding the relationship between athletic and physical activity and circulating female hormone concentrations is critical. Objective: To test the hypotheses that (1) the estradiol-progesterone profile of high school adolescent girls participating in training, conditioning, and competition would differ from that of physically inactive, age-matched adolescent girls throughout a 3-month period; and (2) athletic training and conditioning would alter body composition (muscle, bone), leading to an increasingly greater lean–body-mass to fat–body-mass ratio with accompanying hormonal changes. Design: Cohort study. Settings: Laboratory and participants' homes. Patients or Other Participants: A total of 106 adolescent girls, ages 14–18 years, who had experienced at least 3 menstrual cycles in their lifetime. Main Outcome Measure(s): Participants were prospectively monitored throughout a 13-week period, with weekly physical activity assessments and 15 urine samples for estrogen, luteinizing hormone, creatinine, and progesterone concentrations. Each girl underwent body-composition measurements before and after the study period. Results: Seventy-four of the 98 girls (76%) who completed the study classified themselves as athletes. Body mass index, body mass, and fat measures remained stable, and 17 teenagers had no complete menstrual cycle during the observation period. Mean concentrations of log(estrogen/creatinine) were slightly greater in nonathletes who had cycles of <24 or >35 days. Mean log(progesterone/creatinine) concentrations in nonathletes were less in the first half and greater in the second half of the cycle, but the differences were not statistically significant. Conclusions: A moderate level of athletic or physical activity did not influence urine concentrations of estrogen, progesterone, or luteinizing hormones. However, none of the

Oxysterol-binding Protein Activation at Endoplasmic Reticulum-Golgi Contact Sites Reorganizes Phosphatidylinositol 4-Phosphate Pools*

PubMed Central

Goto, Asako; Charman, Mark; Ridgway, Neale D.

2016-01-01

Oxysterol-binding protein (OSBP) exchanges cholesterol and phosphatidylinositol 4-phosphate (PI-4P) at contact sites between the endoplasmic reticulum (ER) and the trans-Golgi/trans-Golgi network. 25-Hydroxycholesterol (25OH) competitively inhibits this exchange reaction in vitro and causes the constitutive localization of OSBP at the ER/Golgi interface and PI-4P-dependent recruitment of ceramide transfer protein (CERT) for sphingomyelin synthesis. We used PI-4P probes and mass analysis to determine how OSBP controls the availability of PI-4P for this metabolic pathway. Treatment of fibroblasts or Chinese hamster ovary (CHO) cells with 25OH caused a 50–70% reduction in Golgi-associated immunoreactive PI-4P that correlated with Golgi localization of OSBP. In contrast, 25OH caused an OSBP-dependent enrichment in Golgi PI-4P that was detected with a pleckstrin homology domain probe. The cellular mass of phosphatidylinositol monophosphates and Golgi PI-4P measured with an unbiased PI-4P probe (P4M) was unaffected by 25OH and OSBP silencing, indicating that OSBP shifts the distribution of PI-4P upon localization to ER-Golgi contact sites. The PI-4P and sterol binding activities of OSBP were both required for 25OH activation of sphingomyelin synthesis, suggesting that 25OH must be exchanged for PI-4P to be concentrated at contact sites. We propose a model wherein 25OH activation of OSBP promotes the binding and retention of PI-4P at ER-Golgi contact sites. This pool of PI-4P specifically recruits pleckstrin homology domain-containing proteins involved in lipid transfer and metabolism, such as CERT. PMID:26601944

Active-site solvent replenishment observed during human carbonic anhydrase II catalysis.

PubMed

Kim, Jin Kyun; Lomelino, Carrie L; Avvaru, Balendu Sankara; Mahon, Brian P; McKenna, Robert; Park, SangYoun; Kim, Chae Un

2018-01-01

Human carbonic anhydrase II (hCA II) is a zinc metalloenzyme that catalyzes the reversible hydration/dehydration of CO 2 /HCO 3 - . Although hCA II has been extensively studied to investigate the proton-transfer process that occurs in the active site, its underlying mechanism is still not fully understood. Here, ultrahigh-resolution crystallographic structures of hCA II cryocooled under CO 2 pressures of 7.0 and 2.5 atm are presented. The structures reveal new intermediate solvent states of hCA II that provide crystallographic snapshots during the restoration of the proton-transfer water network in the active site. Specifically, a new intermediate water (W I ') is observed next to the previously observed intermediate water W I , and they are both stabilized by the five water molecules at the entrance to the active site (the entrance conduit). Based on these structures, a water network-restructuring mechanism is proposed, which takes place at the active site after the nucleophilic attack of OH - on CO 2 . This mechanism explains how the zinc-bound water (W Zn ) and W1 are replenished, which are directly responsible for the reconnection of the His64-mediated proton-transfer water network. This study provides the first 'physical' glimpse of how a water reservoir flows into the hCA II active site during its catalytic activity.

A concentration-independent micro/nanofluidic active diode using an asymmetric ion concentration polarization layer.

PubMed

Lee, Hyekyung; Kim, Junsuk; Kim, Hyeonsoo; Kim, Ho-Young; Lee, Hyomin; Kim, Sung Jae

2017-08-24

Over the past decade, nanofluidic diodes that rectify ionic currents (i.e. greater current in one direction than in the opposite direction) have drawn significant attention in biomolecular sensing, switching and energy harvesting devices. To obtain current rectification, conventional nanofluidic diodes have utilized complex nanoscale asymmetry such as nanochannel geometry, surface charge density, and reservoir concentration. Avoiding the use of sophisticated nano-asymmetry, micro/nanofluidic diodes using microscale asymmetry have been recently introduced; however, their diodic performance is still impeded by (i) low (even absent) rectification effects at physiological concentrations over 100 mM and strong dependency on the bulk concentration, and (ii) the fact that they possess only passive predefined rectification factors. Here, we demonstrated a new class of micro/nanofluidic diode with an ideal perm-selective nanoporous membrane based on ion concentration polarization (ICP) phenomenon. Thin side-microchannels installed near a nanojunction served as mitigators of the amplified electrokinetic flows generated by ICP and induced convective salt transfer to the nanoporous membrane, leading to actively controlled micro-scale asymmetry. Using this device, current rectifications were successfully demonstrated in a wide range of electrolytic concentrations (10 -5 M to 3 M) as a function of the fluidic resistance of the side-microchannels. Noteworthily, it was confirmed that the rectification factors were independent from the bulk concentration due to the ideal perm-selectivity. Moreover, the rectification of the presenting diode was actively controlled by adjusting the external convective flows, while that of the previous diode was passively determined by invariant nanoscale asymmetry.

Seasonal variations in activity concentrations of 99Tc and 137Cs in the edible meat fraction of crabs and lobsters from the central Irish Sea.

PubMed

Copplestone, D; Jackson, D; Hartnoll, R G; Johnson, M S; McDonald, P; Wood, N

2004-01-01

Discharges of most radionuclides into the Irish Sea from the BNFL site at Sellafield have decreased over the past 20 years or so. For a few radionuclides, however, discharges have peaked more recently. Notably, operation of the Enhanced Actinide Removal Plant (EARP) since 1994 has led to an increase in discharges of (99)Tc, as a result of the treatment of previously stored waste, with consequent increases in (99)Tc activity concentrations in a number of marine species, particularly in crustaceans such as lobsters. Previous research has considered the significance of factors such as sex and body weight on radionuclide concentrations. The current project set out to investigate whether seasonal variations in radionuclide concentrations in crabs and lobsters occur, with particular emphasis on the dynamics of (99)Tc and (137)Cs. Organisms were obtained from a site off the Isle of Man, where radionuclide concentrations were measurable but the site was sufficiently distant from Sellafield that the radionuclides were well mixed in the water column and not likely to be influenced by the pulsed nature of discharges of (99)Tc. Crab and lobster samples were collected monthly, between February 2000 and February 2001. Fifteen or 16 individuals (evenly split as male and female) of each species were collected on each occasion. Seawater samples were also collected over the 12-month period. Activity concentrations of (99)Tc in the edible meat fraction (both brown and white meat) ranged from 0.23 to 2.46 Bq kg(-1) (fresh weight (fw)) in crabs and 124 to 216 Bq kg(-1) (fw) in lobsters, with no observed seasonal variations. Activity concentrations of (137)Cs in both crab and lobster were lower, ranging from <0.16 to 0.85 Bq kg(-1) for crab meat (fw) and <0.3 to 3.3 Bq kg(-1) for lobster meat (fw). A statistically significant increase in activity concentrations of (137)Cs in the meat was observed in the summer months for both crab and lobster. The cause has not been investigated but may

Utilizing patch and site level greenhouse-gas concentration measurements in tandem with the prognostic model, ecosys

NASA Astrophysics Data System (ADS)

Morin, T. H.; Rey Sanchez, C.; Bohrer, G.; Riley, W. J.; Angle, J.; Mekonnen, Z. A.; Stefanik, K. C.; Wrighton, K. C.

2016-12-01

Estimates of wetland greenhouse gas (GHG) budgets currently have large uncertainties. While wetlands are the largest source of natural methane (CH4) emissions worldwide, they are also important carbon dioxide (CO2) sinks. Determining the GHG budget of a wetland is challenging, particularly because wetlands have intrinsically temporally and spatially heterogeneous land cover patterns and complex dynamics of CH4 production and emissions. These issues pose challenges to both measuring and modeling GHG budgets from wetlands. To improve wetland GHG flux predictability, we utilized the ecosys model to predict CH4 fluxes from a natural temperate estuarine wetland in northern Ohio. Multiple patches of terrain (that included Typha spp. and Nelumbo lutea) were represented as separate grid cells in the model. Cells were initialized with measured values but were allowed to dynamically evolve in response to meteorological, hydrological, and thermodynamic conditions. Trace gas surface emissions were predicted as the end result of microbial activity, physical transport, and plant processes. Corresponding to each model gridcell, measurements of dissolved gas concentrations were conducted with pore-water dialysis samplers (peepers). The peeper measurements were taken via a series of tubes, providing an undisturbed observation of the pore water concentrations of in situ dissolved gases along a vertical gradient. Non-steady state chambers and a flux tower provided both patch level and integrated site-level fluxes of CO2 and CH4. New Typha chambers were also developed to enclose entire plants and segregate the plant fluxes from soil/water fluxes. We expect ecosys to predict the seasonal and diurnal fluxes of CH4 from within each land cover type and to resolve where CH4 is generated within the soil column and its transmission mechanisms. We demonstrate the need for detailed information at both the patch and site level when using models to predict whole wetland ecosystem-scale GHG

Estimated fecal coliform bacteria concentrations using near real-time continuous water-quality and streamflow data from five stream sites in Chester County, Pennsylvania, 2007–16

USGS Publications Warehouse

Senior, Lisa A.

2017-09-15

Several streams used for recreational activities, such as fishing, swimming, and boating, in Chester County, Pennsylvania, are known to have periodic elevated concentrations of fecal coliform bacteria, a type of bacteria used to indicate the potential presence of fecally related pathogens that may pose health risks to humans exposed through water contact. The availability of near real-time continuous stream discharge, turbidity, and other water-quality data for some streams in the county presents an opportunity to use surrogates to estimate near real-time concentrations of fecal coliform (FC) bacteria and thus provide some information about associated potential health risks during recreational use of streams.The U.S. Geological Survey (USGS), in cooperation with the Chester County Health Department (CCHD) and the Chester County Water Resources Authority (CCWRA), has collected discrete stream samples for analysis of FC concentrations during March–October annually at or near five gaging stations where near real-time continuous data on stream discharge, turbidity, and water temperature have been collected since 2007 (or since 2012 at 2 of the 5 stations). In 2014, the USGS, in cooperation with the CCWRA and CCHD, began to develop regression equations to estimate FC concentrations using available near real-time continuous data. Regression equations included possible explanatory variables of stream discharge, turbidity, water temperature, and seasonal factors calculated using Julian Day with base-10 logarithmic (log) transformations of selected variables.The regression equations were developed using the data from 2007 to 2015 (101–106 discrete bacteria samples per site) for three gaging stations on Brandywine Creek (West Branch Brandywine Creek at Modena, East Branch Brandywine Creek below Downingtown, and Brandywine Creek at Chadds Ford) and from 2012 to 2015 (37–38 discrete bacteria samples per site) for one station each on French Creek near Phoenixville and

Relation of Nickel Concentrations in Tree Rings to Groundwater Contamination

NASA Astrophysics Data System (ADS)

Yanosky, Thomas M.; Vroblesky, Don A.

1992-08-01

Increment cores were collected from trees growing at two sites where groundwater is contaminated by nickel. Proton-induced X ray emission spectroscopy was used to determine the nickel concentrations in selected individual rings and in parts of individual rings. Ring nickel concentrations were interpreted on the basis of recent concentrations of nickel in aquifers, historical information about site use activities, and model simulations of groundwater flow. Nickel concentrations in rings increased during years of site use but not in trees outside the contaminated aquifers. Consequently, it was concluded that trees may preserve in their rings an annual record of nickel contamination in groundwater. Tulip trees and oaks contained higher concentrations of nickel than did sassafras, sweet gum, or black cherry. No evidence was found that nickel accumulates consistently within parts of individual rings or that nickel is translocated across ring boundaries.

Relation of nickel concentrations in tree rings to groundwater contamination

USGS Publications Warehouse

Yanosky, Thomas M.; Vroblesky, Don A.

1992-01-01

Increment cores were collected from trees growing at two sites where groundwater is contaminated by nickel. Proton-induced X ray emission spectroscopy was used to determine the nickel concentrations in selected individual rings and in parts of individual rings. Ring nickel concentrations were interpreted on the basis of recent concentrations of nickel in aquifers, historical information about site use activities, and model simulations of groundwater flow. Nickel concentrations in rings increased during years of site use but not in trees outside the contaminated aquifers. Consequently, it was concluded that trees may preserve in their rings an annual record of nickel contamination in groundwater. Tulip trees and oaks contained higher concentrations of nickel than did sassafras, sweet gum, or black cherry. No evidence was found that nickel accumulates consistently within parts of individual rings or that nickel is translocated across ring boundaries.

Mechanism and Catalytic Site Atlas (M-CSA): a database of enzyme reaction mechanisms and active sites.

PubMed

Ribeiro, António J M; Holliday, Gemma L; Furnham, Nicholas; Tyzack, Jonathan D; Ferris, Katherine; Thornton, Janet M

2018-01-04

M-CSA (Mechanism and Catalytic Site Atlas) is a database of enzyme active sites and reaction mechanisms that can be accessed at www.ebi.ac.uk/thornton-srv/m-csa. Our objectives with M-CSA are to provide an open data resource for the community to browse known enzyme reaction mechanisms and catalytic sites, and to use the dataset to understand enzyme function and evolution. M-CSA results from the merging of two existing databases, MACiE (Mechanism, Annotation and Classification in Enzymes), a database of enzyme mechanisms, and CSA (Catalytic Site Atlas), a database of catalytic sites of enzymes. We are releasing M-CSA as a new website and underlying database architecture. At the moment, M-CSA contains 961 entries, 423 of these with detailed mechanism information, and 538 with information on the catalytic site residues only. In total, these cover 81% (195/241) of third level EC numbers with a PDB structure, and 30% (840/2793) of fourth level EC numbers with a PDB structure, out of 6028 in total. By searching for close homologues, we are able to extend M-CSA coverage of PDB and UniProtKB to 51 993 structures and to over five million sequences, respectively, of which about 40% and 30% have a conserved active site. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Spatial distribution of and seasonal variations in endosulfan concentrations in soil, air, and biota around a contaminated site.

PubMed

Fang, Yanyan; Nie, Zhiqiang; Yang, Jinzhong; Die, Qingqi; Tian, Yajun; Liu, Feng; He, Jie; Wang, Jianyuan; Huang, Qifei

2018-06-12

Soil, air, tree bark, rice, wheat, invertebrates, and chicken tissues around a typical endosulfan-contaminated site were analyzed in each season in each of two years. The total endosulfan (the sum of α-, β-endosulfan and endosulfan sulfate) were significantly different in soil and air samples collected in the four seasons (P < 0.01) and the mean concentrations were 6.53 ng/g dry weight (d.w.) and 2.40 ng/m 3 , respectively, in autumn, 3.32 ng/g d.w. and 2.48 ng/m 3 , respectively, in winter, 2.10 ng/g d.w. and 0.93 ng/m 3 , respectively, in spring, and 1.03 ng/g d.w. and 0.83 ng/m 3 , respectively, in summer. The total endosulfan concentrations in tree bark, rice, wheat, and invertebrates were 23.0-278 (mean 95.5) ng/g d.w., 7.36-35.5 (mean 17.4) ng/g d.w., 34.3-158 (mean 83.1) ng/g d.w., and 401-4354 (mean 2125) ng/g lipid weight, respectively. The total endosulfan concentrations in the chicken gizzard, heart, liver, and meat samples were 552, 212, 699, and 221 ng/g lipid weight, respectively. The endosulfan concentrations in soil, air, and biota around the site were strongly influenced by endosulfan emissions from the site, and the concentrations had decreased to half the initial concentrations six months after endosulfan production stopped. The invertebrate and chicken bioconcentration and biomagnification factors indicated that endosulfan accumulated in the invertebrates and chicken tissues was slightly biomagnified by chickens. Copyright © 2018 Elsevier Inc. All rights reserved.

[Study on Tritium Content in Soil at Sites of Nuclear Explosions on the Territory of Semipalatinsk Test Site].

PubMed

Timonova, L V; Lyakhova, O N; Lukashenko, S N; Aidarkhanov, A O

2015-01-01

As a result of investigations carried out on the territory of Semipalatinsk Test Site, tritium was found in different environmental objects--surface and ground waters, vegetation, air environment, and snow cover. The analysis of the data obtained has shown that contamination of environmental objects at the Semipalatinsk Test Site with tritium is associated with the places where underground nuclear tests were performed. Since tritium can originate from an activation reaction and be trapped by pock particles during a test, it was decided to examine the soil in the sites where surface and excavation tests took place. It was found that the concentration of tritium in soil correlates with the concentration of europium. Probably, the concentration of tritium in the soil depends on the character and yield of the tests performed. Findings of the study have revealed that tritium can be found in soil in significant amounts not only in sites where underground nuclear tests took place but also in sites where surface and excavation nuclear tests were carried out.

Application of passive sampling for measuring dissolved concentrations of organic contaminants in the water column at three marine superfund sites

EPA Science Inventory

At contaminated sediment sites, including U.S. EPA Superfund sites, it is critical to measure water column concentrations of freely dissolved contaminants to understand the complete exposure of aquatic organisms to hydrophobic organic contaminants (HOCs). However, historically a...

Active Site Gate Dynamics Modulate the Catalytic Activity of the Ubiquitination Enzyme E2-25K.

PubMed

Rout, Manoj K; Lee, Brian L; Lin, Aiyang; Xiao, Wei; Spyracopoulos, Leo

2018-05-03

The ubiquitin proteasome system (UPS) signals for degradation of proteins through attachment of K48-linked polyubiquitin chains, or alterations in protein-protein recognition through attachment of K63-linked chains. Target proteins are ubiquitinated in three sequential chemical steps by a three-component enzyme system. Ubiquitination, or E2 enzymes, catalyze the central step by facilitating reaction of a target protein lysine with the C-terminus of Ub that is attached to the active site cysteine of the E2 through a thioester bond. E2 reactivity is modulated by dynamics of an active site gate, whose central residue packs against the active site cysteine in a closed conformation. Interestingly, for the E2 Ubc13, which specifically catalyzes K63-linked ubiquitination, the central gate residue adopts an open conformation. We set out to determine if active site gate dynamics play a role in catalysis for E2-25K, which adopts the canonical, closed gate conformation, and which selectively synthesizes K48-linked ubiquitin chains. Gate dynamics were characterized using mutagenesis of key residues, combined with enzyme kinetics measurements, and main chain NMR relaxation. The experimental data were interpreted with all atom MD simulations. The data indicate that active site gate opening and closing rates for E2-25K are precisely balanced.

(/sup 3/H)forskolin- and (/sup 3/H)dihydroalprenolol-binding sites and adenylate cyclase activity in heart of rats fed diets containing different oils

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alam, S.Q.; Ren, Y.F.; Alam, B.S.

1988-03-01

The characteristics of the cardiac adenylate cyclase system were studied in rats fed diets containing fish oil (menhaden oil) and other oils. Adenylate cyclase activity generally was higher in cardiac homogenates and membranes of rats fed diet containing 10% menhaden oil than in the other oils. The increase in enzyme activity, especially in forskolin-stimulated activity, was associated with an increase in the concentration of the (/sup 3/H) forskolin-binding sites in cardiac membranes of rats fed menhaden oil. The beta-adrenergic receptor concentration was not significantly altered although the affinity for (/sup 3/H)dihydroalprenolol-binding was lower in membranes of rats fed menhaden oilmore » than those fed the other oils. omega-3 fatty acids from menhaden oil were incorporated into the cardiac membrane phospholipids. The results suggest that the observed increase in myocardial adenylate cyclase activity of rats fed menhaden oil may be due to an increase in the number of the catalytic subunits of the enzyme or due to a greater availability of the forskolin-binding sites.« less

Sedentary behavior, physical activity, and concentrations of insulin among US adults.

PubMed

Ford, Earl S; Li, Chaoyang; Zhao, Guixiang; Pearson, William S; Tsai, James; Churilla, James R

2010-09-01

Time spent watching television has been linked to obesity, metabolic syndrome, and diabetes, all conditions characterized to some degree by hyperinsulinemia and insulin resistance. However, limited evidence relates screen time (watching television or using a computer) directly to concentrations of insulin. We examined the cross-sectional associations between time spent watching television or using a computer, physical activity, and serum concentrations of insulin using data from 2800 participants aged at least 20 years of the 2003-2006 National Health and Nutrition Examination Survey. The amount of time spent watching television and using a computer as well as physical activity was self-reported. The unadjusted geometric mean concentration of insulin increased from 6.2 microU/mL among participants who did not watch television to 10.0 microU/mL among those who watched television for 5 or more hours per day (P = .001). After adjustment for age, sex, race or ethnicity, educational status, concentration of cotinine, alcohol intake, physical activity, waist circumference, and body mass index using multiple linear regression analysis, the log-transformed concentrations of insulin were significantly and positively associated with time spent watching television (P = < .001). Reported time spent using a computer was significantly associated with log-transformed concentrations of insulin before but not after accounting for waist circumference and body mass index. Leisure-time physical activity but not transportation or household physical activity was significantly and inversely associated with log-transformed concentrations of insulin. Sedentary behavior, particularly the amount of time spent watching television, may be an important modifiable determinant of concentrations of insulin. Published by Elsevier Inc.

SABER: A computational method for identifying active sites for new reactions

PubMed Central

Nosrati, Geoffrey R; Houk, K N

2012-01-01

A software suite, SABER (Selection of Active/Binding sites for Enzyme Redesign), has been developed for the analysis of atomic geometries in protein structures, using a geometric hashing algorithm (Barker and Thornton, Bioinformatics 2003;19:1644–1649). SABER is used to explore the Protein Data Bank (PDB) to locate proteins with a specific 3D arrangement of catalytic groups to identify active sites that might be redesigned to catalyze new reactions. As a proof-of-principle test, SABER was used to identify enzymes that have the same catalytic group arrangement present in o-succinyl benzoate synthase (OSBS). Among the highest-scoring scaffolds identified by the SABER search for enzymes with the same catalytic group arrangement as OSBS were l-Ala d/l-Glu epimerase (AEE) and muconate lactonizing enzyme II (MLE), both of which have been redesigned to become effective OSBS catalysts, demonstrated by experiments. Next, we used SABER to search for naturally existing active sites in the PDB with catalytic groups similar to those present in the designed Kemp elimination enzyme KE07. From over 2000 geometric matches to the KE07 active site, SABER identified 23 matches that corresponded to residues from known active sites. The best of these matches, with a 0.28 Å catalytic atom RMSD to KE07, was then redesigned to be compatible with the Kemp elimination using RosettaDesign. We also used SABER to search for potential Kemp eliminases using a theozyme predicted to provide a greater rate acceleration than the active site of KE07, and used Rosetta to create a design based on the proteins identified. PMID:22492397

SABER: a computational method for identifying active sites for new reactions.

PubMed

Nosrati, Geoffrey R; Houk, K N

2012-05-01

A software suite, SABER (Selection of Active/Binding sites for Enzyme Redesign), has been developed for the analysis of atomic geometries in protein structures, using a geometric hashing algorithm (Barker and Thornton, Bioinformatics 2003;19:1644-1649). SABER is used to explore the Protein Data Bank (PDB) to locate proteins with a specific 3D arrangement of catalytic groups to identify active sites that might be redesigned to catalyze new reactions. As a proof-of-principle test, SABER was used to identify enzymes that have the same catalytic group arrangement present in o-succinyl benzoate synthase (OSBS). Among the highest-scoring scaffolds identified by the SABER search for enzymes with the same catalytic group arrangement as OSBS were L-Ala D/L-Glu epimerase (AEE) and muconate lactonizing enzyme II (MLE), both of which have been redesigned to become effective OSBS catalysts, demonstrated by experiments. Next, we used SABER to search for naturally existing active sites in the PDB with catalytic groups similar to those present in the designed Kemp elimination enzyme KE07. From over 2000 geometric matches to the KE07 active site, SABER identified 23 matches that corresponded to residues from known active sites. The best of these matches, with a 0.28 Å catalytic atom RMSD to KE07, was then redesigned to be compatible with the Kemp elimination using RosettaDesign. We also used SABER to search for potential Kemp eliminases using a theozyme predicted to provide a greater rate acceleration than the active site of KE07, and used Rosetta to create a design based on the proteins identified. Copyright © 2012 The Protein Society.

Concentration of (137)Cs in soil across Nebraska.

PubMed

Weesner, Alexandra Palensky; Fairchild, Robert W

2008-06-01

Atmospheric nuclear weapons testing from 1945 through 1980 produced radioactive fallout that was transported by stratospheric winds and deposited unevenly around the world. The accident at Chernobyl in 1986 also contributed to the fallout in some locations. The (137)Cs activity concentration from fallout has been measured as a function of depth in soil samples from five different locations across Nebraska. Soil samples 2-cm thick down to a depth of 30 cm were collected in Brown, Dawes, Lancaster, Red Willow, and Thurston Counties. Samples taken from each of the sites were dried, sieved, and counted using an HPGe gamma spectroscopy system to measure the activity concentration of (137)Cs at each depth in the soil. Activity concentrations as high as 216 Bq kg(-1) were measured in the samples. Dry soil bulk densities were calculated for each site based on soil type and used to calculate the area density of deposition. Area deposition densities up to 13,100 Bq m(-2) were measured, consistent with published estimates.

Targeting smooth emergence: the effect site concentration of remifentanil for preventing cough during emergence during propofol-remifentanil anaesthesia for thyroid surgery.

PubMed

Lee, B; Lee, J-R; Na, S

2009-06-01

The administration of short-acting opioids can be a reliable and safe method to prevent coughing during emergence from anaesthesia but the proper dose or effect site concentration of remifentanil for this purpose has not been reported. We therefore investigated the effect site concentration (Ce) of remifentanil for preventing cough during emergence from anaesthesia with propofol-remifentanil target-controlled infusion. Twenty-three ASA I-II grade female patients, aged 23-66 yr undergoing elective thyroidectomy were enrolled in this study. EC(50) and EC(95) of remifentanil for preventing cough were determined using Dixon's up-and-down method and probit analysis. Propofol effect site concentration at extubation, mean arterial pressure, and heart rate (HR) were compared in patients with smooth emergence and without smooth emergence. Three out of 11 patients with remifentanil Ce of 1.5 ng ml(-1) and all seven patients with Ce of 2.0 ng ml(-1) did not cough during emergence; the EC(50) of remifentanil that suppressed coughing was 1.46 ng ml(-1) by Dixon's up-and-down method, and EC(95) was 2.14 ng ml(-1) by probit analysis. Effect site concentration of propofol at awakening was similar in patients with a smooth emergence and those without smooth emergence, but HR and arterial pressure were higher in those who coughed during emergence. Clinically significant hypoventilation was not seen in any patient. We found that the EC(95) of effect site concentration of remifentanil to suppress coughing at emergence from anaesthesia was 2.14 ng ml(-1). Maintaining an established Ce of remifentanil is a reliable method of abolishing cough and thereby targeting smooth emergence from anaesthesia.

Deepest and hottest hydrothermal activity in the Okinawa Trough: the Yokosuka site at Yaeyama Knoll

PubMed Central

Miyazaki, Junichi; Kawagucci, Shinsuke; Makabe, Akiko; Takahashi, Ayu; Kitada, Kazuya; Torimoto, Junji; Matsui, Yohei; Tasumi, Eiji; Shibuya, Takazo; Nakamura, Kentaro; Horai, Shunsuke; Sato, Shun; Ishibashi, Jun-ichiro; Kanzaki, Hayato; Nakagawa, Satoshi; Hirai, Miho; Takaki, Yoshihiro; Okino, Kyoko; Watanabe, Hiromi Kayama; Kumagai, Hidenori

2017-01-01

Since the initial discovery of hydrothermal vents in 1977, these ‘extreme’ chemosynthetic systems have been a focus of interdisciplinary research. The Okinawa Trough (OT), located in the semi-enclosed East China Sea between the Eurasian continent and the Ryukyu arc, hosts more than 20 known vent sites but all within a relatively narrow depth range (600–1880 m). Depth is a significant factor in determining fluid temperature and chemistry, as well as biological composition. However, due to the narrow depth range of known sites, the actual influence of depth here has been poorly resolved. Here, the Yokosuka site (2190 m), the first OT vent exceeding 2000 m depth is reported. A highly active hydrothermal vent site centred around four active vent chimneys reaching 364°C in temperature, it is the hottest in the OT. Notable Cl depletion (130 mM) and both high H2 and CH4 concentrations (approx. 10 mM) probably result from subcritical phase separation and thermal decomposition of sedimentary organic matter. Microbiota and fauna were generally similar to other sites in the OT, although with some different characteristics. In terms of microbiota, the H2-rich vent fluids in Neuschwanstein chimney resulted in the dominance of hydrogenotrophic chemolithoautotrophs such as Thioreductor and Desulfobacterium. For fauna, the dominance of the deep-sea mussel Bathymodiolus aduloides is surprising given other nearby vent sites are usually dominated by B. platifrons and/or B. japonicus, and a sponge field in the periphery dominated by Poecilosclerida is unusual for OT vents. Our insights from the Yokosuka site implies that although the distribution of animal species may be linked to depth, the constraint is perhaps not water pressure and resulting chemical properties of the vent fluid but instead physical properties of the surrounding seawater. The potential significance of these preliminary results and prospect for future research on this unique site are discussed

Deepest and hottest hydrothermal activity in the Okinawa Trough: the Yokosuka site at Yaeyama Knoll

NASA Astrophysics Data System (ADS)

Miyazaki, Junichi; Kawagucci, Shinsuke; Makabe, Akiko; Takahashi, Ayu; Kitada, Kazuya; Torimoto, Junji; Matsui, Yohei; Tasumi, Eiji; Shibuya, Takazo; Nakamura, Kentaro; Horai, Shunsuke; Sato, Shun; Ishibashi, Jun-ichiro; Kanzaki, Hayato; Nakagawa, Satoshi; Hirai, Miho; Takaki, Yoshihiro; Okino, Kyoko; Watanabe, Hiromi Kayama; Kumagai, Hidenori; Chen, Chong

2017-12-01

Since the initial discovery of hydrothermal vents in 1977, these `extreme' chemosynthetic systems have been a focus of interdisciplinary research. The Okinawa Trough (OT), located in the semi-enclosed East China Sea between the Eurasian continent and the Ryukyu arc, hosts more than 20 known vent sites but all within a relatively narrow depth range (600-1880 m). Depth is a significant factor in determining fluid temperature and chemistry, as well as biological composition. However, due to the narrow depth range of known sites, the actual influence of depth here has been poorly resolved. Here, the Yokosuka site (2190 m), the first OT vent exceeding 2000 m depth is reported. A highly active hydrothermal vent site centred around four active vent chimneys reaching 364°C in temperature, it is the hottest in the OT. Notable Cl depletion (130 mM) and both high H2 and CH4 concentrations (approx. 10 mM) probably result from subcritical phase separation and thermal decomposition of sedimentary organic matter. Microbiota and fauna were generally similar to other sites in the OT, although with some different characteristics. In terms of microbiota, the H2-rich vent fluids in Neuschwanstein chimney resulted in the dominance of hydrogenotrophic chemolithoautotrophs such as Thioreductor and Desulfobacterium. For fauna, the dominance of the deep-sea mussel Bathymodiolus aduloides is surprising given other nearby vent sites are usually dominated by B. platifrons and/or B. japonicus, and a sponge field in the periphery dominated by Poecilosclerida is unusual for OT vents. Our insights from the Yokosuka site implies that although the distribution of animal species may be linked to depth, the constraint is perhaps not water pressure and resulting chemical properties of the vent fluid but instead physical properties of the surrounding seawater. The potential significance of these preliminary results and prospect for future research on this unique site are discussed.

'Unconventional' coordination chemistry by metal chelating fragments in a metalloprotein active site.

PubMed

Martin, David P; Blachly, Patrick G; Marts, Amy R; Woodruff, Tessa M; de Oliveira, César A F; McCammon, J Andrew; Tierney, David L; Cohen, Seth M

2014-04-09

The binding of three closely related chelators: 5-hydroxy-2-methyl-4H-pyran-4-thione (allothiomaltol, ATM), 3-hydroxy-2-methyl-4H-pyran-4-thione (thiomaltol, TM), and 3-hydroxy-4H-pyran-4-thione (thiopyromeconic acid, TPMA) to the active site of human carbonic anhydrase II (hCAII) has been investigated. Two of these ligands display a monodentate mode of coordination to the active site Zn(2+) ion in hCAII that is not recapitulated in model complexes of the enzyme active site. This unprecedented binding mode in the hCAII-thiomaltol complex has been characterized by both X-ray crystallography and X-ray spectroscopy. In addition, the steric restrictions of the active site force the ligands into a 'flattened' mode of coordination compared with inorganic model complexes. This change in geometry has been shown by density functional computations to significantly decrease the strength of the metal-ligand binding. Collectively, these data demonstrate that the mode of binding by small metal-binding groups can be significantly influenced by the protein active site. Diminishing the strength of the metal-ligand bond results in unconventional modes of metal coordination not found in typical coordination compounds or even carefully engineered active site models, and understanding these effects is critical to the rational design of inhibitors that target clinically relevant metalloproteins.

Enhanced enzyme kinetic stability by increasing rigidity within the active site.

PubMed

Xie, Yuan; An, Jiao; Yang, Guangyu; Wu, Geng; Zhang, Yong; Cui, Li; Feng, Yan

2014-03-14

Enzyme stability is an important issue for protein engineers. Understanding how rigidity in the active site affects protein kinetic stability will provide new insight into enzyme stabilization. In this study, we demonstrated enhanced kinetic stability of Candida antarctica lipase B (CalB) by mutating the structurally flexible residues within the active site. Six residues within 10 Å of the catalytic Ser(105) residue with a high B factor were selected for iterative saturation mutagenesis. After screening 2200 colonies, we obtained the D223G/L278M mutant, which exhibited a 13-fold increase in half-life at 48 °C and a 12 °C higher T50(15), the temperature at which enzyme activity is reduced to 50% after a 15-min heat treatment. Further characterization showed that global unfolding resistance against both thermal and chemical denaturation also improved. Analysis of the crystal structures of wild-type CalB and the D223G/L278M mutant revealed that the latter formed an extra main chain hydrogen bond network with seven structurally coupled residues within the flexible α10 helix that are primarily involved in forming the active site. Further investigation of the relative B factor profile and molecular dynamics simulation confirmed that the enhanced rigidity decreased fluctuation of the active site residues at high temperature. These results indicate that enhancing the rigidity of the flexible segment within the active site may provide an efficient method for improving enzyme kinetic stability.

GASS-WEB: a web server for identifying enzyme active sites based on genetic algorithms.

PubMed

Moraes, João P A; Pappa, Gisele L; Pires, Douglas E V; Izidoro, Sandro C

2017-07-03

Enzyme active sites are important and conserved functional regions of proteins whose identification can be an invaluable step toward protein function prediction. Most of the existing methods for this task are based on active site similarity and present limitations including performing only exact matches on template residues, template size restraints, despite not being capable of finding inter-domain active sites. To fill this gap, we proposed GASS-WEB, a user-friendly web server that uses GASS (Genetic Active Site Search), a method based on an evolutionary algorithm to search for similar active sites in proteins. GASS-WEB can be used under two different scenarios: (i) given a protein of interest, to match a set of specific active site templates; or (ii) given an active site template, looking for it in a database of protein structures. The method has shown to be very effective on a range of experiments and was able to correctly identify >90% of the catalogued active sites from the Catalytic Site Atlas. It also managed to achieve a Matthew correlation coefficient of 0.63 using the Critical Assessment of protein Structure Prediction (CASP 10) dataset. In our analysis, GASS was ranking fourth among 18 methods. GASS-WEB is freely available at http://gass.unifei.edu.br/. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Stabilization of Glucocerebrosidase by Active Site Occupancy

PubMed Central

2017-01-01

Glucocerebrosidase (GBA) is a lysosomal β-glucosidase that degrades glucosylceramide. Its deficiency results in Gaucher disease (GD). We examined the effects of active site occupancy of GBA on its structural stability. For this, we made use of cyclophellitol-derived activity-based probes (ABPs) that bind irreversibly to the catalytic nucleophile (E340), and for comparison, we used the potent reversible inhibitor isofagomine. We demonstrate that cyclophellitol ABPs improve the stability of GBA in vitro, as revealed by thermodynamic measurements (Tm increase by 21 °C), and introduce resistance to tryptic digestion. The stabilizing effect of cell-permeable cyclophellitol ABPs is also observed in intact cultured cells containing wild-type GBA, N370S GBA (labile in lysosomes), and L444P GBA (exhibits impaired ER folding): all show marked increases in lysosomal forms of GBA molecules upon exposure to ABPs. The same stabilization effect is observed for endogenous GBA in the liver of wild-type mice injected with cyclophellitol ABPs. Stabilization effects similar to those observed with ABPs were also noted at high concentrations of the reversible inhibitor isofagomine. In conclusion, we provide evidence that the increase in cellular levels of GBA by ABPs and by the reversible inhibitor is in part caused by their ability to stabilize GBA folding, which increases the resistance of GBA against breakdown by lysosomal proteases. These effects are more pronounced in the case of the amphiphilic ABPs, presumably due to their high lipophilic potential, which may promote further structural compactness of GBA through hydrophobic interactions. Our study provides further rationale for the design of chaperones for GBA to ameliorate Gaucher disease. PMID:28485919

A tale of two sites: On defining the carrier concentration in garnet-based ionic conductors for advanced Li batteries

DOE PAGES

Thompson, Travis; Sharafi, Asma; Johannes, Michelle D.; ...

2015-03-21

Solid electrolytes based on the garnet crystal structure have recently been identified as a promising material to enable advance Li battery cell chemistries because of the unprecedented combination of high ionic conductivity and electrochemical stability against metallic Li. To better understand the mechanisms that give rise to high conductivity, the goal of this work is to correlate Li site occupancy with Li-ion transport. Toward this goal, the Li site occupancy is studied in cubic garnet as a function of Li concentration over the compositions range: Li 7-xLa 3Zr 2-xTa xO 12 (x = 0.5, 0.75, and 1.5). The distribution ofmore » Li between the two interstitial sites (24d and 96h) is determined using neutron and synchrotron diffraction. The bulk conductivity is measured on >97% relative density polycrystalline specimens to correlate Li-ion transport as a function of Li site occupancy. It is determined that the conductivity changes nonlinearly with the occupancy of the octahedral (96h) Li site. It is shown that the effective carrier concentration is dependent on the Li site occupancy and suggests that this is a consequence of significant carrier-carrier coulombic interactions. Moreover, the observation of maximum conductivity near Li = 6.5 mol is explained.« less

Comparability of suspended-sediment concentration and total suspended-solids data for two sites on the L'Anguille River, Arkansas, 2001 to 2003

USGS Publications Warehouse

Galloway, Joel M.; Evans, Dennis A.; Green, W. Reed

2005-01-01

Suspended-sediment concentration and total suspended solids data collected with automatic pumping samplers at the L'Anguille River near Colt and the L'Anguille River at Palestine, Arkansas, August 2001 to October 2003 were compared using ordinary least squares regression analyses to determine the relation between the two datasets for each of the two sites. The purpose of this report is to describe the suspended-sediment concentration and total suspended-solids data and examine the comparability of the two datasets for each site. Suspended-sediment concentration and total suspended solids data for the L'Anguille River varied spatially and temporally from August 2001 to October 2003. The site at the L'Anguille River at Palestine represents a larger portion of the L'Anguille River Basin than the site near Colt, and generally had higher median suspended-sediment concentration and total suspended solids and greater ranges in values. The differences between suspended-sediment concentration and total suspended solids data for the L'Anguille River near Colt appeared inversely related to streamflow and not related to time. The relation between suspended-sediment concentration and total suspended solids at the L'Anguille River at Palestine was more variable than at Colt and did not appear to have a relation with flow or time. The relation between suspended-sediment concentration and total suspended solids for the L'Anguille River near Colt shows that total suspended solids increased proportionally as suspended-sediment concentration increased. However, the relation between suspended-sediment concentration and total suspended solids for the L'Anguille River at Palestine showed total suspended solids increased less proportionally as suspended-sediment concentration increased compared to the L'Anguille River near Colt. Differences between the two analytical methods may partially explain differences between the suspended-sediment concentration and total suspended solids data at

Comparison of sources of submicron particle number concentrations measured at two sites in Rochester, NY.

PubMed

Kasumba, John; Hopke, Philip K; Chalupa, David C; Utell, Mark J

2009-09-01

Sources contributing to the submicron particles (100-470 nm) measured between January 2002 and December 2007 at two different New York State Department of Environmental Conservation (NYS DEC) sites in Rochester, NY were identified and apportioned using a bilinear receptor model, positive matrix factorization (PMF). Measurements of aerosol size distributions and number concentrations for particles in the size range of 10-500 nm have been made since December 2001 to date in Rochester. The measurements are being made using a scanning mobility particle sizer (SMPS) consisting of a DMA and a CPC (TSI models 3071 and 3010, respectively). From December 2001 to March 2004, particle measurements were made at the NYS DEC site in downtown Rochester, but it was moved to the eastside of Rochester in May 2004. Each measurement period was divided into three seasons i.e., winter (December, January, and February), summer (June, July, and August), and the transitional periods (March, April, May, September, October, and November) so as to avoid experimental uncertainty resulting from too large season-to-season variability in ambient temperature and solar photon intensity that would lead to unstable/non-stationary size distributions. Therefore, the seasons were analyzed independently for possible sources. Ten sources were identified at both sites and these include traffic, nucleation, residential/commercial heating, industrial emissions, secondary nitrate, ozone- rich secondary aerosol, secondary sulfate, regionally transported aerosol, and a mixed source of nucleation and traffic. These results show that the measured total outdoor particle number concentrations in Rochester generally vary with similar temporal patterns, suggesting that the central monitoring site data can be used to estimate outdoor exposure in other parts of the city.

Facebook, Twitter Activities Sites, Location and Students' Interest in Learning

ERIC Educational Resources Information Center

Igbo, J. N.; Ezenwaji, Ifeyinwa; Ajuziogu, Christiana U.

2018-01-01

This study was carried out to ascertain the influence of social networking sites activities (twitter and Facebook) on secondary school students' interest in learning It also considered the impact of these social networking sites activities on location of the students. Two research questions and two null hypotheses guided the study. Mean and…

Oxysterol-binding Protein Activation at Endoplasmic Reticulum-Golgi Contact Sites Reorganizes Phosphatidylinositol 4-Phosphate Pools.

PubMed

Goto, Asako; Charman, Mark; Ridgway, Neale D

2016-01-15

Oxysterol-binding protein (OSBP) exchanges cholesterol and phosphatidylinositol 4-phosphate (PI-4P) at contact sites between the endoplasmic reticulum (ER) and the trans-Golgi/trans-Golgi network. 25-Hydroxycholesterol (25OH) competitively inhibits this exchange reaction in vitro and causes the constitutive localization of OSBP at the ER/Golgi interface and PI-4P-dependent recruitment of ceramide transfer protein (CERT) for sphingomyelin synthesis. We used PI-4P probes and mass analysis to determine how OSBP controls the availability of PI-4P for this metabolic pathway. Treatment of fibroblasts or Chinese hamster ovary (CHO) cells with 25OH caused a 50-70% reduction in Golgi-associated immunoreactive PI-4P that correlated with Golgi localization of OSBP. In contrast, 25OH caused an OSBP-dependent enrichment in Golgi PI-4P that was detected with a pleckstrin homology domain probe. The cellular mass of phosphatidylinositol monophosphates and Golgi PI-4P measured with an unbiased PI-4P probe (P4M) was unaffected by 25OH and OSBP silencing, indicating that OSBP shifts the distribution of PI-4P upon localization to ER-Golgi contact sites. The PI-4P and sterol binding activities of OSBP were both required for 25OH activation of sphingomyelin synthesis, suggesting that 25OH must be exchanged for PI-4P to be concentrated at contact sites. We propose a model wherein 25OH activation of OSBP promotes the binding and retention of PI-4P at ER-Golgi contact sites. This pool of PI-4P specifically recruits pleckstrin homology domain-containing proteins involved in lipid transfer and metabolism, such as CERT. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Variation in concentrations of three mercury (Hg) forms at a rural and a suburban site in New York State.

PubMed

Choi, Hyun-Deok; Huang, Jiaoyan; Mondal, Sumona; Holsen, Thomas M

2013-03-15

Tekran® Hg speciation systems were used at a rural site (Huntington Forest, NY; HF) and a suburban site (Rochester, NY; ROC) to measure gaseous elemental mercury (GEM), gaseous oxidized mercury (GOM), and fine particulate-bound mercury (PBM2.5) concentrations for two years (December 2007 to November 2009). Ancillary data were also available from the New York State Department of Environmental Conservation and the United States Environmental Protection Agency Clean Air Status and Trends Network. Seasonal GEM concentrations were similar at both sites and influenced by factors such as the planet boundary layer (PBL) height and mercury emissions from snow, soil, and point sources. In some seasons, O3 was negatively correlated with GEM at ROC and positively correlated with GEM at HF. At HF, O3 was correlated with GOM and was typically higher in the afternoon. The cause of this pattern may be photochemical reactions during the day, and the GOM diel pattern may also be due to deposition which is enhanced by dew formation during the night and early morning. PBM2.5 concentrations were higher in winter at both sites. This is indicative of local wood combustion for space heating in winter, increased sorption to particles at lower temperatures, and lower PBL in the winter. At the suburban site, 2 of 12 events with enhanced GEM/CO ratios were poorly correlated with SO2/GOM, implying that these two events were due either to long range transport or regional metallurgical industries in Canada. Copyright © 2012 Elsevier B.V. All rights reserved.

MgATP-concentration dependence of protection of yeast vacuolar V-ATPase from inactivation by 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole supports a bi-site catalytic mechanism of ATP hydrolysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Milgrom, Elena M.; Milgrom, Yakov M., E-mail: milgromy@upstate.edu

2012-06-29

Highlights: Black-Right-Pointing-Pointer MgATP protects V-ATPase from inactivation by 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole. Black-Right-Pointing-Pointer V-ATPase activity saturation with MgATP is not sufficient for complete protection. Black-Right-Pointing-Pointer The results support a bi-site catalytic mechanism for V-ATPase. -- Abstract: Catalytic site occupancy of the yeast vacuolar V-ATPase during ATP hydrolysis in the presence of an ATP-regenerating system was probed using sensitivity of the enzyme to inhibition by 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl). The results show that, regardless of the presence or absence of the proton-motive force across the vacuolar membrane, saturation of V-ATPase activity at increasing MgATP concentrations is accompanied by only partial protection of the enzyme from inhibitionmore » by NBD-Cl. Both in the presence and absence of an uncoupler, complete protection of V-ATPase from inhibition by NBD-Cl requires MgATP concentrations that are significantly higher than those expected from the K{sub m} values for MgATP. The results are inconsistent with a tri-site model and support a bi-site model for a mechanism of ATP hydrolysis by V-ATPase.« less

Active Site Hydrophobicity and the Convergent Evolution of Paraoxonase Activity in Structurally Divergent Enzymes: The Case of Serum Paraoxonase 1

PubMed Central

2016-01-01

Serum paraoxonase 1 (PON1) is a native lactonase capable of promiscuously hydrolyzing a broad range of substrates, including organophosphates, esters, and carbonates. Structurally, PON1 is a six-bladed β-propeller with a flexible loop (residues 70–81) covering the active site. This loop contains a functionally critical Tyr at position 71. We have performed detailed experimental and computational analyses of the role of selected Y71 variants in the active site stability and catalytic activity in order to probe the role of Y71 in PON1’s lactonase and organophosphatase activities. We demonstrate that the impact of Y71 substitutions on PON1’s lactonase activity is minimal, whereas the kcat for the paraoxonase activity is negatively perturbed by up to 100-fold, suggesting greater mutational robustness of the native activity. Additionally, while these substitutions modulate PON1’s active site shape, volume, and loop flexibility, their largest effect is in altering the solvent accessibility of the active site by expanding the active site volume, allowing additional water molecules to enter. This effect is markedly more pronounced in the organophosphatase activity than the lactonase activity. Finally, a detailed comparison of PON1 to other organophosphatases demonstrates that either a similar “gating loop” or a highly buried solvent-excluding active site is a common feature of these enzymes. We therefore posit that modulating the active site hydrophobicity is a key element in facilitating the evolution of organophosphatase activity. This provides a concrete feature that can be utilized in the rational design of next-generation organophosphate hydrolases that are capable of selecting a specific reaction from a pool of viable substrates. PMID:28026940

Active Site Hydrophobicity and the Convergent Evolution of Paraoxonase Activity in Structurally Divergent Enzymes: The Case of Serum Paraoxonase 1.

PubMed

Blaha-Nelson, David; Krüger, Dennis M; Szeler, Klaudia; Ben-David, Moshe; Kamerlin, Shina Caroline Lynn

2017-01-25

Serum paraoxonase 1 (PON1) is a native lactonase capable of promiscuously hydrolyzing a broad range of substrates, including organophosphates, esters, and carbonates. Structurally, PON1 is a six-bladed β-propeller with a flexible loop (residues 70-81) covering the active site. This loop contains a functionally critical Tyr at position 71. We have performed detailed experimental and computational analyses of the role of selected Y71 variants in the active site stability and catalytic activity in order to probe the role of Y71 in PON1's lactonase and organophosphatase activities. We demonstrate that the impact of Y71 substitutions on PON1's lactonase activity is minimal, whereas the k cat for the paraoxonase activity is negatively perturbed by up to 100-fold, suggesting greater mutational robustness of the native activity. Additionally, while these substitutions modulate PON1's active site shape, volume, and loop flexibility, their largest effect is in altering the solvent accessibility of the active site by expanding the active site volume, allowing additional water molecules to enter. This effect is markedly more pronounced in the organophosphatase activity than the lactonase activity. Finally, a detailed comparison of PON1 to other organophosphatases demonstrates that either a similar "gating loop" or a highly buried solvent-excluding active site is a common feature of these enzymes. We therefore posit that modulating the active site hydrophobicity is a key element in facilitating the evolution of organophosphatase activity. This provides a concrete feature that can be utilized in the rational design of next-generation organophosphate hydrolases that are capable of selecting a specific reaction from a pool of viable substrates.

Characterization of the active site properties of CYP4F12.

PubMed

Eksterowicz, John; Rock, Dan A; Rock, Brooke M; Wienkers, Larry C; Foti, Robert S

2014-10-01

Cytochrome P450 4F12 is a drug-metabolizing enzyme that is primarily expressed in the liver, kidney, colon, small intestine, and heart. The properties of CYP4F12 that may impart an increased catalytic selectivity (decreased promiscuity) were explored through in vitro metabolite elucidation, kinetic isotope effect experiments, and computational modeling of the CYP4F12 active site. By using astemizole as a probe substrate for CYP4F12 and CYP3A4, it was observed that although CYP4F12 favored astemizole O-demethylation as the primary route of metabolism, CYP3A4 was capable of metabolizing astemizole at multiple sites on the molecule. Deuteration of astemizole at the site of O-demethylation resulted in an isotope effect of 7.1 as well as an 8.3-fold decrease in the rate of clearance for astemizole by CYP4F12. Conversely, although an isotope effect of 3.8 was observed for the formation of the O-desmethyl metabolite when deuterated astemizole was metabolized by CYP3A4, there was no decrease in the clearance of astemizole. Development of a homology model of CYP4F12 based on the crystal structure of cytochrome P450 BM3 predicted an active site volume for CYP4F12 that was approximately 76% of the active site volume of CYP3A4. As predicted, multiple favorable binding orientations were available for astemizole docked into the active site of CYP3A4, but only a single binding orientation with the site of O-demethylation oriented toward the heme was identified for CYP4F12. Overall, it appears that although CYP4F12 may be capable of binding similar ligands to other cytochrome P450 enzymes such as CYP3A4, the ability to achieve catalytically favorable orientations may be inherently more difficult because of the increased steric constraints of the CYP4F12 active site. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Deletion of loop fragment adjacent to active site diminishes the stability and activity of exo-inulinase.

PubMed

Arjomand, Maryam Rezaei; Habibi-Rezaei, Mehran; Ahmadian, Gholamreza; Hassanzadeh, Malihe; Karkhane, Ali Asghar; Asadifar, Mandana; Amanlou, Massoud

2016-11-01

Inulinases are classified as hydrolases and widely used in the food and medical industries. Here, we report the deletion of a six-membered adjacent active site loop fragment ( 74 YGSDVT 79 sequence) from third Ω-loop of the exo-inulinase containing aspartate residue from Aspergillus niger to study its structural and functional importance. Site-directed mutagenesis was used to create the mutant of the exo-inulinase (Δ6SL). To investigate the stability of the region spanning this loop, MD simulations were performed 80ns for 20-85 residues. Molecular docking was performed to compare the interactions in the active sites of enzymes with fructose as a ligand. Accordingly, the functional thermostability of the exo-inulinase was significantly decreased upon loop fragment deletion. Evaluation of the kinetics parameters (V max , K m , k cat and, k cat /K m ) and activation energy (E a ) of the catalysis of enzymes indicated the importance of the deleted sequence on the catalytic performance of the enzyme. In conclusion, six-membered adjacent active site loop fragment not only plays a crucial role in the stability of the enzyme, but also it involves in the enzyme catalysis through lowering the activation energy of the catalysis and effective improving the catalytic performance. Copyright © 2016. Published by Elsevier B.V.

The Crystal Structure of a Quercetin 2,3-Dioxygenase from Bacillus subtilis Suggests Modulation of Enzyme Activity by a Change in the Metal Ion at the Active Site(s)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gopal, B.; Madan, Lalima L.; Betz, Stephen F.

2010-11-10

Common structural motifs, such as the cupin domains, are found in enzymes performing different biochemical functions while retaining a similar active site configuration and structural scaffold. The soil bacterium Bacillus subtilis has 20 cupin genes (0.5% of the total genome) with up to 14% of its genes in the form of doublets, thus making it an attractive system for studying the effects of gene duplication. There are four bicupins in B. subtilis encoded by the genes yvrK, yoaN, yxaG, and ywfC. The gene products of yvrK and yoaN function as oxalate decarboxylases with a manganese ion at the active site(s),more » whereas YwfC is a bacitracin synthetase. Here we present the crystal structure of YxaG, a novel iron-containing quercetin 2,3-dioxygenase with one active site in each cupin domain. Yxag is a dimer, both in solution and in the crystal. The crystal structure shows that the coordination geometry of the Fe ion is different in the two active sites of YxaG. Replacement of the iron at the active site with other metal ions suggests modulation of enzymatic activity in accordance with the Irving-Williams observation on the stability of metal ion complexes. This observation, along with a comparison with the crystal structure of YvrK determined recently, has allowed for a detailed structure-function analysis of the active site, providing clues to the diversification of function in the bicupin family of proteins.« less

Effects of hexamethonium, phenothiazines, propranolol and ephedrine on acetylcholinesterase carbamylation by physostigmine, aldicarb and carbaryl: interaction between the active site and the functionally distinct peripheral sites in acetylcholinesterase.

PubMed

Singh, A K; Spassova, D

1998-01-01

Physostigmine, aldicarb and carbaryl were potent inhibitors of acetylcholinesterase (AChE). The physostigmine-inhibited AChE fluoresced at 300 nm excitation and 500 nm emission wavelengths, but the aldicarb and carbaryl inhibited enzyme did not. This suggests that the carbamylated active center is not the fluorescing site in AChE. The fluorescence intensity of physostigmine-inhibited AChE decreased with increasing the substrate (acetylthiocholine) concentration, thus indicating that physostigmine binding to the active site is essential for the development of fluorescence. Thus, the physostigmine-inhibited AChE fluoresces due to the binding of trimethylpyrrolo[2,3-b]indol (TMPI) moiety, formed by the hydrolysis of physostigmine, to a peripheral site in AChE. The fluorescence intensity of the physostigmine-inhibited enzyme decreased when the inhibited-enzyme was dialyzed for either 30 min that poorly reactivated the enzyme or 180 min that fully reactivated the enzyme. This suggests that dialysis dissociates the AChE-TMPI complex much faster than it reactivates the carbamylated AChE. Ephedrine, propranolol and phenothiazines including trifluoparazine (TPZ) caused non-competitive inhibition, while hexamethonium caused an uncompetitive inhibition of AChE activity. TPZ, upon binding with AChE, formed a fluorescent TPZ-enzyme complex. The fluorescence intensity of TPZ-AChE complex was effectively decreased by ephedrine, but not by propranolol or hexamethonium. This indicates that TPZ and ephedrine bind to the same site in AChE which is different from the site/or sites to which propranolol or hexamethonium bind. Hexamethonium protected AChE from inhibition by carbamates and decreased the fluorescence intensity of the physostigmine-inhibited AChE. Phenothiazines and ephedrine did not modulate the enzyme inhibition or the fluorescence intensity of the physostigmine-inhibited AChE. Propranolol and TPZ potentiated the enzyme inhibition and increased the fluorescence intensity

Concentrations and content of mercury in bark, wood, and leaves in hardwoods and conifers in four forested sites in the northeastern USA

PubMed Central

Yanai, Ruth D.; Driscoll, Charles T.; Montesdeoca, Mario; Smith, Kevin T.

2018-01-01

Mercury (Hg) is deposited from the atmosphere to remote areas such as forests, but the amount of Hg in trees is not well known. To determine the importance of Hg in trees, we analyzed foliage, bark and bole wood of eight tree species at four sites in the northeastern USA (Huntington Forest, NY; Sleepers River, VT; Hubbard Brook, NH; Bear Brook, ME). Foliar concentrations of Hg averaged 16.3 ng g-1 among the hardwood species, which was significantly lower than values in conifers, which averaged 28.6 ng g-1 (p < 0.001). Similarly, bark concentrations of Hg were lower (p < 0.001) in hardwoods (7.7 ng g-1) than conifers (22.5 ng g-1). For wood, concentrations of Hg were higher in yellow birch (2.1–2.8 ng g-1) and white pine (2.3 ng g-1) than in the other species, which averaged 1.4 ng g-1 (p < 0.0001). Sites differed significantly in Hg concentrations of foliage and bark (p = 0.02), which are directly exposed to the atmosphere, but the concentration of Hg in wood depended more on species (p < 0.001) than site (p = 0.60). The Hg contents of tree tissues in hardwood stands, estimated from modeled biomass and measured concentrations at each site, were higher in bark (mean of 0.10 g ha-1) and wood (0.16 g ha-1) than in foliage (0.06 g ha-1). In conifer stands, because foliar concentrations were higher, the foliar pool tended to be more important. Quantifying Hg in tree tissues is essential to understanding the pools and fluxes of Hg in forest ecosystems. PMID:29684081

Tissue radionuclide concentrations in water birds and upland birds on the Hanford Site (USA) from 1971-2009.

PubMed

Delistraty, Damon; Van Verst, Scott

2011-08-01

Historical operations at the Hanford Site (Washington State, USA) have released a wide array of non-radionuclide and radionuclide contaminants into the environment. As a result, there is a need to characterize contaminant effects on site biota. Within this framework, the main purpose of our study was to evaluate radionuclide concentrations in bird tissue, obtained from the Hanford Environmental Information System (HEIS). The database was sorted by avian group (water bird vs. upland bird), radionuclide (over 20 analytes), tissue (muscle, bone, liver), location (onsite vs. offsite), and time period (1971-1990 vs. 1991-2009). Onsite median concentrations in water birds were significantly higher (Bonferroni P < 0.05) than those in onsite upland birds for Cs-137 in muscle (1971-1990) and Sr-90 in bone (1991-2009), perhaps due to behavioral, habitat, or trophic species differences. Onsite median concentrations in water birds were higher (borderline significance with Bonferroni P = 0.05) than those in offsite birds for Cs-137 in muscle (1971-1990). Onsite median concentrations in the earlier time period were significantly higher (Bonferroni P < 0.05) than those in the later time period for Co-60, Cs-137, Eu-152, and Sr-90 in water bird muscle and for Cs-137 in upland bird muscle tissue. Median concentrations of Sr-90 in bone were significantly higher (Bonferroni P < 0.05) than those in muscle for both avian groups and both locations. Over the time period, 1971-2009, onsite median internal dose was estimated for each radionuclide in water bird and upland bird tissues. However, a meaningful dose comparison between bird groups was not possible, due to a dissimilar radionuclide inventory, mismatch of time periods for input radionuclides, and lack of an external dose estimate. Despite these limitations, our results contribute toward ongoing efforts to characterize ecological risk at the Hanford Site. Copyright © 2011 Elsevier Ltd. All rights reserved.

Hourly elemental concentrations in PM2.5 aerosols sampled simultaneously at urban background and road site during SAPUSS - diurnal variations and PMF receptor modelling

NASA Astrophysics Data System (ADS)

Dall'Osto, M.; Querol, X.; Amato, F.; Karanasiou, A.; Lucarelli, F.; Nava, S.; Calzolai, G.; Chiari, M.

2013-04-01

monitoring sites at similar concentrations regardless their different ventilation conditions; (2) by contrast, local industrial aerosol plumes associated with shipping oil combustion and smelters activities have a higher impact on the more ventilated UB site; (3) a unique source of Pb-Cl (associated with combustion emissions) is found to be the major (82%) source of fine Cl in the urban agglomerate; (4) the mean diurnal variation of PM2.5 primary traffic non-exhaust brake dust (Fe-Cu) suggests that this source is mainly emitted and not resuspended, whereas PM2.5 urban dust (Ca) is found mainly resuspended by both traffic vortex and sea breeze; (5) urban dust (Ca) is found the aerosol source most affected by land wetness, reduced by a factor of eight during rainy days and suggesting that wet roads may be a solution for reducing urban dust concentrations.

Ice dynamics of the Allan Hills meteorite concentration sites revealed by satellite aperture radar interferometry

NASA Astrophysics Data System (ADS)

Coren, F.; Delisle, G.; Sterzai, P.

2003-09-01

The ice flow conditions of a 100 x 100 km area of Victoria Land, Antarctica were analyzed with the synthetic aperture radar (SAR) technique. The area includes a number of meteorite concentration sites, in particular the Allan Hills ice fields. Regional ice flow velocities around the Mid- western and Near-western ice fields and the Allan Hills main ice field are shown to be 2.5 m yr-1. These sites are located on a horseshoe-shaped area that bounds an area characterized by higher ice flow velocities of up to 5 m yr-1. Meteorite find locations on the Elephant Moraine are located in this "high ice flow" area. The SAR derived digital elevation model (DEM) shows atypical low surface slopes for Antarctic conditions, which are the cause for the slow ice movements. Numerous ice rises in the area are interpreted to cap sub-ice obstacles, which were formed by tectonic processes in the past. The ice rises are considered to represent temporary features, which develop only during warm stages when the regional ice stand is lowered. Ice depressions, which develop in warm stages on the lee side of ice rises, may act as the sites of temporary build-up of meteorite concentrations, which turn inoperative during cold stages when the regional ice level rises and the ice rises disappear. Based on a simplified ice flow model, we argue that the regional ice flow in cold stages is reduced by a factor of at least 3.

The role of dust storms in total atmospheric particle concentrations at two sites in the western U.S.

USGS Publications Warehouse

Neff, Jason C.; Reynolds, Richard L.; Munson, Seth M.; Fernandez, Daniel; Belnap, Jayne

2013-01-01

Mineral aerosols are produced during the erosion of soils by wind and are a common source of particles (dust) in arid and semiarid regions. The size of these particles varies widely from less than 2 µm to larger particles that can exceed 50 µm in diameter. In this study, we present two continuous records of total suspended particle (TSP) concentrations at sites in Mesa Verde and Canyonlands National Parks in Colorado and Utah, USA, respectively, and compare those values to measurements of fine and coarse particle concentrations made from nearby samplers. Average annual concentrations of TSP at Mesa Verde were 90 µg m−3 in 2011 and at Canyonlands were 171 µg m−3 in 2009, 113 µg m−3 in 2010, and 134 µg m−3 in 2011. In comparison, annual concentrations of fine (diameter of 2.5 µm and below) and coarse (2.5–10 µm diameter) particles at these sites were below 10 µg m−3 in all years. The high concentrations of TSP appear to be the result of regional dust storms with elevated concentrations of particles greater than 10 µm in diameter. These conditions regularly occur from spring through fall with 2 week mean TSP periodically in excess of 200 µg m−3. Measurement of particles on filters indicates that the median particle size varies between approximately 10 µm in winter and 40 µm during the spring. These persistently elevated concentrations of large particles indicate that regional dust emission as dust storms and events are important determinants of air quality in this region.

Impact of informal electronic waste recycling on metal concentrations in soils and dusts.

PubMed

Ohajinwa, Chimere May; van Bodegom, Peter M; Vijver, Martina G; Peijnenburg, Willie J G M

2018-07-01

Electronic and electrical equipment contains over 1000 different substances, including metals. During informal e-waste recycling some of these substances such as metals, are released into the environment causing environmental pollution. This study assessed the impact of different informal e-waste recycling activities (burning, dismantling, and repairing) on metal concentrations in top soils and various dust. A comparative cross-sectional study design was adopted to assess metal concentrations in top soils and in various dust samples from multiple e-waste recycling sites. Metal concentrations at e-waste recycling sites were compared to the concentrations at control sites in three study locations in Nigeria (Lagos, Ibadan, and Aba). In the three study locations, mean metal concentrations at the e-waste recycling sites exceeded the concentrations at the control sites and the Nigerian standard guideline values by 100 s to 1000 s times. Burning sites showed the highest pollution level, followed by dismantling sites, then repair sites. Our findings show serious environmental and public health concerns. The metal concentrations were also higher than levels reported in other studies at the same locations in Nigeria, indicating that the situation is worsening. This study provides scientific evidence for an urgent need to develop effective strategies to strengthen enforcement of existing e-waste regulations in Nigeria. Copyright © 2018 Elsevier Inc. All rights reserved.

PM2.5 Indoor Air Quality at Two Sites in London Ontario - A Case Study

NASA Astrophysics Data System (ADS)

Mates, A. V.; Xu, X.; Gilliland, J.; Maltby, M. J.

2010-12-01

Studies have shown an association between ambient fine particulate matter (PM2.5) and health impacts, particularly for the elderly and children. As part of a larger study, PM2.5 concentrations were measured using the DustTrak (Model 8520, TSI, St. Paul, MN, USA) at two schools within the city of London, Ontario (Canada). Site A was in a suburban environment while site B was in an urban setting. Monitoring took place for 3 weeks during winter (Feb. 16 - Mar. 8) and 3 weeks during spring (May 05 - 25) of 2010. The winter campaign monitored indoor PM2.5 only, while the spring campaign added outdoor monitors (PM2.5 and CO2) after the first week. Ten min. concentrations were used for analysis. Indoor measurements were split into weekday and weekend. For the same time interval, the outdoor concentrations showed mean values of 18 and 21 μg/m3 for sites A & B, respectively, both under the Canada Wide Standard of 30 μg/m3. Measurements at the two sites showed good associations (R^2 = 0.44), during the spring campaign. This indicates that the outdoor PM2.5 had similar sources. For indoor concentrations, Site B showed a significantly different mean concentration 5 times higher compared to site A during the winter ( 8.1 vs. 1.5 μg/m3 ) and 3 times higher (11.9 vs. 3.7 μg/m3) during the spring campaign. Since the outdoor concentrations were similar the large difference in indoor concentrations could be attributed to the following factors: site B being an older building, and the different physical characteristics between the two sites. The spring measurements showed an increase of 50% from weekday to weekend for site A and 22% for site B. The higher level of PM2.5 during weekends is possibly due to the infiltration of outdoor air while the ventilation/filtration system is shut off. During the winter campaign, Site A showed a 14% higher concentration during weekdays compared to weekends while site B weekend concentrations were 17% higher compared to weekday, which will be

A hybrid QM/MM simulation study of intramolecular proton transfer in the pyridoxal 5'-phosphate in the active site of transaminase: influence of active site interaction on proton transfer.

PubMed

Dutta Banik, Sindrila; Chandra, Amalendu

2014-09-25

Pyridoxal 5'-phosphate (PLP) Schiff base, a versatile cofactor, exhibits a tautomeric equilibrium that involves an intramolecular proton transfer between the N-protonated zwitterionic ketoenamine tautomer and the O-protonated covalent enolimine tautomer. It has been postulated that for the catalytic activity, the PLP has to be in the zwitterionic ketoenamine tautomeric form. However, the exact position of the tautomeric equilibrium of Schiff base in the active site of PLP-dependent enzyme is not known yet. In the present work, we investigated the tautomeric equilibrium for the external aldimine state of PLP aspartate (PLP-Asp) Schiff base in the active site of aspartate aminotransferase (AspAT) using combined quantum mechanical and molecular mechanical simulations. The main focus of the present study is to analyze the factors that control the tautomeric equilibrium in the active sites of various PLP-dependent enzymes. The results show that the ketoenamine tautomer is more preferred than the enolimine tautomer both in the gas and aqueous phases as well as in the active site of AspAT. Current simulations show that the active site of AspAT is more suitable for the ketoenamine tautomer compared to the enolimine tautomer. Interestingly, the Tyr225 acts as a proton donor to the phenolic oxygen in the ketoenamine tautomer, while in the covalent enolimine tautomer, it acts as a proton acceptor to the phenolic oxygen. Finally, the metadynamics study confirms this result. The calculated free energy barrier is about 7.5 kcal/mol. A comparative analysis of the microenvironment created by the active site residues of three different PLP-dependent enzymes (aspartate aminotransferase, Dopa decarboxylase, and Ala-racemase) has been carried out to understand the controlling factor(s) of the tautomeric equilibrium. The analysis shows that the intermolecular hydrogen bonding between active site residues and the phenolic oxygen of PLP shifts the tautomeric equilibrium toward the N

Relationships between waste physicochemical properties, microbial activity and vegetation at coal ash and sludge disposal sites.

PubMed

Woch, Marcin W; Radwańska, Magdalena; Stanek, Małgorzata; Łopata, Barbara; Stefanowicz, Anna M

2018-06-11

The aim of the study was to assess the relationships between vegetation, physicochemical and microbial properties of substrate at coal ash and sludge disposal sites. The study was performed on 32 plots classified into 7 categories: dried ash sedimentation ponds, dominated by a grass Calamagrostis epigejos (AH-Ce), with the admixture of Pinus sylvestris (AH-CePs) or Robinia pseudoacacia (AH-CeRp), dry ash landfill dominated by Betula pendula and Pinus sylvestris (AD-BpPs) or Salix viminalis (AD-Sv) and coal sludge pond with drier parts dominated by Tussilago farfara (CS-Tf) and the wetter ones by Cyperus flavescens (CS-Cf). Ash sites were covered with soil layer imported as a part of technical reclamation. Ash had relatively high concentrations of some alkali and alkaline earth metals, Mn and pH, while coal sludge had high water and C, S, P and K contents. Concentrations of heavy metals were lower than allowable limits in all substrate types. Microbial biomass and, particularly, enzymatic activity in ash and sludge were generally low. The only exception were CS-Tf plots characterized by the highest microbial biomass, presumably due to large deposits of organic matter that became available for aerobic microbial biomass when water level fell. The properties of ash and sludge adversely affected microbial biomass and enzymatic activity as indicated by significant negative correlations between the content of alkali/alkaline earth metals, heavy metals, and macronutrients with enzymatic activity and/or microbial biomass, as well as positive correlations of these parameters with metabolic quotient (qCO 2 ). Plant species richness and cover were relatively high, which may be partly associated with alleviating influence of soil covering the ash. The effect of the admixture of R. pseudoacacia or P. sylvestris to stands dominated by C. epigejos was smaller than expected. The former species increased NNH 4 , NNO 3 and arylsulfatase activity, while the latter reduced activity of

Phytochemical, antimicrobial, and antioxidant activities of different citrus juice concentrates.

PubMed

Oikeh, Ehigbai I; Omoregie, Ehimwenma S; Oviasogie, Faith E; Oriakhi, Kelly

2016-01-01

The search for new antimicrobial compounds is ongoing. Its importance cannot be overemphasized in an era of emerging resistant pathogenic organisms. This study therefore investigated the phytochemical composition and antioxidant and antimicrobial activities of different citrus juice concentrates. Fruit juices of Citrus tangerine (tangerine), Citrus paradisi (grape), Citrus limon (lemon), and Citrus aurantifolia (lime) were evaluated. Antimicrobial activities against five bacterial and three fungal strains were evaluated. The results revealed the presence of alkaloids, flavonoids, steroids, terpenoids, saponins, cardiac glycosides, and reducing sugars in all the juice concentrates. DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging capacities varied with tangerine and grape juices having better scavenging capacities than lemon and lime juices. Grape juice was observed to have a significantly higher (P < 0.05) ferric-reducing antioxidant potential (FRAP) value (364.2 ± 10.25 μmol/L Fe(II)/g of the extract) than the reference antioxidant, ascorbic acid (312.88 ± 5.61 μmol/L). Antimicrobial studies revealed differential antimicrobial activities against different microbial strains. Zones of inhibition ranging from 4 to 26 mm were observed for the antibacterial tests with 0-24 mm for antifungal test. Minimum inhibitory concentrations (MIC) and minimum bacteriostatic concentrations (MBC) for concentrates against bacterial strains ranged from 12.5 to 200 μg/mL. Lemon and lime juice concentrates had lower MIC and MBC values with orange and tangerine having the highest values. Minimum fungicidal concentrations ranged from 50 to 200 μg/mL. The results of this study suggest that these juice concentrates may have beneficial antimicrobial roles that can be exploited in controlling unwanted microbial growth.

Community Update on Site Activities, July 19, 2013

EPA Pesticide Factsheets

In an effort to engage and inform community members interested in the New Bedford Harbor Superfund Site cleanup, EPA will be issuing periodic topic-based fact sheets that will provide background information and updates about ongoing activities.

Structural and Kinetic Analyses of Macrophage Migration Inhibitory Factor Active Site Interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Crichlow, G.; Lubetsky, J; Leng, L

Macrophage migration inhibitory factor (MIF) is a secreted protein expressed in numerous cell types that counters the antiinflammatory effects of glucocorticoids and has been implicated in sepsis, cancer, and certain autoimmune diseases. Interestingly, the structure of MIF contains a catalytic site resembling the tautomerase/isomerase sites of microbial enzymes. While bona fide physiological substrates remain unknown, model substrates have been identified. Selected compounds that bind in the tautomerase active site also inhibit biological functions of MIF. It had previously been shown that the acetaminophen metabolite, N-acetyl-p-benzoquinone imine (NAPQI), covalently binds to the active site of MIF. In this study, kinetic datamore » indicate that NAPQI inhibits MIF both covalently and noncovalently. The structure of MIF cocrystallized with NAPQI reveals that the NAPQI has undergone a chemical alteration forming an acetaminophen dimer (bi-APAP) and binds noncovalently to MIF at the mouth of the active site. We also find that the commonly used protease inhibitor, phenylmethylsulfonyl fluoride (PMSF), forms a covalent complex with MIF and inhibits the tautomerase activity. Crystallographic analysis reveals the formation of a stable, novel covalent bond for PMSF between the catalytic nitrogen of the N-terminal proline and the sulfur of PMSF with complete, well-defined electron density in all three active sites of the MIF homotrimer. Conclusions are drawn from the structures of these two MIF-inhibitor complexes regarding the design of novel compounds that may provide more potent reversible and irreversible inhibition of MIF.« less

Rac1 GTPase activates the WAVE regulatory complex through two distinct binding sites.

PubMed

Chen, Baoyu; Chou, Hui-Ting; Brautigam, Chad A; Xing, Wenmin; Yang, Sheng; Henry, Lisa; Doolittle, Lynda K; Walz, Thomas; Rosen, Michael K

2017-09-26

The Rho GTPase Rac1 activates the WAVE regulatory complex (WRC) to drive Arp2/3 complex-mediated actin polymerization, which underpins diverse cellular processes. Here we report the structure of a WRC-Rac1 complex determined by cryo-electron microscopy. Surprisingly, Rac1 is not located at the binding site on the Sra1 subunit of the WRC previously identified by mutagenesis and biochemical data. Rather, it binds to a distinct, conserved site on the opposite end of Sra1. Biophysical and biochemical data on WRC mutants confirm that Rac1 binds to both sites, with the newly identified site having higher affinity and both sites required for WRC activation. Our data reveal that the WRC is activated by simultaneous engagement of two Rac1 molecules, suggesting a mechanism by which cells may sense the density of active Rac1 at membranes to precisely control actin assembly.

Monocopper active site for partial methane oxidation in Cu-exchanged 8MR zeolites

DOE PAGES

Kulkarni, Ambarish R.; Zhao, Zhi -Jian; Siahrostami, Samira; ...

2016-08-17

Direct conversion of methane to methanol using oxygen is experiencing renewed interest owing to the availability of new natural gas resources. Copper-exchanged zeolites such as mordenite and ZSM-5 have shown encouraging results, and di- and tri-copper species have been suggested as active sites. Recently, small eight-membered ring (8MR) zeolites including SSZ-13, -16, and -39 have been shown to be active for methane oxidation, but the active sites and reaction mechanisms in these 8MR zeolites are not known. In this work, we use density functional theory (DFT) calculations to systematically evaluate monocopper species as active sites for the partial methane oxidationmore » reaction in Cu-exchanged SSZ-13. On the basis of kinetic and thermodynamic arguments, we suggest that [Cu IIOH] + species in the 8MR are responsible for the experimentally observed activity. Furthermore, our results successfully explain the available spectroscopic data and experimental observations including (i) the necessity of water for methanol extraction and (ii) the effect of Si/Al ratio on the catalyst activity. Monocopper species have not yet been suggested as an active site for the partial methane oxidation reaction, and our results suggest that [Cu IIOH] + active site may provide complementary routes for methane activation in zeolites in addition to the known [Cu–O–Cu] 2+ and Cu 3O 3 motifs.« less

What Motivates Young Adults to Talk About Physical Activity on Social Network Sites?

PubMed

Zhang, Ni; Campo, Shelly; Yang, Jingzhen; Eckler, Petya; Snetselaar, Linda; Janz, Kathleen; Leary, Emily

2017-06-22

Electronic word-of-mouth on social network sites has been used successfully in marketing. In social marketing, electronic word-of-mouth about products as health behaviors has the potential to be more effective and reach more young adults than health education through traditional mass media. However, little is known about what motivates people to actively initiate electronic word-of-mouth about health behaviors on their personal pages or profiles on social network sites, thus potentially reaching all their contacts on those sites. This study filled the gap by applying a marketing theoretical model to explore the factors associated with electronic word-of-mouth on social network sites about leisure-time physical activity. A Web survey link was sent to undergraduate students at one of the Midwestern universities and 439 of them completed the survey. The average age of the 439 participants was 19 years (SD=1 year, range: 18-24). Results suggested that emotional engagement with leisure-time physical activity (ie, affective involvement in leisure-time physical activity) predicted providing relevant opinions or information on social network sites. Social network site users who perceived stronger ties with all their contacts were more likely to provide and seek leisure-time physical activity opinions and information. People who provided leisure-time physical activity opinions and information were more likely to seek opinions and information, and people who forwarded information about leisure-time physical activity were more likely to chat about it. This study shed light on the application of the electronic word-of-mouth theoretical framework in promoting health behaviors. The findings can also guide the development of future social marketing interventions using social network sites to promote leisure-time physical activity. ©Ni Zhang, Shelly Campo, Jingzhen Yang, Petya Eckler, Linda Snetselaar, Kathleen Janz, Emily Leary. Originally published in the Journal of Medical

What Motivates Young Adults to Talk About Physical Activity on Social Network Sites?

PubMed Central

Campo, Shelly; Yang, Jingzhen; Eckler, Petya; Snetselaar, Linda; Janz, Kathleen; Leary, Emily

2017-01-01

Background Electronic word-of-mouth on social network sites has been used successfully in marketing. In social marketing, electronic word-of-mouth about products as health behaviors has the potential to be more effective and reach more young adults than health education through traditional mass media. However, little is known about what motivates people to actively initiate electronic word-of-mouth about health behaviors on their personal pages or profiles on social network sites, thus potentially reaching all their contacts on those sites. Objective This study filled the gap by applying a marketing theoretical model to explore the factors associated with electronic word-of-mouth on social network sites about leisure-time physical activity. Methods A Web survey link was sent to undergraduate students at one of the Midwestern universities and 439 of them completed the survey. Results The average age of the 439 participants was 19 years (SD=1 year, range: 18-24). Results suggested that emotional engagement with leisure-time physical activity (ie, affective involvement in leisure-time physical activity) predicted providing relevant opinions or information on social network sites. Social network site users who perceived stronger ties with all their contacts were more likely to provide and seek leisure-time physical activity opinions and information. People who provided leisure-time physical activity opinions and information were more likely to seek opinions and information, and people who forwarded information about leisure-time physical activity were more likely to chat about it. Conclusions This study shed light on the application of the electronic word-of-mouth theoretical framework in promoting health behaviors. The findings can also guide the development of future social marketing interventions using social network sites to promote leisure-time physical activity. PMID:28642215

Concentration and temperature effects on ovostatin activity

NASA Technical Reports Server (NTRS)

Moriarity, Debra M.

1994-01-01

Light scattering experiments performed at Mississippi State University using MSFC ovostatin preparations indicated that at low ovostatin concentrations, below 0.2 mg/ml, the protein was dissociating from a tetramer into dimers. Since the proposed mechanism of action involved the tetrameric form of the protein, we hypothesized that perhaps under the conditions of our assays at various O/T ratios the ovostatin was becoming dissociated into an inactive dimer. To examine this possibility we assayed the ovostatin activity as a function of ovostatin concentration and of temperature of the assay. Data are presented that show the results of these assays at 23 C, 30 C, 37 C and 42 C respectively. The data are highly suggestive that there is a decrease in ovostatin activity as the concentration of the protein falls below 0.06 mg/ml. This may not be of any physiological importance, however, since the concentration of ovostatin in the egg is about 0.5 mg/ml. Curiously, the dissociation of the tetramer into dimers does not show a significant temperature dependence as would be expected for an equilibrium reaction. Whether this is in fact the case, or whether the differences are so small as to not be discerned from the current data remains to be seen. Another aspect to consider is that in the egg the primary role of the ovostatin may or may not be as a protease inhibitor. Although the inhibition of collagenase by ovostatin may be an important aspect of embryogenesis, it is also possible that it functions as a binding protein for some substance. In this regard, all ovostatin preparations from MSFC have shown an approximately 88,000 MW protein associated with the ovostatin. The identity of this protein is not currently known and may be the subject of future studies.

Corrosion Research And Web Site Activities

NASA Technical Reports Server (NTRS)

Heidersbach, Robert H.

2001-01-01

This report covers corrosion-related activities at the NASA Kennedy Space Center during the summer of 2000. The NASA Kennedy Space Center's corrosion web site, corrosion.ksc.nasa.gov, was updated with new information based on feedback over the past two years. The methodology for a two-year atmospheric exposure testing program to study the effectiveness of commercial chemicals sold for rinsing aircraft and other equipment was developed and some preliminary laboratory chemical analyses are presented.

Corrosion Research and Web Site Activities

NASA Technical Reports Server (NTRS)

Heidersbach, Robert H.

2002-01-01

This report covers corrosion-related activities at the NASA Kennedy Space Center during the summer of 2000. The NASA Kennedy Space Center's corrosion web site, corrosion.ksc.nasa.gov, was updated with new information based on feedback over the past two years. The methodology for a two-year atmospheric exposure testing program to study the effectiveness of commercial chemicals sold for rinsing aircraft and other equipment was developed and some preliminary laboratory chemical analyses are presented.

Solution Behavior and Activity of a Halophilic Esterase under High Salt Concentration

PubMed Central

Rao, Lang; Zhao, Xiubo; Pan, Fang; Li, Yin; Xue, Yanfen; Ma, Yanhe; Lu, Jian R.

2009-01-01

Background Halophiles are extremophiles that thrive in environments with very high concentrations of salt. Although the salt reliance and physiology of these extremophiles have been widely investigated, the molecular working mechanisms of their enzymes under salty conditions have been little explored. Methodology/Principal Findings A halophilic esterolytic enzyme LipC derived from archeaon Haloarcula marismortui was overexpressed from Escherichia coli BL21. The purified enzyme showed a range of hydrolytic activity towards the substrates of p-nitrophenyl esters with different alkyl chains (n = 2−16), with the highest activity being observed for p-nitrophenyl acetate, consistent with the basic character of an esterase. The optimal esterase activities were found to be at pH 9.5 and [NaCl] = 3.4 M or [KCl] = 3.0 M and at around 45°C. Interestingly, the hydrolysis activity showed a clear reversibility against changes in salt concentration. At the ambient temperature of 22°C, enzyme systems working under the optimal salt concentrations were very stable against time. Increase in temperature increased the activity but reduced its stability. Circular dichroism (CD), dynamic light scattering (DLS) and small angle neutron scattering (SANS) were deployed to determine the physical states of LipC in solution. As the salt concentration increased, DLS revealed substantial increase in aggregate sizes, but CD measurements revealed the maximal retention of the α-helical structure at the salt concentration matching the optimal activity. These observations were supported by SANS analysis that revealed the highest proportion of unimers and dimers around the optimal salt concentration, although the coexistent larger aggregates showed a trend of increasing size with salt concentration, consistent with the DLS data. Conclusions/Significance The solution α-helical structure and activity relation also matched the highest proportion of enzyme unimers and dimers. Given that

The proper effect site concentration of remifentanil for prevention of myoclonus after etomidate injection

PubMed Central

Ri, Hyun Su; Kim, Tae Kyun; Baik, Seung Wan; Yoon, Ji Uk; Byeon, Gyeong Jo

2011-01-01

Background Etomidate frequently induces myoclonus when administered intravenously with bolus injection during anesthetic induction. This can be bothersome for the anesthesiologist. The dose of remifentanil appropriate for preventing myoclonus without side effects was investigated. Methods All patients with American Society of Anesthesiologists (ASA) physical status I-III were divided into three groups (n = 33 per group) according to the pretreatment effect site concentration of remifentanil (Ultiva, Glaxo-Wellcome, München, Germany) of 0, 2 or 4 ng/ml (Group N: 0 ng/ml, Group R: 2 ng/ml, Group Q: 4 ng/ml) by a target controlled infusion (TCI) system. After a 0.3 mg/kg dose of etomidate was injected intravenously for over 1 minute for anesthetic induction, myoclonus was observed. Before the etomidate injection, the patients were pretreated with remifentanil and their side effects were monitored. Results The number of patients showing myoclonus was significantly different among the groups. The incidence of myoclonus was 81%, 12% and 0% (groups N, R, and Q, respectively, P < 0.01). Side effects including bradycardia and hypotension did not occur in either Group R or Q. Chest wall rigidity occured in 45% of patients in Group Q. Conclusions Administration with a 2 ng/ml effect site concentration of remifentanil could reduce the incidence of myoclonus caused by etomidate bolus injection without chest wall rigidity. PMID:21927682

Cooperative activation of cardiac transcription through myocardin bridging of paired MEF2 sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Anderson, Courtney M.; Hu, Jianxin; Thomas, Reuben

2017-03-28

Enhancers frequently contain multiple binding sites for the same transcription factor. These homotypic binding sites often exhibit synergy, whereby the transcriptional output from two or more binding sites is greater than the sum of the contributions of the individual binding sites alone. Although this phenomenon is frequently observed, the mechanistic basis for homotypic binding site synergy is poorly understood. Here in this paper, we identify a bona fide cardiac-specific Prkaa2 enhancer that is synergistically activated by homotypic MEF2 binding sites. We show that two MEF2 sites in the enhancer function cooperatively due to bridging of the MEF2C-bound sites by themore » SAP domain-containing co-activator protein myocardin, and we show that paired sites buffer the enhancer from integration site-dependent effects on transcription in vivo. Paired MEF2 sites are prevalent in cardiac enhancers, suggesting that this might be a common mechanism underlying synergy in the control of cardiac gene expression in vivo.« less

Concentration evolution of pharmaceutically active compounds in raw urban and industrial wastewater.

PubMed

Camacho-Muñoz, Dolores; Martín, Julia; Santos, Juan Luis; Aparicio, Irene; Alonso, Esteban

2014-09-01

The distribution of pharmaceutically active compounds in the environment has been reported in several works in which wastewater treatment plants have been identified as the main source of these compounds to the environment. The concentrations of these compounds in influent wastewater can vary widely not only during the day but also along the year, because of the seasonal-consumption patterns of some pharmaceuticals. However, only few studies have attempted to assess the hourly variability of the concentrations of pharmaceutically active compounds in wastewater. In this work, the distribution and seasonal and hourly variability of twenty-one pharmaceuticals, belonging to seven therapeutic groups, have been investigated in urban and industrial wastewater. The highest concentrations of pharmaceutically active compounds, except salicylic acid, were found in urban wastewater, especially in the case of anti-inflammatory drugs and caffeine. The highest concentrations of salicylic acid were measured in industrial wastewater, reaching concentration levels up to 3295μgL(-)(1). The studied pharmaceutically active compounds showed different distribution patterns during winter and summer periods. Temporal variability of pharmaceutically active compounds during a 24-h period showed a distribution in concordance with their consumption and excretion patterns, in the case of urban wastewater, and with the schedule of industrial activities, in the case of industrial wastewater. Copyright © 2014 Elsevier Ltd. All rights reserved.

Active Site Conformational Dynamics in Human Uridine Phosphorylase 1

PubMed Central

Roosild, Tarmo P.; Castronovo, Samantha

2010-01-01

Uridine phosphorylase (UPP) is a central enzyme in the pyrimidine salvage pathway, catalyzing the reversible phosphorolysis of uridine to uracil and ribose-1-phosphate. Human UPP activity has been a focus of cancer research due to its role in activating fluoropyrimidine nucleoside chemotherapeutic agents such as 5-fluorouracil (5-FU) and capecitabine. Additionally, specific molecular inhibitors of this enzyme have been found to raise endogenous uridine concentrations, which can produce a cytoprotective effect on normal tissues exposed to these drugs. Here we report the structure of hUPP1 bound to 5-FU at 2.3 Å resolution. Analysis of this structure reveals new insights as to the conformational motions the enzyme undergoes in the course of substrate binding and catalysis. The dimeric enzyme is capable of a large hinge motion between its two domains, facilitating ligand exchange and explaining observed cooperativity between the two active sites in binding phosphate-bearing substrates. Further, a loop toward the back end of the uracil binding pocket is shown to flexibly adjust to the varying chemistry of different compounds through an “induced-fit” association mechanism that was not observed in earlier hUPP1 structures. The details surrounding these dynamic aspects of hUPP1 structure and function provide unexplored avenues to develop novel inhibitors of this protein with improved specificity and increased affinity. Given the recent emergence of new roles for uridine as a neuron protective compound in ischemia and degenerative diseases, such as Alzheimer's and Parkinson's, inhibitors of hUPP1 with greater efficacy, which are able to boost cellular uridine levels without adverse side-effects, may have a wide range of therapeutic applications. PMID:20856879

Application of nonparametric regression methods to study the relationship between NO2 concentrations and local wind direction and speed at background sites.

PubMed

Donnelly, Aoife; Misstear, Bruce; Broderick, Brian

2011-02-15

Background concentrations of nitrogen dioxide (NO(2)) are not constant but vary temporally and spatially. The current paper presents a powerful tool for the quantification of the effects of wind direction and wind speed on background NO(2) concentrations, particularly in cases where monitoring data are limited. In contrast to previous studies which applied similar methods to sites directly affected by local pollution sources, the current study focuses on background sites with the aim of improving methods for predicting background concentrations adopted in air quality modelling studies. The relationship between measured NO(2) concentration in air at three such sites in Ireland and locally measured wind direction has been quantified using nonparametric regression methods. The major aim was to analyse a method for quantifying the effects of local wind direction on background levels of NO(2) in Ireland. The method was expanded to include wind speed as an added predictor variable. A Gaussian kernel function is used in the analysis and circular statistics employed for the wind direction variable. Wind direction and wind speed were both found to have a statistically significant effect on background levels of NO(2) at all three sites. Frequently environmental impact assessments are based on short term baseline monitoring producing a limited dataset. The presented non-parametric regression methods, in contrast to the frequently used methods such as binning of the data, allow concentrations for missing data pairs to be estimated and distinction between spurious and true peaks in concentrations to be made. The methods were found to provide a realistic estimation of long term concentration variation with wind direction and speed, even for cases where the data set is limited. Accurate identification of the actual variation at each location and causative factors could be made, thus supporting the improved definition of background concentrations for use in air quality modelling

Maturation of the [Ni-4Fe-4S] active site of carbon monoxide dehydrogenases.

PubMed

Merrouch, Mériem; Benvenuti, Martino; Lorenzi, Marco; Léger, Christophe; Fourmond, Vincent; Dementin, Sébastien

2018-02-14

Nickel-containing enzymes are diverse in terms of function and active site structure. In many cases, the biosynthesis of the active site depends on accessory proteins which transport and insert the Ni ion. We review and discuss the literature related to the maturation of carbon monoxide dehydrogenases (CODH) which bear a nickel-containing active site consisting of a [Ni-4Fe-4S] center called the C-cluster. The maturation of this center has been much less studied than that of other nickel-containing enzymes such as urease and NiFe hydrogenase. Several proteins present in certain CODH operons, including the nickel-binding proteins CooT and CooJ, still have unclear functions. We question the conception that the maturation of all CODH depends on the accessory protein CooC described as essential for nickel insertion into the active site. The available literature reveals biological variations in CODH active site biosynthesis.

CASPASE-9 CARD:CORE DOMAIN INTERACTIONS REQUIRE A PROPERLY-FORMED ACTIVE SITE

PubMed Central

Huber, Kristen L.; Serrano, Banyuhay P.; Hardy, Jeanne A.

2018-01-01

Caspase-9 is a critical factor in the initiation of apoptosis, and as a result is tightly regulated by a number of mechanisms. Caspase-9 contains a Caspase Activation and Recruitment Domain (CARD), which enables caspase-9 to form a tight interaction with the apoptosome, a heptameric activating platform. The caspase-9 CARD has been thought to be principally involved in recruitment to the apoptosome, but its roles outside this interaction have yet to be uncovered. In this work we show that the CARD is involved in physical interactions with the catalytic core of caspase-9 in the absence of the apoptosome; this interaction requires a properly formed caspase-9 active site. The active sites of caspases are composed of four extremely mobile loops. When the active-site loops are not properly ordered, the CARD and core domains of caspase-9 do not interact and behave independently, like loosely tethered beads. When the active-site loop bundle is properly ordered, the CARD domain interacts with the catalytic core, forming a single folding unit. Together these findings provide mechanistic insight into a new level of caspase-9 regulation, prompting speculation that the CARD may also play a role in the recruitment or recognition of substrate. PMID:29500231

Prediction of active sites of enzymes by maximum relevance minimum redundancy (mRMR) feature selection.

PubMed

Gao, Yu-Fei; Li, Bi-Qing; Cai, Yu-Dong; Feng, Kai-Yan; Li, Zhan-Dong; Jiang, Yang

2013-01-27

Identification of catalytic residues plays a key role in understanding how enzymes work. Although numerous computational methods have been developed to predict catalytic residues and active sites, the prediction accuracy remains relatively low with high false positives. In this work, we developed a novel predictor based on the Random Forest algorithm (RF) aided by the maximum relevance minimum redundancy (mRMR) method and incremental feature selection (IFS). We incorporated features of physicochemical/biochemical properties, sequence conservation, residual disorder, secondary structure and solvent accessibility to predict active sites of enzymes and achieved an overall accuracy of 0.885687 and MCC of 0.689226 on an independent test dataset. Feature analysis showed that every category of the features except disorder contributed to the identification of active sites. It was also shown via the site-specific feature analysis that the features derived from the active site itself contributed most to the active site determination. Our prediction method may become a useful tool for identifying the active sites and the key features identified by the paper may provide valuable insights into the mechanism of catalysis.

A multi-site analysis of the association between black carbon concentrations and vehicular idling, traffic, background pollution, and meteorology during school dismissals.

PubMed

Richmond-Bryant, J; Bukiewicz, L; Kalin, R; Galarraga, C; Mirer, F

2011-05-01

A study was performed to assess the relationship between black carbon (BC), passing traffic, and vehicular idling outside New York City (NYC) schools during student dismissal. Monitoring was performed at three school sites in East Harlem, the Bronx, and Brooklyn for 1month per year over a two-year period from November 2006-October 2008. Monitoring at each site was conducted before and after the Asthma Free School Zone (AFSZ) asthma reduction education program was administered. Real-time equipment with a one-minute averaging interval was used to obtain the BC data, while volume counts of idling and passing school busses, trucks, and automobiles were collected each minute by study staff. These data were matched to ambient PM(2.5) and meteorology data obtained from the New York State Department of Environmental Conservation. A generalized additive model (GAM) model was run to examine the relationship between BC concentration and each variable while accounting for site-to-site differences. F-tests were employed to assess the significance of each of the predictor variables. The model results suggested that variability in ambient PM(2.5) concentration contributed 24% of the variability in transformed BC concentration, while variability in the number of idling busses and trucks on the street during dismissal contributed 20% of the variability in transformed BC concentration. The results of this study suggest that a combination of urban scale and local traffic control approaches in combination with cessation of school bus idling will produce improved local BC concentration outside schools. Published by Elsevier B.V.

Effect-site concentration of remifentanil required to blunt haemodynamic responses during tracheal intubation: A randomized comparison between single- and double-lumen tubes.

PubMed

Kim, Tae Kyong; Hong, Deok Man; Lee, Seo Hee; Paik, Hyesun; Min, Se Hee; Seo, Jeong-Hwa; Jung, Chul-Woo; Bahk, Jae-Hyon

2018-01-01

Objective To investigate the effect-site concentration of remifentanil required to blunt haemodynamic responses during tracheal intubation with a single-lumen tube (SLT) or a double-lumen tube (DLT). Methods Patients scheduled for thoracic surgery requiring one-lung ventilation were randomly allocated to either the SLT or DLT group. All patients received a target-controlled infusion of propofol and a predetermined concentration of remifentanil. Haemodynamic parameters during intubation were recorded. The effect-site concentration of remifentanil was determined using a delayed up-and-down sequential allocation method. Results A total of 92 patients were enrolled in the study. The effective effect-site concentrations of remifentanil required to blunt haemodynamic responses in 50% of patients (EC 50 ) estimated by isotonic regression with bootstrapping was higher in the DLT than the SLT group (8.5 ng/ml [95% confidence interval (CI) 8.0-9.5 ng/ml] versus 6.5 ng/ml [95% CI 5.6-6.7 ng/ml], respectively). Similarly, the effective effect-site concentrations of remifentanil in 95% of patients in the DLT group was higher than the SLT group (9.9 ng/ml [95% CI 9.8-10.0 ng/ml] versus 7.0 ng/ml [95% CI 6.9-7.0 ng/ml], respectively). Conclusions This study demonstrated that a DLT requires a 30% higher EC 50 of remifentanil than does an SLT to blunt haemodynamic responses during tracheal intubation when combined with a target-controlled infusion of propofol. Trial registration Clinicaltrials.gov identifier: NCT01542099.

Active site densities, oxygen activation and adsorbed reactive oxygen in alcohol activation on npAu catalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Lu-Cun; Friend, C. M.; Fushimi, Rebecca

The activation of molecular O 2as well as the reactivity of adsorbed oxygen species is of central importance in aerobic selective oxidation chemistry on Au-based catalysts. Herein, we address the issue of O 2activation on unsupported nanoporous gold (npAu) catalysts by applying a transient pressure technique, a temporal analysis of products (TAP) reactor, to measure the saturation coverage of atomic oxygen, its collisional dissociation probability, the activation barrier for O 2dissociation, and the facility with which adsorbed O species activate methanol, the initial step in the catalytic cycle of esterification. The results from these experiments indicate that molecular O 2dissociationmore » is associated with surface silver, that the density of reactive sites is quite low, that adsorbed oxygen atoms do not spill over from the sites of activation onto the surrounding surface, and that methanol reacts quite facilely with the adsorbed oxygen atoms. In addition, the O species from O 2dissociation exhibits reactivity for the selective oxidation of methanol but not for CO. The TAP experiments also revealed that the surface of the npAu catalyst is saturated with adsorbed O under steady state reaction conditions, at least for the pulse reaction.« less

Active site densities, oxygen activation and adsorbed reactive oxygen in alcohol activation on npAu catalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Lu-Cun; Friend, C. M.; Fushimi, Rebecca

2016-01-01

The activation of molecular O 2as well as the reactivity of adsorbed oxygen species is of central importance in aerobic selective oxidation chemistry on Au-based catalysts. Herein, we address the issue of O 2activation on unsupported nanoporous gold (npAu) catalysts by applying a transient pressure technique, a temporal analysis of products (TAP) reactor, to measure the saturation coverage of atomic oxygen, its collisional dissociation probability, the activation barrier for O 2dissociation, and the facility with which adsorbed O species activate methanol, the initial step in the catalytic cycle of esterification. The results from these experiments indicate that molecular O 2dissociationmore » is associated with surface silver, that the density of reactive sites is quite low, that adsorbed oxygen atoms do not spill over from the sites of activation onto the surrounding surface, and that methanol reacts quite facilely with the adsorbed oxygen atoms. In addition, the O species from O 2dissociation exhibits reactivity for the selective oxidation of methanol but not for CO. The TAP experiments also revealed that the surface of the npAu catalyst is saturated with adsorbed O under steady state reaction conditions, at least for the pulse reaction.« less

Observations of fine particulate nitrated phenols in four sites in northern China: concentrations, source apportionment, and secondary formation

NASA Astrophysics Data System (ADS)

Wang, Liwei; Wang, Xinfeng; Gu, Rongrong; Wang, Hao; Yao, Lan; Wen, Liang; Zhu, Fanping; Wang, Weihao; Xue, Likun; Yang, Lingxiao; Lu, Keding; Chen, Jianmin; Wang, Tao; Zhang, Yuanghang; Wang, Wenxing

2018-03-01

Filter samples of fine particulate matters were collected at four sites in northern China (urban, rural, and mountain) in summer and winter, and the contents of nine nitrated phenols were quantified in the laboratory with the use of ultra-high-performance liquid chromatography coupled with mass spectrometry. During the sampling periods, the concentrations of particulate nitrated phenols exhibited distinct temporal and spatial variation. On average, the total concentration of particulate nitrated phenols in urban Jinan in the wintertime reached 48.4 ng m-3, and those in the summertime were 9.8, 5.7, 5.9, and 2.5 ng m-3 in urban Jinan, rural Yucheng and Wangdu, and Mt. Tai, respectively. The elevated concentrations of nitrated phenols in wintertime and in urban areas demonstrate the apparent influences of anthropogenic sources. The positive matrix factorization receptor model was then applied to determine the origins of particulate nitrated phenols in northern China. The five major source factors were traffic, coal combustion, biomass burning, secondary formation, and aged coal combustion plume. Among them, coal combustion played a vital role, especially at the urban site in the wintertime, with a contribution of around 55 %. In the summertime, the observed nitrated phenols were highly influenced by aged coal combustion plumes at all of the sampling sites. Meanwhile, in remote areas, contributions from secondary formation were significant. Further correlation analysis indicates that nitrosalicylic acids were produced mostly from secondary formation that was dominated by NO2 nitration.

Tracing of ca 800 yr old mining activity in peat bog using Pb elemental concentrations and isotope compositions.

NASA Astrophysics Data System (ADS)

Baron, S.; Carignan, J.; Ploquin, A.

2003-04-01

Sixty sites of slags have been documented on the Mont-Lozère in southern France. The petrographic analysis shows that slags are metallurgical wastes (800 to 850 yr BP) which certainly result from smelting activity for lead and silver extraction (Ploquin et al., 2001). The aims of this study are: 1) to trace the source of Pb ores which supplied the smelting sites, by using the Pb isotopic composition of several surrounding Pb deposits, 2) to evaluate the actual pollution caused by these slags, by using elemental and isotopic compositions of soils, water and vegetation, and 3) to document the pollution history of the region, by using elemental and isotopic compositions of peat bog cores collected in the neighbourhood of the historical smelting sites. The lead isotopic composition of galena collected in most surrounding ores is very similar to that of different slag samples. On the other hand, the high precision of the results allowed us to select the mineralised areas which were probably the ore sources. The Pb isotopic composition of slags is even more homogeneous: 208/206 Pb: 2.092±0.002; 206/207 Pb: 1.179±0.001; 208/204 Pb: 38.663±0.025; 207/204 Pb: 15.665±0.006; 206/204 Pb: 18.476±0.023, and will allow source tracing in the environment. The "Narses Mortes" peat bog, around which two smelting sites have been reported, is strongly minerotrophic and contains 8 to 60% ash. A 1.40 m core have been retrieved and divided into 58 individual samples. Minerotrophic peat bog records both atmospheric deposition, soils leaching and the grounwater influence. The measured metal concentrations are normalised to Al contents of peat bog samples and the metal/Al ratios are compared to that of the Mont-Lozère granite: relative excess in metal concentrations are found in peat bog samples. An increasing excess of most metals (Pb, Zn, Cd...) was measured for surface samples, from 55 cm depth to the top of the core (23 cm depth). This profil might be attributed to atmospheric

Brownian aggregation rate of colloid particles with several active sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nekrasov, Vyacheslav M.; Yurkin, Maxim A.; Chernyshev, Andrei V., E-mail: chern@ns.kinetics.nsc.ru

2014-08-14

We theoretically analyze the aggregation kinetics of colloid particles with several active sites. Such particles (so-called “patchy particles”) are well known as chemically anisotropic reactants, but the corresponding rate constant of their aggregation has not yet been established in a convenient analytical form. Using kinematic approximation for the diffusion problem, we derived an analytical formula for the diffusion-controlled reaction rate constant between two colloid particles (or clusters) with several small active sites under the following assumptions: the relative translational motion is Brownian diffusion, and the isotropic stochastic reorientation of each particle is Markovian and arbitrarily correlated. This formula was shownmore » to produce accurate results in comparison with more sophisticated approaches. Also, to account for the case of a low number of active sites per particle we used Monte Carlo stochastic algorithm based on Gillespie method. Simulations showed that such discrete model is required when this number is less than 10. Finally, we applied the developed approach to the simulation of immunoagglutination, assuming that the formed clusters have fractal structure.« less

Airborne measurement of submicron aerosol number concentration and CCN activity in and around the Korean Peninsula and their comparison to ground measurement in Seoul

NASA Astrophysics Data System (ADS)

Park, M.; Kim, N.; Yum, S. S.

2016-12-01

Aerosols exert impact not only on human health and visibility but also on climate change directly by scattering or absorbing solar radiation and indirectly by acting as cloud condensation nuclei (CCN) and thus altering cloud radiative and microphysical properties. Aerosol indirect effects on climate has been known to have large uncertainty because of insufficient measurement data on aerosol and CCN activity distribution. Submicron aerosol number concentration (NCN, TSI CPC) and CCN number concentration (NCCN, DMT CCNC) were measured on board the NASA DC-8 research aircraft and at a ground site at Olympic Park in Seoul from May 2nd to June 10th, 2016. CCNC on the airborne platform was operated with the fixed internal supersaturation of 0.6% and CCNC at the ground site was operated with the five different supersaturations (0.2%, 0.4%, 0.6%, 0.8%, and 1.0%). The NASA DC-8 conducted 20 research flights (about 150 hours) in and around the Korean Peninsula and the ground measurement at Olympic Park was continuously made during the measurement period. Both airborne and ground measurements showed spatially and temporally varied aerosol number concentration and CCN activity. Aerosol number concentration in the boundary layer measured on airborne platform was highly affected by pollution sources on the ground. The average diurnal distribution of ground aerosol number concentration showed distinct peaks are located at about 0800, 1500, and 2000. The middle peak indicates that new particle formation events frequently occurred during the measurement period. CCN activation ratio at 0.6% supersaturation (NCCN/NCN) of the airborne measurement ranged from 0.1 to 0.9, indicating that aerosol properties in and around the Korean Peninsula varied so much (e. g. size, hygroscopicity). Comprehensive analysis results will be shown at the conference.

Stepwise Loop Insertion Strategy for Active Site Remodeling to Generate Novel Enzyme Functions.

PubMed

Hoque, Md Anarul; Zhang, Yong; Chen, Liuqing; Yang, Guangyu; Khatun, Mst Afroza; Chen, Haifeng; Hao, Liu; Feng, Yan

2017-05-19

The remodeling of active sites to generate novel biocatalysts is an attractive and challenging task. We developed a stepwise loop insertion strategy (StLois), in which randomized residue pairs are inserted into active site loops. The phosphotriesterase-like lactonase from Geobacillus kaustophilus (GkaP-PLL) was used to investigate StLois's potential for changing enzyme function. By inserting six residues into active site loop 7, the best variant ML7-B6 demonstrated a 16-fold further increase in catalytic efficiency toward ethyl-paraoxon compared with its initial template, that is a 609-fold higher, >10 7 fold substrate specificity shift relative to that of wild-type lactonase. The remodeled variants displayed 760-fold greater organophosphate hydrolysis activity toward the organophosphates parathion, diazinon, and chlorpyrifos. Structure and docking computations support the source of notably inverted enzyme specificity. Considering the fundamental importance of active site loops, the strategy has potential for the rapid generation of novel enzyme functions by loop remodeling.

Orthogonal use of a human tRNA synthetase active site to achieve multifunctionality.

PubMed

Zhou, Quansheng; Kapoor, Mili; Guo, Min; Belani, Rajesh; Xu, Xiaoling; Kiosses, William B; Hanan, Melanie; Park, Chulho; Armour, Eva; Do, Minh-Ha; Nangle, Leslie A; Schimmel, Paul; Yang, Xiang-Lei

2010-01-01

Protein multifunctionality is an emerging explanation for the complexity of higher organisms. In this regard, aminoacyl tRNA synthetases catalyze amino acid activation for protein synthesis, but some also act in pathways for inflammation, angiogenesis and apoptosis. It is unclear how these multiple functions evolved and how they relate to the active site. Here structural modeling analysis, mutagenesis and cell-based functional studies show that the potent angiostatic, natural fragment of human tryptophanyl-tRNA synthetase (TrpRS) associates via tryptophan side chains that protrude from its cognate cellular receptor vascular endothelial cadherin (VE-cadherin). VE-cadherin's tryptophan side chains fit into the tryptophan-specific active site of the synthetase. Thus, specific side chains of the receptor mimic amino acid substrates and expand the functionality of the active site of the synthetase. We propose that orthogonal use of the same active site may be a general way to develop multifunctionality of human tRNA synthetases and other proteins.

Intramuscular pressure and torque during isometric, concentric and eccentric muscular activity

NASA Technical Reports Server (NTRS)

Styf, J.; Ballard, R.; Aratow, M.; Crenshaw, A.; Watenpaugh, D.; Hargens, A. R.

1995-01-01

Intramuscular pressures, electromyography (EMG) and torque generation during isometric, concentric and eccentric maximal isokinetic muscle activity were recorded in 10 healthy volunteers. Pressure and EMG activity were continuously and simultaneously measured side by side in the tibialis anterior and soleus muscles. Ankle joint torque and position were monitored continuously by an isokinetic dynamometer during plantar flexion and dorsiflexion of the foot. The increased force generation during eccentric muscular activity, compared with other muscular activity, was not accompanied by higher intramuscular pressure. Thus, this study demonstrated that eccentric muscular activity generated higher torque values for each increment of intramuscular pressure. Intramuscular pressures during antagonistic co-activation were significantly higher in the tibilis anterior muscle (42-46% of maximal agonistic activity) compared with the soleus muscle (12-29% of maximal agonistic activity) and was largely due to active recruitment of muscle fibers. In summary, eccentric muscular activity creates higher torque values with no additional increase of the intramuscular pressure compared with concentric and isometric muscular activity.

In silico analysis of Pycnoporus cinnabarinus laccase active site with toxic industrial dyes.

PubMed

Prasad, Nirmal K; Vindal, Vaibhav; Narayana, Siva Lakshmi; Ramakrishna, V; Kunal, Swaraj Priyaranjan; Srinivas, M

2012-05-01

Laccases belong to multicopper oxidases, a widespread class of enzymes implicated in many oxidative functions in various industrial oxidative processes like production of fine chemicals to bioremediation of contaminated soil and water. In order to understand the mechanisms of substrate binding and interaction between substrates and Pycnoporus cinnabarinus laccase, a homology model was generated. The resulted model was further validated and used for docking studies with toxic industrial dyes- acid blue 74, reactive black 5 and reactive blue 19. Interactions of chemical mediators with the laccase was also examined. The docking analysis showed that the active site always cannot accommodate the dye molecules, due to constricted nature of the active site pocket and steric hindrance of the residues whereas mediators are relatively small and can easily be accommodated into the active site pocket, which, thereafter leads to the productive binding. The binding properties of these compounds along with identification of critical active site residues can be used for further site-directed mutagenesis experiments in order to identify their role in activity and substrate specificity, ultimately leading to improved mutants for degradation of these toxic compounds.

Active Iron Sites of Disordered Mesoporous Silica Catalyst FeKIL-2 in the Oxidation of Volatile Organic Compounds (VOC)

PubMed Central

Rangus, Mojca; Mazaj, Matjaž; Dražić, Goran; Popova, Margarita; Tušar, Nataša Novak

2014-01-01

Iron-functionalized disordered mesoporous silica (FeKIL-2) is a promising, environmentally friendly, cost-effective and highly efficient catalyst for the elimination of volatile organic compounds (VOCs) from polluted air via catalytic oxidation. In this study, we investigated the type of catalytically active iron sites for different iron concentrations in FeKIL-2 catalysts using advanced characterization of the local environment of iron atoms by a combination of X-ray Absorption Spectroscopy Techniques (XANES, EXAFS) and Atomic-Resolution Scanning Transmission Electron Microscopy (AR STEM). We found that the molar ratio Fe/Si ≤ 0.01 leads to the formation of stable, mostly isolated Fe3+ sites in the silica matrix, while higher iron content Fe/Si > 0.01 leads to the formation of oligonuclear iron clusters. STEM imaging and EELS techniques confirmed the existence of these clusters. Their size ranges from one to a few nanometers, and they are unevenly distributed throughout the material. The size of the clusters was also found to be similar, regardless of the nominal concentration of iron (Fe/Si = 0.02 and Fe/Si = 0.05). From the results obtained from sample characterization and model catalytic tests, we established that the enhanced activity of FeKIL-2 with the optimal Fe/Si = 0.01 ratio can be attributed to: (1) the optimal concentration of stable isolated Fe3+ in the silica support; and (2) accelerated diffusion of the reactants in disordered mesoporous silica (FeKIL-2) when compared to ordered mesoporous silica materials (FeSBA-15, FeMCM-41). PMID:28788674

Temporal and spatial variations of polycyclic aromatic hydrocarbon concentrations around a coke oven plant.

PubMed

Stella, Anna; Piccardo, Maria Teresa; Pala, Mauro; Balducci, Daniele; Cipolla, Massimo; Ceppi, Marcello; Valerio, Federico

2012-09-01

From 1995 to 2004, in Genoa, Italy, daily concentrations of twelve polycyclic aromatic hydrocarbons (PAHs) were measured in particulate phase (PM10), around a coke oven plant in operation from the 1950s and closed in 2002. The study permitted to identify the coke oven as the main PAH source in Genoa, causing constant exceeding of benzo(a)pyrene (BaP) air quality target (1.0 ng/m3) in the urban area till 1,900 meters distance downwind the plant. For this reason the plant was closed. Distance and daily hours downwind the coke plant were the main sources of variability of toxic BaP equivalent (BaPeq) concentrations and equations that best fitted these variables were experimentally obtained. During full plant activity, annual average BaPeq concentrations, measured in the three sampling sites aligned downwind to the summer prevalent winds, were: 85 ng/m3 at 40 m (site 2, industrial area), 13.2 ng/m3 at 300 m (site 3, residential area) and 5.6 ng/m3 at 575 m (site 4, residential area). Soon after the coke oven's closure (February 2002) BaPeq concentrations (annual average) measured in residential area, decreased drastically: 0.2 ng/m3 at site 3, 0.4 ng/m3 at site 4. Comparing 1998 and 2003 data, BaPeq concentrations decreased 97.6% in site 3 and 92.8% in site 4. Samples collected at site 3, during the longest downwind conditions, provided a reliable PAH profile of fugitive coke oven emissions. This profile was significantly different from the PAH profile, contemporary found at site 5, near the traffic flow. This study demonstrates that risk assessment based only on distance of residences from a coke plant can be heavily inaccurate and confirmed that seasonal variability of BaPeq concentrations and high variability of fugitive emissions of PAHs during coke oven activities require at least one year of frequent and constant monitoring (10-15 samples each month). Around a coking plant, polycyclic aromatic hydrocarbons (PAHs), concentrations depend mainly on downwind hours

Thermodynamic compensation upon binding to exosite 1 and the active site of thrombin

PubMed Central

Treuheit, Nicholas A.; Beach, Muneera A.; Komives, Elizabeth A.

2011-01-01

Several lines of experimental evidence including amide exchange and NMR suggest that ligands binding to thrombin cause reduced backbone dynamics. Binding of the covalent inhibitor dPhe-Pro-Arg chloromethylketone to the active site serine, as well as non-covalent binding of a fragment of the regulatory protein, thrombomodulin, to exosite 1 on the back side of the thrombin molecule both cause reduced dynamics. However, the reduced dynamics do not appear to be accompanied by significant conformational changes. In addition, binding of ligands to the active site does not change the affinity of thrombomodulin fragments binding to exosite 1, however, the thermodynamic coupling between exosite 1 and the active site has not been fully explored. We present isothermal titration calorimetry experiments that probe changes in enthalpy and entropy upon formation of binary ligand complexes. The approach relies on stringent thrombin preparation methods and on the use of dansyl-L-arginine-(3-methyl-1,5-pantanediyl) amide and a DNA aptamer as ligands with ideal thermodynamic signatures for binding to the active site and to exosite 1. Using this approach, the binding thermodynamic signatures of each ligand alone as well as the binding signatures of each ligand when the other binding site was occupied were measured. Different exosite 1 ligands with widely varied thermodynamic signatures cause the same reduction in ΔH and a concomitantly lower entropy cost upon DAPA binding at the active site. The results suggest a general phenomenon of enthalpy-entropy compensation consistent with reduction of dynamics/increased folding of thrombin upon ligand binding to either the active site or to exosite 1. PMID:21526769

Thermodynamic compensation upon binding to exosite 1 and the active site of thrombin.

PubMed

Treuheit, Nicholas A; Beach, Muneera A; Komives, Elizabeth A

2011-05-31

Several lines of experimental evidence including amide exchange and NMR suggest that ligands binding to thrombin cause reduced backbone dynamics. Binding of the covalent inhibitor dPhe-Pro-Arg chloromethyl ketone to the active site serine, as well as noncovalent binding of a fragment of the regulatory protein, thrombomodulin, to exosite 1 on the back side of the thrombin molecule both cause reduced dynamics. However, the reduced dynamics do not appear to be accompanied by significant conformational changes. In addition, binding of ligands to the active site does not change the affinity of thrombomodulin fragments binding to exosite 1; however, the thermodynamic coupling between exosite 1 and the active site has not been fully explored. We present isothermal titration calorimetry experiments that probe changes in enthalpy and entropy upon formation of binary ligand complexes. The approach relies on stringent thrombin preparation methods and on the use of dansyl-l-arginine-(3-methyl-1,5-pantanediyl)amide and a DNA aptamer as ligands with ideal thermodynamic signatures for binding to the active site and to exosite 1. Using this approach, the binding thermodynamic signatures of each ligand alone as well as the binding signatures of each ligand when the other binding site was occupied were measured. Different exosite 1 ligands with widely varied thermodynamic signatures cause a similar reduction in ΔH and a concomitantly lower entropy cost upon DAPA binding at the active site. The results suggest a general phenomenon of enthalpy-entropy compensation consistent with reduction of dynamics/increased folding of thrombin upon ligand binding to either the active site or exosite 1.

Active-Site Hydration and Water Diffusion in Cytochrome P450cam: A Highly Dynamic Process

PubMed Central

Miao, Yinglong; Baudry, Jerome

2011-01-01

Long-timescale molecular dynamics simulations (300 ns) are performed on both the apo- (i.e., camphor-free) and camphor-bound cytochrome P450cam (CYP101). Water diffusion into and out of the protein active site is observed without biased sampling methods. During the course of the molecular dynamics simulation, an average of 6.4 water molecules is observed in the camphor-binding site of the apo form, compared to zero water molecules in the binding site of the substrate-bound form, in agreement with the number of water molecules observed in crystal structures of the same species. However, as many as 12 water molecules can be present at a given time in the camphor-binding region of the active site in the case of apo-P450cam, revealing a highly dynamic process for hydration of the protein active site, with water molecules exchanging rapidly with the bulk solvent. Water molecules are also found to exchange locations frequently inside the active site, preferentially clustering in regions surrounding the water molecules observed in the crystal structure. Potential-of-mean-force calculations identify thermodynamically favored trans-protein pathways for the diffusion of water molecules between the protein active site and the bulk solvent. Binding of camphor in the active site modifies the free-energy landscape of P450cam channels toward favoring the diffusion of water molecules out of the protein active site. PMID:21943431

Salicylic Acid and Jasmonic Acid Pathways are Activated in Spatially Different Domains Around the Infection Site During Effector-Triggered Immunity in Arabidopsis thaliana.

PubMed

Betsuyaku, Shigeyuki; Katou, Shinpei; Takebayashi, Yumiko; Sakakibara, Hitoshi; Nomura, Nobuhiko; Fukuda, Hiroo

2018-01-01

The innate immune response is, in the first place, elicited at the site of infection. Thus, the host response can be different among the infected cells and the cells surrounding them. Effector-triggered immunity (ETI), a form of innate immunity in plants, is triggered by specific recognition between pathogen effectors and their corresponding plant cytosolic immune receptors, resulting in rapid localized cell death known as hypersensitive response (HR). HR cell death is usually limited to a few cells at the infection site, and is surrounded by a few layers of cells massively expressing defense genes such as Pathogenesis-Related Gene 1 (PR1). This virtually concentric pattern of the cellular responses in ETI is proposed to be regulated by a concentration gradient of salicylic acid (SA), a phytohormone accumulated around the infection site. Recent studies demonstrated that jasmonic acid (JA), another phytohormone known to be mutually antagonistic to SA in many cases, is also accumulated in and required for ETI, suggesting that ETI is a unique case. However, the molecular basis for this uniqueness remained largely to be solved. Here, we found that, using intravital time-lapse imaging, the JA signaling pathway is activated in the cells surrounding the central SA-active cells around the infection sites in Arabidopsis thaliana. This distinct spatial organization explains how these two phythormone pathways in a mutually antagonistic relationship can be activated simultaneously during ETI. Our results re-emphasize that the spatial consideration is a key strategy to gain mechanistic insights into the apparently complex signaling cross-talk in immunity. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

Active Site Mutations Change the Cleavage Specificity of Neprilysin

PubMed Central

Sexton, Travis; Hitchcook, Lisa J.; Rodgers, David W.; Bradley, Luke H.; Hersh, Louis B.

2012-01-01

Neprilysin (NEP), a member of the M13 subgroup of the zinc-dependent endopeptidase family is a membrane bound peptidase capable of cleaving a variety of physiological peptides. We have generated a series of neprilysin variants containing mutations at either one of two active site residues, Phe563 and Ser546. Among the mutants studied in detail we observed changes in their activity towards leucine5-enkephalin, insulin B chain, and amyloid β1–40. For example, NEPF563I displayed an increase in preference towards cleaving leucine5-enkephalin relative to insulin B chain, while mutant NEPS546E was less discriminating than neprilysin. Mutants NEPF563L and NEPS546E exhibit different cleavage site preferences than neprilysin with insulin B chain and amyloid ß1–40 as substrates. These data indicate that it is possible to alter the cleavage site specificity of neprilysin opening the way for the development of substrate specific or substrate exclusive forms of the enzyme with enhanced therapeutic potential. PMID:22384224

Carbinolamine Formation and Dehydration in a DNA Repair Enzyme Active Site

PubMed Central

Dodson, M. L.; Walker, Ross C.; Lloyd, R. Stephen

2012-01-01

In order to suggest detailed mechanistic hypotheses for the formation and dehydration of a key carbinolamine intermediate in the T4 pyrimidine dimer glycosylase (T4PDG) reaction, we have investigated these reactions using steered molecular dynamics with a coupled quantum mechanics–molecular mechanics potential (QM/MM). We carried out simulations of DNA abasic site carbinolamine formation with and without a water molecule restrained to remain within the active site quantum region. We recovered potentials of mean force (PMF) from thirty replicate reaction trajectories using Jarzynski averaging. We demonstrated feasible pathways involving water, as well as those independent of water participation. The water–independent enzyme–catalyzed reaction had a bias–corrected Jarzynski–average barrier height of approximately for the carbinolamine formation reaction and ) for the reverse reaction at this level of representation. When the proton transfer was facilitated with an intrinsic quantum water, the barrier height was approximately in the forward (formation) reaction and for the reverse. In addition, two modes of unsteered (free dynamics) carbinolamine dehydration were observed: in one, the quantum water participated as an intermediate proton transfer species, and in the other, the active site protonated glutamate hydrogen was directly transferred to the carbinolamine oxygen. Water–independent unforced proton transfer from the protonated active site glutamate carboxyl to the unprotonated N–terminal amine was also observed. In summary, complex proton transfer events, some involving water intermediates, were studied in QM/MM simulations of T4PDG bound to a DNA abasic site. Imine carbinolamine formation was characterized using steered QM/MM molecular dynamics. Dehydration of the carbinolamine intermediate to form the final imine product was observed in free, unsteered, QM/MM dynamics simulations, as was unforced acid-base transfer between the active site

Immobilization of alkaline phosphatase on solid surface through self-assembled monolayer and by active-site protection.

PubMed

Gao, En-Feng; Kang, Kyung Lhi; Kim, Jeong Hee

2014-06-01

Retaining biological activity of a protein after immobilization is an important issue and many studies reported to enhance the activity of proteins after immobilization. We recently developed a new immobilization method of enzyme using active-site protection and minimization of the cross-links between enzyme and surface with a DNA polymerase as a model system. In this study, we extended the new method to an enzyme with a small mono-substrate using alkaline phosphatase (AP) as another model system. A condition to apply the new method is that masking agents, in this case its own substrate needs to stay at the active-site of the enzyme to be immobilized in order to protect the active-site during the harsh immobilization process. This could be achieved by removal of essential divalent ion, Zn2+ that is required for full enzyme activity of AP from the masking solution while active-site of AP was protected with p-nitrophenyl phosphate (pNPP). Approximately 40% of the solution-phase activity was acquired with active-site protected immobilized AP. In addition to protection active-site of AP, the number of immobilization links was kinetically controlled. When the mole fraction of the activated carboxyl group of the linker molecule in self-assembled monolayer (SAM) of 12-mercaptododecanoic acid and 6-mercapto-1-ethanol was varied, 10% of 12-mercaptododecanoic acid gave the maximum enzyme activity. Approximately 51% increase in enzyme activity of the active-site protected AP was observed compared to that of the unprotected group. It was shown that the concept of active-site protection and kinetic control of the number of covalent immobilization bonds can be extended to enzymes with small mono-substrates. It opens the possibility of further extension of the new methods of active-site protection and kinetic control of immobilization bond to important enzymes used in research and industrial fields.

Mercury concentrations in water and mercury and selenium concentrations in fish from Brownlee Reservoir and selected sites in the Boise and Snake Rivers, Idaho and Oregon, 2013–15

USGS Publications Warehouse

Williams, Marshall L.; MacCoy, Dorene E.

2016-06-30

Mercury (Hg) analyses were conducted on samples of sport fish and water collected from selected sampling sites in Brownlee Reservoir and the Boise and Snake Rivers to meet National Pollution Discharge and Elimination System (NPDES) permit requirements for the City of Boise, Idaho, between 2013 and 2015. City of Boise personnel collected water samples from six sites between October and November 2013 and 2015, with one site sampled in 2014. Total Hg concentrations in unfiltered water samples ranged from 0.48 to 8.8 nanograms per liter (ng/L), with the highest value in Brownlee Reservoir in 2013. All Hg concentrations in water samples were less than the U.S. Environmental Protection Agency (USEPA) Hg chronic aquatic life criterion of 12 ng/L.The USEPA recommended a water-quality criterion of 0.30 milligrams per kilogram (mg/kg) methylmercury (MeHg) expressed as a fish-tissue residue value (wet-weight MeHg in fish tissue). The Idaho Department of Environmental Quality adopted the USEPA’s fish-tissue criterion and established a reasonable potential to exceed (RPTE) threshold 20 percent lower than the criterion or greater than 0.24 mg/kg Hg based on an average concentration of 10 fish from a receiving waterbody. NPDES permitted discharge to waters with fish having Hg concentrations exceeding 0.24 mg/kg are said to have a reasonable potential to exceed the water-quality criterion and thus are subject to additional permit obligations, such as requirements for increased monitoring and the development of a Hg minimization plan. The Idaho Fish Consumption Advisory Program (IFCAP) issues fish advisories to protect general and sensitive populations of fish consumers and has developed an action level of 0.22 mg/kg Hg in fish tissue. Fish consumption advisories are water body- and species-specific and are used to advise allowable fish consumption from specific water bodies. The geometric mean Hg concentration of 10 fish of a single species collected from a single water body

Ultrafast Light-Driven Substrate Expulsion from the Active Site of a Photoswitchable Catalyst.

PubMed

Pescher, Manuel D; van Wilderen, Luuk J G W; Grützner, Susanne; Slavov, Chavdar; Wachtveitl, Josef; Hecht, Stefan; Bredenbeck, Jens

2017-09-25

The photoswitchable piperidine general base catalyst is a prototype structure for light control of catalysis. Its azobenzene moiety moves sterically shielding groups to either protect or expose the active site, thereby changing the basicity and hydrogen-bonding affinity of the compound. The reversible switching dynamics of the catalyst is probed in the infrared spectral range by monitoring hydrogen bond (HB) formation between its active site and methanol (MeOH) as HB donor. Steady-state infrared (IR) and ultrafast IR and UV/Vis spectroscopies are used to uncover ultrafast expulsion of MeOH from the active site within a few picoseconds. Thus, the force generated by the azobenzene moiety even in the final phase of its isomerization is sufficient to break a strong HB within 3 ps and to shut down access to the active site. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Dynamic Perturbation of the Active Site Determines Reversible Thermal Inactivation in Glycoside Hydrolase Family 12.

PubMed

Jiang, Xukai; Li, Wen; Chen, Guanjun; Wang, Lushan

2017-02-27

The temperature dependence of enzyme catalysis is highly debated. Specifically, how high temperatures induce enzyme inactivation has broad implications for both fundamental and applied science. Here, we explored the mechanism of the reversible thermal inactivation in glycoside hydrolase family 12 (GH12) using comparative molecular dynamics simulations. First, we investigated the distribution of structural flexibility over the enzyme and found that the active site was the general thermal-sensitive region in GH12 cellulases. The dynamic perturbation of the active site before enzyme denaturation was explored through principal-component analysis, which indicated that variations in the collective motion and conformational ensemble of the active site may precisely correspond to enzyme transition from its active form to the inactive form. Furthermore, the degree of dynamic perturbation of the active site was found to be negatively correlated with the melting temperatures of GH12 enzymes, further proving the importance of the dynamic stability of the active site. Additionally, analysis of the residue-interaction network revealed that the active site in thermophilic enzyme was capable of forming additional contacts with other amino acids than those observed in the mesophilic enzyme. These interactions are likely the key mechanisms underlying the differences in rigidity of the active site. These findings provide further biophysical insights into the reversible thermal inactivation of enzymes and potential applications in future protein engineering.

Seasonal and spatial trends in particle number concentrations and size distributions at the children's health study sites in Southern California.

PubMed

Singh, Manisha; Phuleria, Harish C; Bowers, Kenneth; Sioutas, Constantinos

2006-01-01

Continuous measurements of particle number (PN), particle mass (PM(10)) and gaseous copollutants (NO(x), CO and O3) were obtained at eight sites (urban, suburban and remote) in Southern California during years 2002 and 2003 in support of University of Southern California Children's Health Study. We report the spatial and temporal variation of PNs and size distributions within these sites. Higher average total PN concentrations are found in winter (November to February), compared to summer (July to September) and spring (March to June) in all urban sites. Contribution of local vehicular emissions is most evident in cooler months, whereas effects of long-range transport of particles are enhanced during warmer periods. The particle size profile is most represented by a combination of the spatial effects, for example, sources, atmospheric processes and meteorological conditions prevalent at each location. Afternoon periods in the warmer months are characterized by elevated number concentrations that either coincide or follow a peak in ozone concentrations, suggesting the formation of new particles by photochemistry. Results show no meaningful correlation between PN and mass, indicating that mass based standards may not be effective in controlling ultrafine particles. The study of the impact of the Union worker's strike at port of Long Beach in October 2002 revealed statistically significant increase in PN concentrations in the 60-200 nm range (P<0.001), which are indicative of contributions of emissions from the idling ships at the port.

Refining the site conceptual model at a former uranium mill site in Riverton, Wyoming, USA

DOE PAGES

Dam, William; Campbell, Sam; Johnson, Ray; ...

2015-07-07

Milling activities at a former uranium mill site near Riverton, Wyoming, USA, contaminated the shallow groundwater beneath and downgradient of the site. Although the mill operated for <6 years (1958-1963), its impact remains an environmental liability. Groundwater modeling predicted that contaminant concentrations were declining steadily, which confirmed the conceptual site model (CSM). However, local flooding in 2010 mobilized contaminants that migrated downgradient from the Riverton site and resulted in a dramatic increase in groundwater contaminant concentrations. This observation indicated that the original CSM was inadequate to explain site conditions and needed to be refined. In response to the new observationsmore » after the flood, a collaborative investigation to better understand site conditions and processes commenced. This investigation included installing 103 boreholes to collect soil and groundwater samples, sampling and analysis of evaporite minerals along the bank of the Little Wind River, an analysis of evaportranspiration in the shallow aquifer, and sampling naturally organic-rich sediments near groundwater discharge areas. The enhanced characterization revealed that the existing CSM did not account for high uranium concentrations in groundwater remaining on the former mill site and groundwater plume stagnation near the Little Wind River. Observations from the flood and subsequent investigations indicate that additional characterization is still needed to continue refining the CSM and determine the viability of the natural flushing compliance strategy. Additional sampling, analysis, and testing of soil and groundwater are necessary to investigate secondary contaminant sources, mobilization of contaminants during floods, geochemical processes, contaminant plume stagnation, distribution of evaporite minerals and organic-rich sediments, and mechanisms and rates of contaminant transfer from soil to groundwater. Future data collection will be used to

Airborne metal concentrations in shipyard environment.

PubMed

Alebić-Juretić, A; Matković, N

2000-06-01

Protection against corrosion in the shipyard is a source of airborne particles. From October 1996 to September 1997 samples of suspended particles (1 site) and dustfall (6 sites) were collected in the vicinity of a repairs shipyard situated in the martinsćica Cove, east of the city of Rijeka, Croatia. Collected samples were analysed for lead, cadmium, iron, copper, and zinc content. Though annual mean concentrations of suspended particles, lead, and cadmium kept below the guideline values, the metal contents were generally higher than values measured in the city centre. The correlation between the quantity of abrasives used at the shipyard and monthly mean concentrations of all parameters except cadmium suggests that the shipyard was the main source of those pollutants. The annual mean, as well as maximum monthly amount of dustfall at the site next to the shipyard zone exceeded the national limit values, indicating considerable pollution of this area with coarse particles. The annual mean quantity of lead in dustfall exceeded the guideline values at the same site. The content of metals occasionally observed in dustfall at particular sites surrounding the shipyard depended on the location of corrosion protection activities and meterological conditions within the Martinsćica Cove.

Non-active site mutation (Q123A) in New Delhi metallo-β-lactamase (NDM-1) enhanced its enzyme activity.

PubMed

Ali, Abid; Azam, Mohd W; Khan, Asad U

2018-06-01

New Delhi metallo β-lactamase-1 is one of the carbapenemases, causing hydrolysis of almost all β-lactamase antibiotics. Seventeen different NDM variants have been reported so far, they varied in their sequences either by single or multiple amino acid substitutions. Hence, it is important to understand its structural and functional relation. In the earlier studies role of active site residues has been studied but non-active site residues has not studied in detail. Therefore, we have initiated to further comprehend its structure and function relation by mutating some of its non-active site residues. A laboratory mutant of NDM-1 was generated by PCR-based site-directed mutagenesis, replacing Q to A at 123 position. The MICs of imipenem and meropenem for NDM-1 Q123A were found increased by 2 fold as compare to wild type and so the hydrolytic activity was enhanced (Kcat/Km) as compared to NDM-1 wild type. GOLD fitness scores were also found in favour of kinetics data. Secondary structure for α-helical content was determined by Far-UV circular dichroism (CD), which showed significant conformational changes. We conclude a noteworthy role of non-active-site amino acid residues in the catalytic activity of NDM-1. This study also provides an insight of emergence of new variants through natural evolution. Copyright © 2018 Elsevier B.V. All rights reserved.

pH-Dependent Binding of Chloride to a Marine Alkaline Phosphatase Affects the Catalysis, Active Site Stability, and Dimer Equilibrium.

PubMed

Hjörleifsson, Jens G; Ásgeirsson, Bjarni

2017-09-26

The effect of ionic strength on enzyme activity and stability varies considerably between enzymes. Ionic strength is known to affect the catalytic activity of some alkaline phosphatases (APs), such as Escherichia coli AP, but how ions affect APs is debated. Here, we studied the effect of various ions on a cold-adapted AP from Vibrio splendidus (VAP). Previously, we have found that the active form of VAP is extremely unstable at low ionic strengths. Here we show that NaCl increased the activity and stability of VAP and that the effect was pH-dependent in the range of pH 7-10. The activity profile as a function of pH formed two maxima, indicating a possible conformational change. Bringing the pH from the neutral to the alkaline range was accompanied by a large increase in both the K i for inorganic phosphate (product inhibition) and the K M for p-nitrophenyl phosphate. The activity transitions observed as the pH was varied correlated with structural changes as monitored by tryptophan fluorescence. Thermal and urea-induced inactivation was shown to be accompanied by neither dissociation of the active site metal ions nor dimer dissociation. This would suggest that the inactivation involved subtle changes in active site conformation. Furthermore, the VAP dimer equilibrium was studied for the first time and shown to highly favor dimerization, which was dependent on pH and NaCl concentration. Taken together, the data support a model in which anions bind to some specific acceptor in the active site of VAP, resulting in great stabilization and catalytic rate enhancement, presumably through a different mechanism.

Prediction of suspended-sediment concentrations at selected sites in the Fountain Creek watershed, Colorado, 2008-09

USGS Publications Warehouse

Stogner, Sr., Robert W.; Nelson, Jonathan M.; McDonald, Richard R.; Kinzel, Paul J.; Mau, David P.

2013-01-01

In 2008, the U.S. Geological Survey (USGS), in cooperation with Pikes Peak Area Council of Governments, Colorado Water Conservation Board, Colorado Springs City Engineering, and the Lower Arkansas Valley Water Conservancy District, began a small-scale pilot study to evaluate the effectiveness of the use of a computational model of streamflow and suspended-sediment transport for predicting suspended-sediment concentrations and loads in the Fountain Creek watershed in Colorado. Increased erosion and sedimentation damage have been identified by the Fountain Creek Watershed Plan as key problems within the watershed. A recommendation in the Fountain Creek Watershed plan for management of the basin is to establish measurable criteria to determine if progress in reducing erosion and sedimentation damage is being made. The major objective of this study was to test a computational method to predict local suspended-sediment loads at two sites with different geomorphic characteristics in order to evaluate the feasibility of using such an approach to predict local suspended-sediment loads throughout the entire watershed. Detailed topographic surveys, particle-size data, and suspended-sediment samples were collected at two gaged sites: Monument Creek above Woodmen Road at Colorado Springs, Colorado (USGS gage 07103970), and Sand Creek above mouth at Colorado Springs, Colorado (USGS gage 07105600). These data were used to construct three-dimensional computational models of relatively short channel reaches at each site. The streamflow component of these models predicted a spatially distributed field of water-surface elevation, water velocity, and bed shear stress for a range of stream discharges. Using the model predictions, along with measured particle sizes, the sediment-transport component of the model predicted the suspended-sediment concentration throughout the reach of interest. These computed concentrations were used with predicted flow patterns and channel morphology to

Multiple active site residues are important for photochemical efficiency in the light-activated enzyme protochlorophyllide oxidoreductase (POR).

PubMed

Menon, Binuraj R K; Hardman, Samantha J O; Scrutton, Nigel S; Heyes, Derren J

2016-08-01

Protochlorophyllide oxidoreductase (POR) catalyzes the light-driven reduction of protochlorophyllide (Pchlide), an essential, regulatory step in chlorophyll biosynthesis. The unique requirement of the enzyme for light has provided the opportunity to investigate how light energy can be harnessed to power biological catalysis and enzyme dynamics. Excited state interactions between the Pchlide molecule and the protein are known to drive the subsequent reaction chemistry. However, the structural features of POR and active site residues that are important for photochemistry and catalysis are currently unknown, because there is no crystal structure for POR. Here, we have used static and time-resolved spectroscopic measurements of a number of active site variants to study the role of a number of residues, which are located in the proposed NADPH/Pchlide binding site based on previous homology models, in the reaction mechanism of POR. Our findings, which are interpreted in the context of a new improved structural model, have identified several residues that are predicted to interact with the coenzyme or substrate. Several of the POR variants have a profound effect on the photochemistry, suggesting that multiple residues are important in stabilizing the excited state required for catalysis. Our work offers insight into how the POR active site geometry is finely tuned by multiple active site residues to support enzyme-mediated photochemistry and reduction of Pchlide, both of which are crucial to the existence of life on Earth. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Effective method of continuous rocuronium administration based on effect-site concentrations using a pharmacokinetic/pharmacodynamic model during propofol-remifentanil anesthesia.

PubMed

Moriyama, Takahiro; Matsunaga, Akira; Nagata, Osamu; Enohata, Kei; Kamikawaji, Tomomi; Uchino, Erika; Kanmura, Yuichi

2015-08-01

Rocuronium bromide (Rb) is a rapid onset, intermediate-acting neuromuscular blocking agent that is suitable for continuous administration. The appropriate rate of rocuronium administration is, however, difficult to determine due to large interindividual differences in sensitivity to rocuronium. The aim of this study was to clarify whether the simulated rocuronium concentration at the time of recovery to %T1 > 0 % after the initial administration of rocuronium is a good indicator of optimal effect-site concentrations during continuous rocuronium administration. Twenty-one patients were anesthetized with propofol. After induction, Rb 0.6 mg/kg was administered intravenously, and nerve stimulation using the single stimulation mode was conducted every 15 s. When %T1 recovered to >0 % after the initial administration of Rb, the effect-site concentration of rocuronium, calculated by pharmacokinetic simulation with Wierda's set of parameters, was recorded and defined as the recovery concentration (Rb r.c.). The administration rate of rocuronium was adjusted to maintain the Rb r.c. during surgery. Rb administration was discontinued just before the end of surgery, and the recovery time until %T1 > 25 % was recorded. Plasma Rb concentrations were measured at 1 and 3 h after the initiation of continuous Rb administration. The mean Rb r.c. was 1.56 ± 0.35 μg/ml, with minimum and maximum values of 1.09 and 2.08 μg/ml, respectively. The %T1 did not increase above 10 % in any of the patients during continuous administration of Rb, and the recovery period to %T1 > 25 % ranged from 9 to 29 min. The effect-site concentrations of Rb calculated with Wierda's parameters significantly correlated with plasma concentrations (P < 0.01) at both 1 and 3 h after the initial administration of Rb. The results suggest that our method may be one of the most reliable protocols for the continuous administration of Rb described to date for maintaining suitable muscle relaxation during surgery

Ginseng gintonin activates the human cardiac delayed rectifier K+ channel: involvement of Ca2+/calmodulin binding sites.

PubMed

Choi, Sun-Hye; Lee, Byung-Hwan; Kim, Hyeon-Joong; Jung, Seok-Won; Kim, Hyun-Sook; Shin, Ho-Chul; Lee, Jun-Hee; Kim, Hyoung-Chun; Rhim, Hyewhon; Hwang, Sung-Hee; Ha, Tal Soo; Kim, Hyun-Ji; Cho, Hana; Nah, Seung-Yeol

2014-09-01

Gintonin, a novel, ginseng-derived G protein-coupled lysophosphatidic acid (LPA) receptor ligand, elicits [Ca(2+)]i transients in neuronal and non-neuronal cells via pertussis toxin-sensitive and pertussis toxin-insensitive G proteins. The slowly activating delayed rectifier K(+) (I(Ks)) channel is a cardiac K(+) channel composed of KCNQ1 and KCNE1 subunits. The C terminus of the KCNQ1 channel protein has two calmodulin-binding sites that are involved in regulating I(Ks) channels. In this study, we investigated the molecular mechanisms of gintonin-mediated activation of human I(Ks) channel activity by expressing human I(Ks) channels in Xenopus oocytes. We found that gintonin enhances IKs channel currents in concentration- and voltage-dependent manners. The EC50 for the I(Ks) channel was 0.05 ± 0.01 μg/ml. Gintonin-mediated activation of the I(Ks) channels was blocked by an LPA1/3 receptor antagonist, an active phospholipase C inhibitor, an IP3 receptor antagonist, and the calcium chelator BAPTA. Gintonin-mediated activation of both the I(Ks) channel was also blocked by the calmodulin (CaM) blocker calmidazolium. Mutations in the KCNQ1 [Ca(2+)]i/CaM-binding IQ motif sites (S373P, W392R, or R539W)blocked the action of gintonin on I(Ks) channel. However, gintonin had no effect on hERG K(+) channel activity. These results show that gintonin-mediated enhancement of I(Ks) channel currents is achieved through binding of the [Ca(2+)]i/CaM complex to the C terminus of KCNQ1 subunit.

Heterogeneous Electrocatalyst with Molecular Cobalt Ions Serving as the Center of Active Sites.

PubMed

Wang, Jiong; Ge, Xiaoming; Liu, Zhaolin; Thia, Larissa; Yan, Ya; Xiao, Wei; Wang, Xin

2017-02-08

Molecular Co 2+ ions were grafted onto doped graphene in a coordination environment, resulting in the formation of molecularly well-defined, highly active electrocatalytic sites at a heterogeneous interface for the oxygen evolution reaction (OER). The S dopants of graphene are suggested to be one of the binding sites and to be responsible for improving the intrinsic activity of the Co sites. The turnover frequency of such Co sites is greater than that of many Co-based nanostructures and IrO 2 catalysts. Through a series of carefully designed experiments, the pathway for the evolution of the Co cation-based molecular catalyst for the OER was further demonstrated on such a single Co-ion site for the first time. The Co 2+ ions were successively oxidized to Co 3+ and Co 4+ states prior to the OER. The sequential oxidation was coupled with the transfer of different numbers of protons/hydroxides and generated an active Co 4+ ═O fragment. A side-on hydroperoxo ligand of the Co 4+ site is proposed as a key intermediate for the formation of dioxygen.

Rubisco activity in Mediterranean species is regulated by the chloroplastic CO2 concentration under water stress

PubMed Central

Galmés, Jeroni; Ribas-Carbó, Miquel; Medrano, Hipólito; Flexas, Jaume

2011-01-01

Water stress decreases the availability of the gaseous substrate for ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) by decreasing leaf conductance to CO2. In spite of limiting photosynthetic carbon assimilation, especially in those environments where drought is the predominant factor affecting plant growth and yield, the effects of water deprivation on the mechanisms that control Rubisco activity are unclear. In the present study, 11 Mediterranean species, representing different growth forms, were subject to increasing levels of drought stress, the most severe one followed by rewatering. The results confirmed species-specific patterns in the decrease in the initial activity and activation state of Rubisco as drought stress and leaf dehydration intensified. Nevertheless, all species followed roughly the same trend when Rubisco activity was related to stomatal conductance (gs) and chloroplastic CO2 concentration (Cc), suggesting that deactivation of Rubisco sites could be induced by low Cc, as a result of water stress. The threshold level of Cc that triggered Rubisco deactivation was dependent on leaf characteristics and was related to the maximum attained for each species under non-stressing conditions. Those species adapted to low Cc were more capable of maintaining active Rubisco as drought stress intensified. PMID:21115663

Radiochemical Analyses of the Filter Cake, Granular Activated Carbon, and Treated Ground Water from the DTSC Stringfellow Superfund Site Pretreatment Plant

DOE Office of Scientific and Technical Information (OSTI.GOV)

Esser, B K; McConachie, W; Fischer, R

2005-09-16

The Department of Toxic Substance Control (DTSC) requested that Lawrence Livermore National Laboratory (LLNL) evaluate the treatment process currently employed at the Department's Stringfellow Superfund Site Pretreatment Plant (PTP) site to determine if wastes originating from the site were properly managed with regards to their radioactivity. In order to evaluate the current management strategy, LLNL suggested that DTSC characterize the effluents from the waste treatment system for radionuclide content. A sampling plan was developed; samples were collected and analyzed for radioactive constituents. Following is brief summary of those results and what implications for waste characterization may be made. (1) Themore » sampling and analysis provides strong evidence that the radionuclides present are Naturally Occurring Radioactive Material (NORM). (2) The greatest source of radioactivity in the samples was naturally occurring uranium. The sample results indicate that the uranium concentration in the filter cake is higher than the Granular Activated Carbon (GAC) samples. (11 -14 and 2-6 ppm respectively). (3) No radiologic background for geologic materials has been established for the Stringfellow site, and comprehensive testing of the process stream has not been conducted. Without site-specific testing of geologic materials and waste process streams, it is not possible to conclude if filter cake and spent GAC samples contain radioactivity concentrated above natural background levels, or if radionuclides are being concentrated by the waste treatment process. Recommendation: The regulation of Technologically Enhanced, Naturally Occurring Radioactive Materials (T-NORM) is complex. Since the results of this study do not conclusively demonstrate that natural radioactive materials have not been concentrated by the treatment process it is recommended that the DTSC consult with the Department of Health Services (DHS) Radiological Health Branch to determine if any further

Sublethal effects in Avocet and Stilt hatchlings from selenium-contaminated sites

USGS Publications Warehouse

Hoffman, D.J.; Marn, C.M.; Marois, Katherine C.; Sproul, E.; Dunne, M.; Skorupa, J.P.

2002-01-01

Excess selenium (Se) in the aquatic food chain is embryotoxic and teratogenic to avocets, stilts, and other waterbirds. American avocet (Recurvirostra americana) and black-necked stilt (Himantopus mexicanus) eggs were collected from three sites in the Tulare Lake Basin of California, USA, and hatched in the laboratory. These sites included the Tulare Lake Drainage District?north (TLDD-N, water 2.5 ppb Se), TLDD?south (TLDD-S, water 8.6 ppb Se), and Westfarmers (WF, water 190 ppb Se). Highest egg Se concentrations occurred at WF (geometric mean 31.4 ppm dry wt for avocets and 20.5 ppm dry wt for stilts). Mean egg Se concentrations were 6.7 ppm for avocets and 8.4 ppm for stilts at TLDD-S, and 3.3 ppm for avocets and 2.3 ppm for stilts at TLDD-N. Hatching success and incidence of malformations did not differ among sites, but yolk sac?free hatching weights and bone lengths were less for avocets at the WF site, whereas liver weights and liver to body weight ratios were greater at that site. With increasing Se concentration, oxidative stress was most apparent in avocet hatchlings from WF: hepatic glutathione (GSH) peroxidase activity increased, glucose-6-phosphate dehydrogenase activity decreased, and oxidized glutathione (GSSG) concentration as well as the ratio of GSSG to reduced GSH concentration increased. In stilts, hepatic GSH concentration was lower in WF hatchlings. In conclusion, our findings of Se-impaired embryo growth and hepatotoxicity in avocet hatchlings suggest that oxidative stress observed in hatchlings may be related to these biological effects and may serve as a potential bioindicator of subsequent impaired functions.

The Activity of Trypsin

ERIC Educational Resources Information Center

Russo, Salvatore F.; Holzman, Tom

1977-01-01

Describes an experiment that illustrates the following points concerning the experimental determination of trypsin activity: (1) there is a difference in basing enzyme concentration on weight, absorbance, or active sites; and (2) the method of expressing enzyme concentration determines the value of specific, molecular, and catalytic center…

Effect of site treatments on soil temperature and moisture and oak and pine growth and nutrient concentrations

Treesearch

Felix, Jr. Ponder

2003-01-01

Five years after planting, measurements of soil moisture and temperature, leaf nutrient concentrations and growth, were compared for plots of northern red oak, white oak, and shortleaf pine for treatment combinations that included two levels each of harvesting intensity (organic matter removal), site disturbance (soil compaction), and weed control (control of the...

Catalysis-dependent selenium incorporation and migration in the nitrogenase active site iron-molybdenum cofactor.

PubMed

Spatzal, Thomas; Perez, Kathryn A; Howard, James B; Rees, Douglas C

2015-12-16

Dinitrogen reduction in the biological nitrogen cycle is catalyzed by nitrogenase, a two-component metalloenzyme. Understanding of the transformation of the inert resting state of the active site FeMo-cofactor into an activated state capable of reducing dinitrogen remains elusive. Here we report the catalysis dependent, site-selective incorporation of selenium into the FeMo-cofactor from selenocyanate as a newly identified substrate and inhibitor. The 1.60 Å resolution structure reveals selenium occupying the S2B site of FeMo-cofactor in the Azotobacter vinelandii MoFe-protein, a position that was recently identified as the CO-binding site. The Se2B-labeled enzyme retains substrate reduction activity and marks the starting point for a crystallographic pulse-chase experiment of the active site during turnover. Through a series of crystal structures obtained at resolutions of 1.32-1.66 Å, including the CO-inhibited form of Av1-Se2B, the exchangeability of all three belt-sulfur sites is demonstrated, providing direct insights into unforeseen rearrangements of the metal center during catalysis.

Relationship between peroxisome proliferator-activated receptor alpha activity and cellular concentration of 14 perfluoroalkyl substances in HepG2 cells.

PubMed

Rosenmai, Anna Kjerstine; Ahrens, Lutz; le Godec, Théo; Lundqvist, Johan; Oskarsson, Agneta

2018-02-01

Peroxisome proliferator-activated receptor alpha (PPARα) is a molecular target for perfluoroalkyl substances (PFASs). Little is known about the cellular uptake of PFASs and how it affects the PPARα activity. We investigated the relationship between PPARα activity and cellular concentration in HepG2 cells of 14 PFASs, including perfluoroalkyl carboxylates (PFCAs), perfluoroalkyl sulfonates and perfluorooctane sulfonamide (FOSA). Cellular concentrations were determined by high-performance liquid chromatography-tandem mass spectrometry and PPARα activity was determined in transiently transfected cells by reporter gene assay. Cellular uptake of the PFASs was low (0.04-4.1%) with absolute cellular concentrations in the range 4-2500 ng mg -1 protein. Cellular concentration of PFCAs increased with perfluorocarbon chain length up to perfluorododecanoate. PPARα activity of PFCAs increased with chain length up to perfluorooctanoate. The maximum induction of PPARα activity was similar for short-chain (perfluorobutanoate and perfluoropentanoate) and long-chain PFCAs (perfluorododecanoate and perfluorotetradecanoate) (approximately twofold). However, PPARα activities were induced at lower cellular concentrations for the short-chain homologs compared to the long-chain homologs. Perfluorohexanoate, perfluoroheptanoate, perfluorooctanoate, perfluorononanoate (PFNA) and perfluorodecanoate induced PPARα activities >2.5-fold compared to controls. The concentration-response relationships were positive for all the tested compounds, except perfluorooctane sulfonate PFOS and FOSA, and were compound-specific, as demonstrated by differences in the estimated slopes. The relationships were steeper for PFCAs with chain lengths up to and including PFNA than for the other studied PFASs. To our knowledge, this is the first report establishing relationships between PPARα activity and cellular concentration of a broad range of PFASs. Copyright © 2017 John Wiley & Sons, Ltd.

Photovoltaic concentrator assembly with optically active cover

DOEpatents

Plesniak, Adam P

2014-01-21

A photovoltaic concentrator assembly that includes a housing that defines an internal volume and includes a rim, wherein the rim defines an opening into the internal volume, a photovoltaic cell positioned in the internal volume, and an optical element that includes an optically active body and a flange extending outward from the body, wherein the flange is sealingly engaged with the rim of the housing to enclose the internal volume.

Freezing-Out Technique Applied to the Concentration of Biologically Active Materials

PubMed Central

Wilson, T. E.; Evans, D. J.; Theriot, Mary L.

1964-01-01

When applied to a dilute solution of folic acid and glucose, a freezing-out (with agitation) technique was shown to be an effective method of achieving a 20-fold reduction in volume with a loss of 10% of the active material being concentrated. Concentration of a stimulatory factor for Lactobacillus casei produced by Candida albicans in a complex medium was limited by the total solute concentration. Salts in the medium were concentrated to levels inhibitory for L. casei. The process is not selective and all solutes are concentrated. Images FIG. 2 FIG. 3 PMID:14131370

Comparison of PM10 concentrations and metal content in three different sites of the Venice Lagoon: an analysis of possible aerosol sources.

PubMed

Contini, Daniele; Belosi, Franco; Gambaro, Andrea; Cesari, Daniela; Stortini, Angela Maria; Bove, Maria Chiara

2012-01-01

The Venice Lagoon is exposed to atmospheric pollutants from industrial activities, thermoelectric power plants, petrochemical plants, incinerator, domestic heating, ship traffic, glass factories and vehicular emissions on the mainland. In 2005, construction began on the mobile dams (MOSE), one dam for each channel connecting the lagoon to the Adriatic Sea as a barrier against high tide. These construction works could represent an additional source of pollutants. PM10 samples were taken on random days between 2007 and 2010 at three different sites: Punta Sabbioni, Chioggia and Malamocco, located near the respective dam construction worksites. Chemical analyses of V, Cr, Fe, Co, Ni, Cu, Zn, As, Mo, Cd, Sb, Tl and Pb in PM10 samples were performed by Inductively coupled plasma-quadrupole mass spectrometry (ICP-QMS) and results were used to identify the main aerosol sources. The correlation of measured data with meteorology, and source apportionment, failed to highlight a contribution specifically associated to the emissions of the MOSE construction works. The comparison of the measurements at the three sites showed a substantial homogeneity of metal concentrations in the area. Source apportionment with principal component analysis (PCA) and positive matrix factorization (PMF) showed that a four principal factors model could describe the sources of metals in PM10. Three of them were assigned to specific sources in the area and one was characterised as a source of mixed origin (anthropogenic and crustal). A specific anthropogenic source of PM10 rich in Ni and Cr, active at the Chioggia site, was also identified.

Application of Passive Sampling for Measuring Dissolved Concentrations of Organic Contaminants in the Water Column at Three U.S. EPA Marine Superfund Sites

EPA Science Inventory

At contaminated sediment sites, including U.S. EPA Superfund sites, it is critical to measure water column concentrations of freely dissolved contaminants to understand the complete exposure of aquatic organisms to hydrophobic organic contaminants (HOCs). Historically acquiring ...

Application of Passive Sampling for Measuring Dissolved Concentrations of Organic Contaminants in the Water Column at Three U.S. EPA Marine Superfund Sites.

EPA Science Inventory

At contaminated sediment sites, including U.S. EPA Superfund sites, it is critical to measure water column concentrations of freely dissolved contaminants to understand the complete exposure of aquatic organisms to hydrophobic organic contaminants (HOCs). Historically, acquiring...

Methylation of an alpha-foetoprotein gene intragenic site modulates gene activity.

PubMed Central

Opdecamp, K; Rivière, M; Molné, M; Szpirer, J; Szpirer, C

1992-01-01

By comparing the methylation pattern of Mspl/Hpall sites in the 5' region of the mouse alpha-foetoprotein (AFP) gene of different cells (hepatoma cells, foetal and adult liver, fibroblasts), we found a correlation between gene expression and unmethylation of a site located in the first intron of the gene. Other sites did not show this correlation. In transfection experiments of unmethylated and methylated AFP-CAT chimeric constructions, we then showed that methylation of the intronic site negatively modulates expression of CAT activity. We also found that a DNA segment centered on this site binds nuclear proteins; however methylation did not affect protein binding. Images PMID:1371343

[Effect of ear point embedding on plasma and effect site concentrations of propofol-remifentanil in elderly patients after target-controlled induction].

PubMed

Zheng, Xiaochun; Wan, Liling; Gao, Fei; Chen, Jianghu; Tu, Wenshao

2017-08-12

To observe the clinical effect of ear point embedding on plasma and effect site concentrations of propofol-remifentanil in elderly patients who underwent abdominal external hernia surgery at the time of consciousness and pain disappearing by target-controlled infusion (TCI) and bispectral index (BIS). Fifty patients who underwent elective abdominal hernia surgery were randomly assigned into an observation group and a control group, 25 cases in each one. In the observation group, 30 minutes before anesthesia induction, Fugugou (Extra), Gan (CO 12 ), Pizhixia (AT 4 ), and Shenmen (TF 4 ) were embedded by auricular needles until the end of surgery, 10 times of counter press each point. In the control group, the same amount of auricular tape was applied until the end of surgery at the same points without stimulation 30 minutes before anesthesia induction. Patients in the two groups were given total intravenous anesthesia, and BIS was monitored by BIS anesthesia depth monitor. Propofol was infused by TCI at a beginning concentration of 1.5μg/L and increased by 0.3μg/L every 30s until the patients lost their consciousness. After that, remifentanil was infused by TCI at a beginning concentration of 2.0μg/L and increased by 0.3μg/L every 30s until the patients had no body reaction to pain stimulation (orbital reflex). Indices were recorded, including mean arterial pressure (MAP), heart rate (HR) and the BIS values, at the time of T 0 (entering into the operation room), T 1 (losing consciousness) and T 2 (pain relief), the plasma and effect site concentrations of propofol at T 1 , the plasma and effect site concentrations of remifentanil at T 2 . After surgery we recorded the total amounts of propofol and remifentanil, surgery time and anesthesia time. At T 1 and T 2 , MAP and HR of the observation group were higher than those of the control group ( P <0.05, P <0.01). At T 1 , the plasma and effect site concentrations of propofol in the observation group were

Does canopy mean N concentration explain differences in light use efficiency in 14 eddy-covariance sites?

NASA Astrophysics Data System (ADS)

Peltoniemi, Mikko; Pulkkinen, Minna; Kolari, Pasi; Mäkelä, Annikki

2010-05-01

Production efficiency models aim at explaining variation of vegetation productivity with climatic input and information on vegetation cover often obtained from satellite observations. It has been acknowledged that different plant species differ in their potential to assimilate carbon dioxide per unit of PAR (i.e light use efficiency, LUE). Subsequently, some LUE-based models apply different LUE-coefficients for different plant functional types. Leaf N concentrations differ between plant species, and related differences in light saturated photosynthesis rate (A_max) have been detected. How much these differences affect the ecosystem production or LUE is more obscure. Canopies acclimate to prevailing environmental conditions, which causes variation e.g. in the proportion of leaves exposed to direct sunlight, leaf morphology, structure,orientation, and vertical N distibution. Furthermore, a fair proportion of photosynthesis occurs during cloudy days, in which case high A_max is unessential, and number of these days differs by location. We studied if canopy mean N concentration could explain differences in LUE derived for 14 forest sites using eddy-covariance measurements. The largest actual LUE was estimated for each site directly as an upper percentile of the ratio of Gross Primary Production (GPP) to absorbed PAR. Potential LUE for each site, on the other hand, was estimated by parameterising a LUE-based production efficiency model (Prelued), which accounts for daily changes in weather (temperature, VPD, PAR). In this model structure, the LUE-parameter for each site, can be interpreted as the potential LUE under optimal environmental conditions, i.e when the environment is not limiting production at all. Averages of the largest actual LUE and potential LUE were higher in deciduous sites than in conifer sites. Canopy mean N correlated weakly with both the largest actual and potential LUE, and the correlation was also significant in conifer subset in the former case

Relationship between leptin concentrations and disease activity in patients with rheumatoid arthritis.

PubMed

Batún-Garrido, José Antonio de Jesús; Salas-Magaña, Marisol; Juárez-Rojop, Isela Esther; Hernández-Núñez, Eúfrates; Olán, Francisco

2018-05-11

Multiple studies have found a direct relationship between leptin concentrations and disease activity in rheumatoid arthritis. We studied 77 patients with the diagnosis of rheumatoid arthritis; the leptin determination was through an enzyme immunoassay. Disease activity was assessed by the DAS-28 CRP. A multivariate logistic regression model was used to determine the association between significant variables and leptin concentrations. 40.3% of the patients were in remission, 41.6% were mildly active, 11.7% were moderately active and 6.5% were severely active. The results show an independent association between higher concentrations of leptin and disease activity (OR 1.7; 95% CI 1.4-3.2; p .03), the number of swollen joints (OR 4.6; 95% CI 1.7-8.3; p .000), the number of painful joints (OR 3.4; 95% CI 1.6-4.6; p .000), and the presence of metabolic syndrome (OR 1.3; 95% IC 1.2-1,9; p .045). The data suggest that serum leptin is elevated in patients with active RA. Copyright © 2017 Elsevier España, S.L.U. All rights reserved.

Passive sampling to measure baseline dissolved persistent organic pollutant concentrations in the water column of the Palos Verdes Shelf Superfund site.

PubMed

Fernandez, Loretta A; Lao, Wenjian; Maruya, Keith A; White, Carmen; Burgess, Robert M

2012-11-06

Passive sampling was used to deduce water concentrations of persistent organic pollutants (POPs) in the vicinity of a marine Superfund site on the Palos Verdes Shelf, California, USA. Precalibrated solid phase microextraction (SPME) fibers and polyethylene (PE) strips that were preloaded with performance reference compounds (PRCs) were codeployed for 32 d along an 11-station gradient at bottom, surface, and midwater depths. Retrieved samplers were analyzed for DDT congeners and their breakdown products (DDE, DDD, DDMU, and DDNU) and 43 PCB congeners using GC-EI- and NCI-MS. PRCs were used to calculate compound-specific fractional equilibration achieved in situ for the PE samplers, using both an exponential approach to equilibrium (EAE) and numerical integration of Fickian diffusion (NI) models. The highest observed concentrations were for p,p'-DDE, with 2200 and 990 pg/L deduced from PE and SPME, respectively. The difference in these estimates could be largely attributed to uncertainty in equilibrium partition coefficients, unaccounted for disequilibrium between samplers and water, or different time scales over which the samplers average. The concordance between PE and SPME estimated concentrations for DDE was high (R(2) = 0.95). PCBs were only detected in PE samplers, due to their much larger size. Near-bottom waters adjacent to and down current from sediments with the highest bulk concentrations exhibited aqueous concentrations of DDTs and PCBs that exceeded Ambient Water Quality Criteria (AWQC) for human and aquatic health, indicating the need for future monitoring to determine the effectiveness of remedial activities taken to reduce adverse effects of contaminated surface sediments.

Strontium-90 and caesium-137 activity concentrations in bats in the Chernobyl exclusion zone.

PubMed

Gashchak, Sergey; Beresford, Nicholas Anthony; Maksimenko, Andrey; Vlaschenko, Anton S

2010-11-01

Bats are a protected species and as such may be an object of protection in radiological assessments of the environment. However, there have previously been only few radioecological studies of species of bats. In this paper, results for >140 measurements of (90)Sr and (137)Cs in 10 species of bats collected within the Chernobyl zone are presented. There was some indication of a decreasing transfer of (90)Sr with increasing deposition, although this was inconsistent across species and explained little of the observed variability. There was no difference between male and female bats in the transfer (expressed as the ratio of whole-body activity concentrations to those in soil) of either radionuclide. There was considerable variability in transfer across all species groups. At two sites where there were sufficient data, Eptesicus serotinus was found to have higher transfer than other species.

Molecular imprint of enzyme active site by camel nanobodies: rapid and efficient approach to produce abzymes with alliinase activity.

PubMed

Li, Jiang-Wei; Xia, Lijie; Su, Youhong; Liu, Hongchun; Xia, Xueqing; Lu, Qinxia; Yang, Chunjin; Reheman, Kalbinur

2012-04-20

Screening of inhibitory Ab1 antibodies is a critical step for producing catalytic antibodies in the anti-idiotypic approach. However, the incompatible surface of the active site of the enzyme and the antigen-binding site of heterotetrameric conventional antibodies become the limiting step. Because camelid-derived nanobodies possess the potential to preferentially bind to the active site of enzymes due to their small size and long CDR3, we have developed a novel approach to produce antibodies with alliinase activities by exploiting the molecular mimicry of camel nanobodies. By screening the camelid-derived variable region of the heavy chain cDNA phage display library with alliinase, we obtained an inhibitory nanobody VHHA4 that recognizes the active site. Further screening with VHHA4 from the same variable domain of the heavy chain of a heavy-chain antibody library led to a higher incidence of anti-idiotypic Ab2 abzymes with alliinase activities. One of the abzymes, VHHC10, showed the highest activity that can be inhibited by Ab1 VHHA4 and alliinase competitive inhibitor penicillamine and significantly suppressed the B16 tumor cell growth in the presence of alliin in vitro. The results highlight the feasibility of producing abzymes via anti-idiotypic nanobody approach.

Synthesis of a molecularly defined single-active site heterogeneous catalyst for selective oxidation of N-heterocycles.

PubMed

Zhang, Yujing; Pang, Shaofeng; Wei, Zhihong; Jiao, Haijun; Dai, Xingchao; Wang, Hongli; Shi, Feng

2018-04-13

Generally, a homogeneous catalyst exhibits good activity and defined active sites but it is difficult to recycle. Meanwhile, a heterogeneous catalyst can easily be reused but its active site is difficult to reveal. It is interesting to bridge the gap between homogeneous and heterogeneous catalysis via controllable construction of a heterogeneous catalyst containing defined active sites. Here, we report that a molecularly defined, single-active site heterogeneous catalyst has been designed and prepared via the oxidative polymerization of maleimide derivatives. These polymaleimide derivatives can be active catalysts for the selective oxidation of heterocyclic compounds to quinoline and indole via the recycling of -C=O and -C-OH groups, which was confirmed by tracing the reaction with GC-MS using maleimide as the catalyst and by FT-IR analysis with polymaleimide as the catalyst. These results might promote the development of heterogeneous catalysts with molecularly defined single active sites exhibiting a comparable activity to homogeneous catalysts.

Biological role of site-specific O-glycosylation in cell adhesion activity and phosphorylation of osteopontin.

PubMed

Oyama, Midori; Kariya, Yoshinobu; Kariya, Yukiko; Matsumoto, Kana; Kanno, Mayumi; Yamaguchi, Yoshiki; Hashimoto, Yasuhiro

2018-05-09

Osteopontin (OPN) is an extracellular glycosylated phosphoprotein that promotes cell adhesion by interacting with several integrin receptors. We previously reported that an OPN mutant lacking five O-glycosylation sites (Thr 134 /Thr 138 /Thr 143 /Thr 147 /Thr 152 ) in the threonine/proline-rich region increased cell adhesion activity and phosphorylation compared with the wild type. However, the role of O-glycosylation in cell adhesion activity and phosphorylation of OPN remains to be clarified. Here, we show that site-specific O-glycosylation in the threonine/proline-rich region of OPN affects its cell adhesion activity and phosphorylation independently and/or synergistically. Using site-directed mutagenesis, we found that OPN mutants with substitution sets of Thr 134 /Thr 138 or Thr 143 /Thr 147 /Thr 152 had decreased and increased cell adhesion activity, respectively. In contrast, the introduction of a single mutation into the O-glycosylation sites had no effect on OPN cell adhesion activity. An adhesion assay using function-blocking antibodies against αvβ3 and β1 integrins, as well as αvβ3 integrin-overexpressing A549 cells, revealed that site-specific O-glycosylation affected the association of OPN with the two integrins. Phosphorylation analyses using phos-tag and LC-MS/MS indicated that phosphorylation levels and sites were influenced by the O-glycosylation status, although the number of O-glycosylation sites was not correlated with the phosphorylation level in OPN. Furthermore, a correlation analysis between phosphorylation level and cell adhesion activity in OPN mutants with the site-specific O-glycosylation showed that they were not always correlated. These results provide conclusive evidence of a novel regulatory mechanism of cell adhesion activity and phosphorylation of OPN by site-specific O-glycosylation. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Magnetic Measurements and Heavy Metal Concentrations at Formosa Mine Superfund Site, Douglas County, OR

NASA Astrophysics Data System (ADS)

Upton, T. L.

2015-12-01

Advances in the field of environmental magnetism have led to exciting new applications for this field. Magnetic minerals are ubiquitous in the environment and tend to have an affinity for heavy metals. It has been demonstrated that magnetic properties are often significantly related to concentrations of heavy metals and/or pollution loading index (PLI). As a result, magnetic techniques have been used as proxy for determining hot spots of several types of pollution produced from a diversity of anthropogenic sources. Magnetic measurements are non-destructive and relatively inexpensive compared to geochemical analyses. The utility of environmental magnetic methods varies widely depending on biological, chemical and physical processes that create and transform soils and sediments. Applications in the direction of mapping heavy metals have been studied and shown to be quite useful in countries such as China and India but to date, little research has been done in the US. As such, there is need to expand the scope of research to a wider range of soil types and land uses, especially within the US. This study investigates the application of environmental magnetic techniques to mapping of heavy metal concentrations and PLI at the Formosa Mine Superfund Site, an abandoned mine about 25 miles southwest of Roseburg, OR. Using hotspot analysis, correlation and cluster analyses, interactions between metals and magnetic parameters are examined in relation to environmental factors such as proximity to seeps and adits. Preliminary results suggest significant correlation of magnetic susceptibility with certain heavy metals, signifying that magnetic methods may be useful in mapping heavy metal hotspots at this site.

Interactive flare sites within an active region complex

NASA Technical Reports Server (NTRS)

Poletto, G.; Gary, G. A.; Machado, M. E.

1993-01-01

We examine here a set of images of an active region complex, acquired on June 24-25, 1980, by the Hard X-ray Imaging Spectrometer on SMM, with the purpose of establishing whether there was any interplay between the frequent activity observed at different sites in the activity center and, in such a case, how the interaction was established. By analyzing both quiet and active orbits we show that, as a rule, activity originating in one region triggers the other region's activity. However, we find little unambiguous evidence for the presence of large-scale interconnecting loops. A comparison of X-ray images with magnetic field observations suggested that we interpret the active region behavior in terms of the interaction between different loop systems, in a scenario quite analogous to the interacting bipole representation of individual flares. We conclude that active region interplay provides an easily observable case to study the time-dependent topology and the mechanisms for the spreading of activity in transient events over all energy scales.

Ultrafast infrared spectroscopy reveals water-mediated coherent dynamics in an enzyme active site.

PubMed

Adamczyk, Katrin; Simpson, Niall; Greetham, Gregory M; Gumiero, Andrea; Walsh, Martin A; Towrie, Michael; Parker, Anthony W; Hunt, Neil T

2015-01-01

Understanding the impact of fast dynamics upon the chemical processes occurring within the active sites of proteins and enzymes is a key challenge that continues to attract significant interest, though direct experimental insight in the solution phase remains sparse. Similar gaps in our knowledge exist in understanding the role played by water, either as a solvent or as a structural/dynamic component of the active site. In order to investigate further the potential biological roles of water, we have employed ultrafast multidimensional infrared spectroscopy experiments that directly probe the structural and vibrational dynamics of NO bound to the ferric haem of the catalase enzyme from Corynebacterium glutamicum in both H 2 O and D 2 O. Despite catalases having what is believed to be a solvent-inaccessible active site, an isotopic dependence of the spectral diffusion and vibrational lifetime parameters of the NO stretching vibration are observed, indicating that water molecules interact directly with the haem ligand. Furthermore, IR pump-probe data feature oscillations originating from the preparation of a coherent superposition of low-frequency vibrational modes in the active site of catalase that are coupled to the haem ligand stretching vibration. Comparisons with an exemplar of the closely-related peroxidase enzyme family shows that they too exhibit solvent-dependent active-site dynamics, supporting the presence of interactions between the haem ligand and water molecules in the active sites of both catalases and peroxidases that may be linked to proton transfer events leading to the formation of the ferryl intermediate Compound I. In addition, a strong, water-mediated, hydrogen bonding structure is suggested to occur in catalase that is not replicated in peroxidase; an observation that may shed light on the origins of the different functions of the two enzymes.

Effects of Low Phytanic Acid-Concentrated DHA on Activated Microglial Cells: Comparison with a Standard Phytanic Acid-Concentrated DHA.

PubMed

Ruiz-Roso, María Belén; Olivares-Álvaro, Elena; Quintela, José Carlos; Ballesteros, Sandra; Espinosa-Parrilla, Juan F; Ruiz-Roso, Baltasar; Lahera, Vicente; de Las Heras, Natalia; Martín-Fernández, Beatriz

2018-05-30

Docosahexaenoic acid (DHA, 22:6 n-3) is an essential omega-3 (ω-3) long chain polyunsaturated fatty acid of neuronal membranes involved in normal growth, development, and function. DHA has been proposed to reduce deleterious effects in neurodegenerative processes. Even though, some inconsistencies in findings from clinical and pre-clinical studies with DHA could be attributed to the presence of phytanic acid (PhA) in standard DHA treatments. Thus, the aim of our study was to analyze and compare the effects of a low PhA-concentrated DHA with a standard PhA-concentrated DHA under different neurotoxic conditions in BV-2 activated microglial cells. To this end, mouse microglial BV-2 cells were stimulated with either lipopolysaccharide (LPS) or hydrogen peroxide (H 2 O 2 ) and co-incubated with DHA 50 ppm of PhA (DHA (PhA:50)) or DHA 500 ppm of PhA (DHA (PhA:500)). Cell viability, superoxide anion (O 2 - ) production, Interleukin 6 (L-6), cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), glutathione peroxidase (GtPx), glutathione reductase (GtRd), Caspase-3, and the brain-derived neurotrophic factor (BDNF) protein expression were explored. Low PhA-concentrated DHA protected against LPS or H 2 O 2 -induced cell viability reduction in BV-2 activated cells and O 2 - production reduction compared to DHA (PhA:500). Low PhA-concentrated DHA also decreased COX-2, IL-6, iNOS, GtPx, GtRd, and SOD-1 protein expression when compared to DHA (PhA:500). Furthermore, low PhA-concentrated DHA increased BDNF protein expression in comparison to DHA (PhA:500). The study provides data supporting the beneficial effect of low PhA-concentrated DHA in neurotoxic injury when compared to a standard PhA-concentrated DHA in activated microglia.

Unmasking tandem site interaction in human acetylcholinesterase. Substrate activation with a cationic acetanilide substrate.

PubMed

Johnson, Joseph L; Cusack, Bernadette; Davies, Matthew P; Fauq, Abdul; Rosenberry, Terrone L

2003-05-13

Acetylcholinesterase (AChE) contains a narrow and deep active site gorge with two sites of ligand binding, an acylation site (or A-site) at the base of the gorge, and a peripheral site (or P-site) near the gorge entrance. The P-site contributes to catalytic efficiency by transiently binding substrates on their way to the acylation site, where a short-lived acyl enzyme intermediate is produced. A conformational interaction between the A- and P-sites has recently been found to modulate ligand affinities. We now demonstrate that this interaction is of functional importance by showing that the acetylation rate constant of a substrate bound to the A-site is increased by a factor a when a second molecule of substrate binds to the P-site. This demonstration became feasible through the introduction of a new acetanilide substrate analogue of acetylcholine, 3-(acetamido)-N,N,N-trimethylanilinium (ATMA), for which a = 4. This substrate has a low acetylation rate constant and equilibrates with the catalytic site, allowing a tractable algebraic solution to the rate equation for substrate hydrolysis. ATMA affinities for the A- and P-sites deduced from the kinetic analysis were confirmed by fluorescence titration with thioflavin T as a reporter ligand. Values of a >1 give rise to a hydrolysis profile called substrate activation, and the AChE site-specific mutant W86F, and to a lesser extent wild-type human AChE itself, showed substrate activation with acetylthiocholine as the substrate. Substrate activation was incorporated into a previous catalytic scheme for AChE in which a bound P-site ligand can also block product dissociation from the A-site, and two additional features of the AChE catalytic pathway were revealed. First, the ability of a bound P-site ligand to increase the substrate acetylation rate constant varied with the structure of the ligand: thioflavin T accelerated ATMA acetylation by a factor a(2) of 1.3, while propidium failed to accelerate. Second, catalytic rate

Effect-site concentration of propofol required for LMA-Supreme™ insertion with and without remifentanil: a randomized controlled trial.

PubMed

Zaballos, Matilde; Bastida, Emilia; Agustí, Salomé; Portas, Maite; Jiménez, Consuelo; López-Gil, Maite

2015-10-06

A new supraglottic device, the LMA-Supreme™, has recently become available for clinical use. Information on anaesthetic and co-adjuvant requirements for insertion of the LMA-Supreme™ is limited. The present study aimed to evaluate the optimal effect-site concentration of propofol in 50 % (EC50) of adults necessary for successful insertion of the LMA-Supreme™ and to examine remifentanil's effect on propofol requirements. Fifty-eight elective patients (aged 18-60 years; ASA (American Society Anaesthesiologists) physical status classification I and II) scheduled for day surgery were randomly assigned to one of two groups: propofol with saline or propofol with remifentanil. Anaesthesia was induced by target-controlled infusion according to predetermined effect-site concentrations of propofol and remifentanil (5 ng.mL(-1)). The EC50 was calculated using Dixon's up-and-down method. Ten minutes following drug administration, LMA-Supreme™ insertion was attempted without the use of muscle relaxant drugs. In the propofol + saline group, the EC50 of propofol required for LMA-Supreme™ insertion was 6.32 ± 0.67 μg.mL(-1) (95 % CI, 5.69-6.94 μg.mL(-1)). With the addition of remifentanil at an effect-site concentration of 5 ng.mL(-1), the EC50 of propofol required for LMA-Supreme™ insertion was 2.50 ± 0.80 μg.mL(-1) (95 % CI, 1.82-3.17 μg.mL(-1); p < 0.0001). The propofol requirement for smooth insertion of the LMA-Supreme™ was 60 % less when remifentanil (5 ng.mL(-1)) was co-administered. Identified as NCT01974648 at www.clinicaltrials.gov .

Binding Site Concentration Explains the Differential Susceptibility of Chilo suppressalis and Sesamia inferens to Cry1A-Producing Rice

PubMed Central

Han, Chao; Liu, Zewen; Chen, Fajun; Hou, Maolin; Peng, Yufa

2014-01-01

Chilo suppressalis and Sesamia inferens are two important lepidopteran rice pests that occur concurrently during outbreaks in paddy fields in the main rice-growing areas of China. Previous and current field tests demonstrate that the transgenic rice line Huahui 1 (HH1) producing a Cry1Ab-Cry1Ac hybrid toxin from the bacterium Bacillus thuringiensis reduces egg and larval densities of C. suppressalis but not of S. inferens. This differential susceptibility to HH1 rice correlates with the reduced susceptibility to Cry1Ab and Cry1Ac toxins in S. inferens larvae compared to C. suppressalis larvae. The goal of this study was to identify the mechanism responsible for this differential susceptibility. In saturation binding assays, both Cry1Ab and Cry1Ac toxins bound with high affinity and in a saturable manner to midgut brush border membrane vesicles (BBMV) from C. suppressalis and S. inferens larvae. While binding affinities were similar, a dramatically lower concentration of Cry1A toxin binding sites was detected for S. inferens BBMV than for C. suppressalis BBMV. In contrast, no significant differences between species were detected for Cry1Ca toxin binding to BBMV. Ligand blotting detected BBMV proteins binding Cry1Ac or Cry1Ca toxins, some of them unique to C. suppressalis or S. inferens. These data support that reduced Cry1A binding site concentration is associated with a lower susceptibility to Cry1A toxins and HH1 rice in S. inferens larvae than in C. suppressalis larvae. Moreover, our data support Cry1Ca as a candidate for pyramiding efforts with Cry1A-producing rice to extend the activity range and durability of this technology against rice stem borers. PMID:24928872

Binding site concentration explains the differential susceptibility of Chilo suppressalis and Sesamia inferens to Cry1A-producing rice.

PubMed

Han, Lanzhi; Han, Chao; Liu, Zewen; Chen, Fajun; Jurat-Fuentes, Juan Luis; Hou, Maolin; Peng, Yufa

2014-08-01

Chilo suppressalis and Sesamia inferens are two important lepidopteran rice pests that occur concurrently during outbreaks in paddy fields in the main rice-growing areas of China. Previous and current field tests demonstrate that the transgenic rice line Huahui 1 (HH1) producing a Cry1Ab-Cry1Ac hybrid toxin from the bacterium Bacillus thuringiensis reduces egg and larval densities of C. suppressalis but not of S. inferens. This differential susceptibility to HH1 rice correlates with the reduced susceptibility to Cry1Ab and Cry1Ac toxins in S. inferens larvae compared to C. suppressalis larvae. The goal of this study was to identify the mechanism responsible for this differential susceptibility. In saturation binding assays, both Cry1Ab and Cry1Ac toxins bound with high affinity and in a saturable manner to midgut brush border membrane vesicles (BBMV) from C. suppressalis and S. inferens larvae. While binding affinities were similar, a dramatically lower concentration of Cry1A toxin binding sites was detected for S. inferens BBMV than for C. suppressalis BBMV. In contrast, no significant differences between species were detected for Cry1Ca toxin binding to BBMV. Ligand blotting detected BBMV proteins binding Cry1Ac or Cry1Ca toxins, some of them unique to C. suppressalis or S. inferens. These data support that reduced Cry1A binding site concentration is associated with a lower susceptibility to Cry1A toxins and HH1 rice in S. inferens larvae than in C. suppressalis larvae. Moreover, our data support Cry1Ca as a candidate for pyramiding efforts with Cry1A-producing rice to extend the activity range and durability of this technology against rice stem borers. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Cytochrome P450 and contaminant concentrations in nestling black-crowned night-herons and their interrelation with sibling embryos

USGS Publications Warehouse

Rattner, Barnett A.; Melancon, Mark J.; Custer, Thomas W.; Hothem, Roger L.

1996-01-01

Hepatic cytochrome P450-associated monooxygenase activities were measured in 11-d-old nestling black-crowned night-herons (Nycticorax nycticorax) collected from a reference site (next to Chincoteague National Wildlife Refuge, VA, USA) and three contaminated sites (Cat Island, Green Bay, WI, USA; Bair Island, San Francisco Bay, CA, USA; and West Marin Island, San Francisco Bay, CA, USA). Arylhydrocarbon hydroxylase and benzyloxyresorufin-O-dealkylase activities of nestlings from contaminated sites were only slightly elevated (less than threefold) compared with the reference site. Organochlorine pesticide and total polychlorinated biphenyl (PCB) concentrations in nestlings were greatest at contaminated sites, although much lower than found in concurrently collected eggs and pipping embryos. Pollutant concentrations of nestlings were rarely associated with monooxygenase activity. In contrast, concurrently collected pipping heron embryos (often siblings of the nestlings) exhibited pronounced monooxygenase induction (means at contaminated sites were elevated up to sevenfold and values of some embryos exceeded 25-fold induction). Furthermore, monooxygenase activity of pipping embryos was significantly correlated with total PCBs, arylhydrocarbon receptor-active PCB congeners, and toxic equivalents. The modest monooxygenase responses of heron nestlings suggest that this biomarker may have only limited value during this rapid-growth life stage.

Engineering streptokinase for generation of active site-labeled plasminogen analogs*

PubMed Central

Laha, Malabika; Panizzi, Peter; Nahrendorf, Matthias; Bock, Paul E.

2011-01-01

We previously demonstrated that streptokinase (SK) can be used to generate active site-labeled fluorescent analogs of plasminogen (Pg) by virtue of its non-proteolytic activation of the zymogen. The method is versatile and allows for stoichiometric and active site-specific incorporation of any one of many molecular probes. The limitation of the labeling approach is that it is both time-consuming and low yield. Here we demonstrate an improved method for the preparation of labeled Pg analogs by the use of an engineered SK mutant fusion protein with both COOH- and NH2-terminal His6-tags. The NH2-terminal tag is followed by a tobacco etch virus proteinase cleavage site to ensure that the SK Ile1 residue, essential for conformational activation of Pg, is preserved. The SK COOH-terminal Lys414 residue and residues Arg253-Leu260 in the SK β-domain were deleted to prevent cleavage by plasmin (Pm), and to disable Pg substrate binding to the SK·Pg*/Pm catalytic complexes, respectively. Near-elimination of Pm generation with the SKΔ(R253-L260)ΔK414-His6 mutant increased the yield of labeled Pg 2.6-fold and reduced the time required >2-fold. The versatility of the labeling method was extended to the application of Pg labeled with a near-infrared probe to quantitate Pg receptors on immune cells by flow cytometry. PMID:21570944

Effect of acute stress on plasma beta-corticosterone, estradiol-17 beta and testosterone concentrations in juvenile American alligators collected from three sites within the Kissimmee-Everglades drainage basin in Florida (USA).

PubMed

Gunderson, M P; Kools, S A E; Milnes, M R; Guillette, L J

2003-07-01

The effect of acute stress on plasma beta-corticosterone (B), testosterone (T) and estradiol-17beta (E2) concentrations in juvenile alligators collected from sites with varying sediment contaminants was examined in this study. Dramatic increases in plasma B concentrations were observed in alligators from all of the sites after 2 h of capture although females from the intermediate contaminant site exhibited a significantly lower percentage increase in B than females from the other two sites. Males from the site with the highest contaminant levels exhibited elevated initial B concentrations relative to the other sites. This pattern was not observed after 2 h of restraint. Females from the highest contaminant site exhibited depressed initial T when compared to the other sites although this pattern was not observed after 2 h of restraint. Neither E2 nor T decreased after 2 h in females, whereas T concentrations decreased in all males over the same time period. The variance associated with these endpoints was also examined to determine whether it could serve as a more sensitive marker for perturbations of the endocrine system and stress response. Females from the higher and intermediate contaminant sites exhibited the lowest and highest standard errors (respectively) associated with 2 h plasma B concentrations with no differences among mean concentrations suggesting a perturbation of the stress response in these animals that was not detected by examining the means. We concluded that the environmental contaminants could be acting as stressors, leading to the observed differences.

Correlation of endothelin-1 concentration and angiotensin-converting enzyme activity with the staging of liver fibrosis.

PubMed

Kardum, Dusko; Fabijanić, Damir; Lukić, Anita; Romić, Zeljko; Petrovecki, Mladen; Bogdanović, Zoran; Jurić, Klara; Urek-Crncević, Marija; Banić, Marko

2012-06-01

Increased serum angiotensin-converting enzyme (SACE) activity and serum concentration of endothelin-1 (ET-1) were found in liver cirrhosis. We investigated a correlation between the different stages of liver fibrosis and SACE activity and serum ET-1 concentration. Seventy patients with pathohistologically established chronic liver disease were divided in three groups according to Ishak criteria for liver fibrosis: minimal fibrosis (Ishak score 0-1, n =20), medium fibrosis (Ishak score 2-5, n=20) and cirrhosis (Ishak score 6, n=30). SACE activity and ET-1 concentration were determined using commercial ELISA kits. SACE activity and ET-1 concentrations were proportional to the severity of disease, the highest being in patients with liver cirrhosis. Maximal increase in SACE activity was found between minimal and medium fibrosis while maximal increase in ET-1 concentration was revealed between medium fibrosis and cirrhosis. The analysis of the Receiver Operating Characteristic (ROC) curve for SACE activity suggested a cut-off value to separate minimal from medium fibrosis at 59.00 U/L (sensitivity 100%, specificity 64.7%). The cut-off value for serum ET-1 concentration to separate medium fibrosis from cirrhosis was 12.4 pg/mL (sensitivity 96.8%, specificity 94.4%). A positive correlation between SACE activity and ET-1 concentration was registered (Spearman's ñ = 0.438, p = 0.004). Both SACE activity and ET-1 concentration were increased in all stages of liver fibrosis. Cut-off points for SACE activity and ET-1 concentration could be a biochemical marker for the progression of fibrosis. Positive correlation between SACE activity and ET-1 concentration might indicate their interaction in the development of liver cirrhosis.

2011 Groundwater Monitoring and Inspection Report Gnome-Coach Site, New Mexico

DOE Office of Scientific and Technical Information (OSTI.GOV)

None

2012-02-01

Gnome-Coach was the site of a 3-kiloton underground nuclear test in 1961. Surface and subsurface contamination resulted from the underground nuclear testing, post-test drilling, and groundwater tracer test performed at the site. The State of New Mexico is currently proceeding with a conditional certificate of completion for the surface. As for the subsurface, monitoring activities that include hydraulic head monitoring and groundwater sampling of the wells onsite are conducted as part of the annual site inspection. These activities were conducted on January 19, 2011. The site roads, monitoring well heads, and the monument at surface ground zero were observed asmore » being in good condition at the time of the site inspection. An evaluation of the hydraulic head data obtained from the site indicates that water levels in wells USGS-4 and USGS-8 appear to respond to the on/off cycling of the dedicated pump in well USGS-1 and that water levels in wells LRL-7 and DD-1 increased during this annual monitoring period. Analytical results obtained from the sampling indicate that concentrations of tritium, strontium-90, and cesium-137 were consistent with concentrations from historical sampling events.« less

Reactive sites influence in PMMA oligomers reactivity: a DFT study

NASA Astrophysics Data System (ADS)

Paz, C. V.; Vásquez, S. R.; Flores, N.; García, L.; Rico, J. L.

2018-01-01

In this work, we present a theoretical study of methyl methacrylate (MMA) living anionic polymerization. The study was addressed to understanding two important experimental observations made for Michael Szwarc in 1956. The unexpected effect of reactive sites concentration in the propagation rate, and the self-killer behavior of MMA (deactivating of living anionic polymerization). The theoretical calculations were performed by density functional theory (DFT) to obtain the frontier molecular orbitals values. These values were used to calculate and analyze the chemical interaction descriptors in DFT-Koopmans’ theorem. As a result, it was observed that the longest chain-length species (related with low concentration of reactive sites) exhibit the highest reactivity (behavior associated with the increase of the propagation rate). The improvement in this reactivity was attributed to the crosslinking produced in the polymethyl methacrylate chains. Meanwhile, the self-killer behavior was associated with the intermolecular forces present in the reactive sites. This behavior was associated to an obstruction in solvation, since the active sites remained active through all propagation species. The theoretical results were in good agreement with the Szwarc experiments.

Quantifying the density and utilization of active sites in non-precious metal oxygen electroreduction catalysts

PubMed Central

Sahraie, Nastaran Ranjbar; Kramm, Ulrike I.; Steinberg, Julian; Zhang, Yuanjian; Thomas, Arne; Reier, Tobias; Paraknowitsch, Jens-Peter; Strasser, Peter

2015-01-01

Carbon materials doped with transition metal and nitrogen are highly active, non-precious metal catalysts for the electrochemical conversion of molecular oxygen in fuel cells, metal air batteries, and electrolytic processes. However, accurate measurement of their intrinsic turn-over frequency and active-site density based on metal centres in bulk and surface has remained difficult to date, which has hampered a more rational catalyst design. Here we report a successful quantification of bulk and surface-based active-site density and associated turn-over frequency values of mono- and bimetallic Fe/N-doped carbons using a combination of chemisorption, desorption and 57Fe Mössbauer spectroscopy techniques. Our general approach yields an experimental descriptor for the intrinsic activity and the active-site utilization, aiding in the catalyst development process and enabling a previously unachieved level of understanding of reactivity trends owing to a deconvolution of site density and intrinsic activity. PMID:26486465

Tunable multiple emissions in manganese-concentrated sulfide through simultaneous tailoring of Mn-site coordination and Mn-Mn pair geometry

NASA Astrophysics Data System (ADS)

Chen, Zitao; Song, Enhai; Ye, Shi; Zhang, Qinyuan

2017-12-01

In contrast to generally single-band visible emission feature from Mn2+, simultaneous visible (VIS) and near-infrared (NIR) multiple emissions are demonstrated in Mn2+ concentrated sulfide (MnS) by only involving a single crystallographic site. Upon varying the Mn2+-site coordination and/or Mn-Mn pairs geometry in different structural MnS, the multiple emissions from divalent manganese can be easily tuned from 575 to 720 nm (VIS) or from 880 to 900 or 1380 nm (NIR), respectively. The excitation spectroscopy and the luminescent decay, together with crystal structural analyses, are employed to investigate the electronic transition and the excited state dynamics of these Mn2+ concentrated systems. It is found that the VIS and NIR emissions can be ascribed to the isolated Mn2+ ion and exchange coupled Mn-Mn pair center, respectively. The effect of crystal field and bridging geometry, as well as temperature on the exchange coupled Mn2+ pairs NIR emissive center, is also investigated in detail. This work not only provides keen insights into the de-excitation pathway of Mn2+-concentrated material, but also offers the possibilities of designing a novel NIR emitting source for various photonic applications.

Exploring the Active Site of the Tungsten, Iron-Sulfur Enzyme Acetylene Hydratase▿ †

PubMed Central

tenBrink, Felix; Schink, Bernhard; Kroneck, Peter M. H.

2011-01-01

The soluble tungsten, iron-sulfur enzyme acetylene hydratase (AH) from mesophilic Pelobacter acetylenicus is a member of the dimethyl sulfoxide (DMSO) reductase family. It stands out from its class as it catalyzes a nonredox reaction, the addition of H2O to acetylene (H—C☰C—H) to form acetaldehyde (CH3CHO). Caught in its active W(IV) state, the high-resolution three-dimensional structure of AH offers an excellent starting point to tackle its unique chemistry and to identify catalytic amino acid residues within the active site cavity: Asp13 close to W(IV) coordinated to two molybdopterin-guanosine-dinucleotide ligands, Lys48 which couples the [4Fe-4S] cluster to the W site, and Ile142 as part of a hydrophobic ring at the end of the substrate access channel designed to accommodate the substrate acetylene. A protocol was developed to express AH in Escherichia coli and to produce active-site variants which were characterized with regard to activity and occupancy of the tungsten and iron-sulfur centers. By this means, fusion of the N-terminal chaperone binding site of the E. coli nitrate reductase NarG to the AH gene improved the yield and activity of AH and its variants significantly. Results from site-directed mutagenesis of three key residues, Asp13, Lys48, and Ile142, document their important role in catalysis of this unusual tungsten enzyme. PMID:21193613

Comparison of the active-site design of molybdenum oxo-transfer enzymes by quantum mechanical calculations.

PubMed

Li, Jilai; Ryde, Ulf

2014-11-17

There are three families of mononuclear molybdenum enzymes that catalyze oxygen atom transfer (OAT) reactions, named after a typical example from each family, viz., dimethyl sulfoxide reductase (DMSOR), sulfite oxidase (SO), and xanthine oxidase (XO). These families differ in the construction of their active sites, with two molybdopterin groups in the DMSOR family, two oxy groups in the SO family, and a sulfido group in the XO family. We have employed density functional theory calculations on cluster models of the active sites to understand the selection of molybdenum ligands in the three enzyme families. Our calculations show that the DMSOR active site has a much stronger oxidative power than the other two sites, owing to the extra molybdopterin ligand. However, the active sites do not seem to have been constructed to make the OAT reaction as exergonic as possible, but instead to keep the reaction free energy close to zero (to avoid excessive loss of energy), thereby making the reoxidation (SO and XO) or rereduction of the active sites (DMSOR) after the OAT reaction facile. We also show that active-site models of the three enzyme families can all catalyze the reduction of DMSO and that the DMSOR model does not give the lowest activation barrier. Likewise, all three models can catalyze the oxidation of sulfite, provided that the Coulombic repulsion between the substrate and the enzyme model can be overcome, but for this harder reaction, the SO model gives the lowest activation barrier, although the differences are not large. However, only the XO model can catalyze the oxidation of xanthine, owing to its sulfido ligand.

Enzyme/non-enzyme discrimination and prediction of enzyme active site location using charge-based methods.

PubMed

Bate, Paul; Warwicker, Jim

2004-07-02

Calculations of charge interactions complement analysis of a characterised active site, rationalising pH-dependence of activity and transition state stabilisation. Prediction of active site location through large DeltapK(a)s or electrostatic strain is relevant for structural genomics. We report a study of ionisable groups in a set of 20 enzymes, finding that false positives obscure predictive potential. In a larger set of 156 enzymes, peaks in solvent-space electrostatic properties are calculated. Both electric field and potential match well to active site location. The best correlation is found with electrostatic potential calculated from uniform charge density over enzyme volume, rather than from assignment of a standard atom-specific charge set. Studying a shell around each molecule, for 77% of enzymes the potential peak is within that 5% of the shell closest to the active site centre, and 86% within 10%. Active site identification by largest cleft, also with projection onto a shell, gives 58% of enzymes for which the centre of the largest cleft lies within 5% of the active site, and 70% within 10%. Dielectric boundary conditions emphasise clefts in the uniform charge density method, which is suited to recognition of binding pockets embedded within larger clefts. The variation of peak potential with distance from active site, and comparison between enzyme and non-enzyme sets, gives an optimal threshold distinguishing enzyme from non-enzyme. We find that 87% of the enzyme set exceeds the threshold as compared to 29% of the non-enzyme set. Enzyme/non-enzyme homologues, "structural genomics" annotated proteins and catalytic/non-catalytic RNAs are studied in this context.

Bactericidal activity of electrolyzed acid water from solution containing sodium chloride at low concentration, in comparison with that at high concentration.

PubMed

Kiura, Hiromasa; Sano, Kouichi; Morimatsu, Shinichi; Nakano, Takashi; Morita, Chizuko; Yamaguchi, Masaki; Maeda, Toyoyuki; Katsuoka, Yoji

2002-05-01

Electrolyzed strong acid water (ESW) containing free chlorine at various concentrations is becoming to be available in clinical settings as a disinfectant. ESW is prepared by electrolysis of a NaCl solution, and has a corrosive activity against medical instruments. Although lower concentrations of NaCl and free chlorine are desired to eliminate corrosion, the germicidal effect of ESW with low NaCl and free-chlorine concentrations (ESW-L) has not been fully clarified. In this study, we demonstrated that ESW-L possesses bactericidal activity against Mycobacteria and spores of Bacillus subtilis. The effect was slightly weaker than that of ESW containing higher NaCl and free-chlorine concentrations (ESW-H), but acceptable as a disinfectant. To clarify the mechanism of the bactericidal activity, we investigated ESW-L-treated Pseudomonas aeruginosa by transmission electron microscopy, a bacterial enzyme assay and restriction fragment length polymorphism pattern (RFLP) assay. Since the bacterium, whose growth was completely inhibited by ESW-L, revealed the inactivation of cytoplasmic enzyme, blebs and breaks in its outer membrane and remained complete RFLP of DNA, damage of the outer membrane and inactivation of cytoplasmic enzyme are the important determinants of the bactericidal activity.

The production of high efficiency Ziegler-Natta catalyst with dual active sites nature using cyclohexyl chloride as promoter with super activity and produced superior polyethylene with controllable molecular weight distribution.

PubMed

Seifali Abbas-Abadi, Mehrdad

2017-01-01

In the previous studies, the several halocarbons (HC) were tested as promoters for a Ti-based Ziegler-Natta (ZN) catalyst at different polymerization conditions. The Results showed that chloro cyclohexane has the best operation in catalyst activity, polymer particle size growth, hydrogen responsibility and wax reduction too. For the first time in this study, the effect of Al/Ti ratio on the optimum HC/Ti ratio has been considered and the results showed that the optimum HC/Ti ratio depends on the Al/Ti ratio directly. In the optimum HC/Ti ratio, the catalyst activity and hydrogen responsibility ratio of the catalyst increase up to 125 and 55% respectively. The acceptable growth of polymer powder up to 46%, lower flow rate ratio (FRR) up to 19% and decrease of wax amount up to 12%, completed the promotion results. Furthermore, in the next part of this study and as key note, a little dose of halocarbon was used in the catalyst preparation to produce the special catalysts with dual active sites. In the catalyst preparation, the concentration of each active sites depends on the halocarbon amount and it can control the molecular weight distribution of the produced polyethylene; because each active sites have different response to hydrogen. The halocarbon based catalysts showed the remarkable effect on the catalyst activity, the molecular weight and especially molecular weight distribution (MWD). The flow rate ratio and MWD could be increased up to 77 and 88% respectively as the main result of halocarbon addition during the catalyst preparation.

The production of high efficiency Ziegler–Natta catalyst with dual active sites nature using cyclohexyl chloride as promoter with super activity and produced superior polyethylene with controllable molecular weight distribution

PubMed Central

Seifali Abbas-Abadi, Mehrdad

2017-01-01

Abstract In the previous studies, the several halocarbons (HC) were tested as promoters for a Ti-based Ziegler–Natta (ZN) catalyst at different polymerization conditions. The Results showed that chloro cyclohexane has the best operation in catalyst activity, polymer particle size growth, hydrogen responsibility and wax reduction too. For the first time in this study, the effect of Al/Ti ratio on the optimum HC/Ti ratio has been considered and the results showed that the optimum HC/Ti ratio depends on the Al/Ti ratio directly. In the optimum HC/Ti ratio, the catalyst activity and hydrogen responsibility ratio of the catalyst increase up to 125 and 55% respectively. The acceptable growth of polymer powder up to 46%, lower flow rate ratio (FRR) up to 19% and decrease of wax amount up to 12%, completed the promotion results. Furthermore, in the next part of this study and as key note, a little dose of halocarbon was used in the catalyst preparation to produce the special catalysts with dual active sites. In the catalyst preparation, the concentration of each active sites depends on the halocarbon amount and it can control the molecular weight distribution of the produced polyethylene; because each active sites have different response to hydrogen. The halocarbon based catalysts showed the remarkable effect on the catalyst activity, the molecular weight and especially molecular weight distribution (MWD). The flow rate ratio and MWD could be increased up to 77 and 88% respectively as the main result of halocarbon addition during the catalyst preparation. PMID:29491824

Torque, power and muscle activation of eccentric and concentric isokinetic cycling.

PubMed

Green, David J; Thomas, Kevin; Ross, Emma Z; Green, Steven C; Pringle, Jamie S M; Howatson, Glyn

2018-06-01

This study aimed to establish the effect of cycling mode and cadence on torque, external power output, and lower limb muscle activation during maximal, recumbent, isokinetic cycling. After familiarisation, twelve healthy males completed 6 × 10 s of maximal eccentric (ECC) and concentric (CON) cycling at 20, 40, 60, 80, 100, and 120 rpm with five minutes recovery. Vastus lateralis, medial gastrocnemius, rectus femoris, and biceps femoris surface electromyography was recorded throughout. As cadence increased, peak torque linearly decreased during ECC (350-248 N·m) and CON (239-117 N·m) and peak power increased in a parabolic manner. Crank angle at peak torque increased with cadence in CON (+13°) and decreased in ECC (-9.0°). At all cadences, peak torque (mean +129 N·m, range 111-143 N·m), and power (mean +871 W, range 181-1406 W), were greater during ECC compared to CON. For all recorded muscles the crank angle at peak muscle activation was greater during ECC compared to CON. This difference increased with cadence in all muscles except the vastus lateralis. Additionally, peak vastus laterallis and biceps femoris activation was greater during CON compared to ECC. Eccentric cycling offers a greater mechanical stimulus compared to concentric cycling but the effect of cadence is similar between modalities. Markers of technique (muscle activation, crank angle at peak activation and torque) were different between eccentric and concentric cycling and respond differently to changes in cadence. Such data should be considered when comparing between, and selecting cadences for, recumbent, isokinetic, eccentric and concentric cycling. Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.

Low concentrations of bilirubin inhibit activation of hepatic stellate cells in vitro.

PubMed

Tang, Yinhe; Zhang, Qiyu; Zhu, Yefan; Chen, Gang; Yu, Fuxiang

2017-04-01

Hepatic stellate cell (HSC) activation serves a key role in liver fibrosis, and is associated with chronic liver diseases. Bilirubin, a product of heme degradation, has been demonstrated to have antioxidant properties. The present study investigated the effects of physiological concentrations of bilirubin on rat HSC activation. Rat HSCs were isolated and cultured for several generations to induce activation. The activated HSCs were subsequently treated with 0, 1, 10 or 20 mg/l bilirubin and assayed for parameters of cell activation. As the bilirubin concentration increased, HSCs demonstrated reduced production of reactive oxygen species, reduced protein expression levels of α‑smooth muscle actin, a decreased mRNA expression ratio of tissue inhibitor of matrix metalloproteinase‑1/matrix metalloproteinase‑2, decreased proliferation and increased apoptosis. In conclusion, elevated bilirubin levels, within its physiological concentration range, appeared to inhibit HSC activation. These findings suggested a potential role for bilirubin in the treatment of fibrosis that requires further investigation.

Active Site Flexibility of Mycobacterium tuberculosis Isocitrate Lyase in Dimer Form.

PubMed

Lee, Yie-Vern; Choi, Sy Bing; Wahab, Habibah A; Choong, Yee Siew

2017-09-25

Tuberculosis (TB) still remains a global threat due to the emergence of a drug-resistant strain. Instead of focusing on the drug target of active stage TB, we are highlighting the isocitrate lyase (ICL) at the dormant stage TB. ICL is one of the persistent factors for Mycobacterium tuberculosis (MTB) to survive during the dormant phase. In addition, the absence of ICL in human has made ICL a potential drug target for TB therapy. However, the dynamic details of ICL which could give insights to the ICL-ligand interaction have yet to be solved. Therefore, a series of ICL dimer dynamics studies through molecular dynamics simulation were performed in this work. The ICL active site entrance gate closure is contributed to by hydrogen bonding and electrostatic interactions with the C-terminal. Analysis suggested that the open-closed behavior of the ICL active site entrance depends on the type of ligand present in the active site. We also observed four residues (Ser91, Asp108, Asp153, and Cys191) which could possibly be the nucleophiles for nucleophilic attack on the cleavage of isocitrate at the C 2 -C 3 bond. We hope that the elucidation of ICL dynamics can benefit future works such as lead identification or antibody design against ICL for TB therapeutics.

Blogs and Social Network Sites as Activity Systems: Exploring Adult Informal Learning Process through Activity Theory Framework

ERIC Educational Resources Information Center

Heo, Gyeong Mi; Lee, Romee

2013-01-01

This paper uses an Activity Theory framework to explore adult user activities and informal learning processes as reflected in their blogs and social network sites (SNS). Using the assumption that a web-based space is an activity system in which learning occurs, typical features of the components were investigated and each activity system then…

Counting Active Sites on Titanium Oxide-Silica Catalysts for Hydrogen Peroxide Activation through In Situ Poisoning with Phenylphosphonic Acid

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eaton, Todd R.; Boston, Andrew M.; Thompson, Anthony B.

2015-06-04

Quantifying specific active sites in supported catalysts improves our understanding and assists in rational design. Supported oxides can undergo significant structural changes as surface densities increase from site-isolated cations to monolayers and crystallites, which changes the number of kinetically relevant sites. Herein, TiO x domains are titrated on TiO x–SiO 2 selectively with phenylphosphonic acid (PPA). An ex situ method quantifies all fluid-accessible TiO x, whereas an in situ titration during cis-cyclooctene epoxidation provides previously unavailable values for the number of tetrahedral Ti sites on which H 2O 2 activation occurs. We use this method to determine the active sitemore » densities of 22 different catalysts with different synthesis methods, loadings, and characteristic spectra and find a single intrinsic turnover frequency for cis-cyclooctene epoxidation of (40±7) h -1. This simple method gives molecular-level insight into catalyst structure that is otherwise hidden when bulk techniques are used.« less

Concentrated dispersions of equilibrium protein nanoclusters that reversibly dissociate into active monomers

NASA Astrophysics Data System (ADS)

Truskett, Thomas M.; Johnston, Keith; Maynard, Jennifer; Borwankar, Ameya; Miller, Maria; Wilson, Brian; Dinin, Aileen; Khan, Tarik; Kaczorowski, Kevin

2012-02-01

Stabilizing concentrated protein solutions is of wide interest in drug delivery. However, a major challenge is how to reliably formulate concentrated, low viscosity (i.e., syringeable) solutions of biologically active proteins. Unfortunately, proteins typically undergo irreversible aggregation at intermediate concentrations of 100-200 mg/ml. In this talk, I describe how they can effectively avoid these intermediate concentrations by reversibly assembling into nanoclusters. Nanocluster assembly is achieved by balancing short-ranged, cosolute-induced attractions with weak, longer-ranger electrostatic repulsions near the isoelectric point. Theory predicts that native proteins are stabilized by a self-crowding mechanism within the concentrated environment of the nanoclusters, while weak cluster-cluster interactions can result in colloidally-stable dispersions with moderate viscosities. I present experimental results where this strategy is used to create concentrated antibody dispersions (up to 260 mg/ml) comprising nanoclusters of proteins [monoclonal antibody 1B7, polyclonal sheep Immunoglobin G and bovine serum albumin], which upon dilution in vitro or administration in vivo, are conformationally stable and retain activity.

Quantitative Analysis of Uranium Accumulation on Sediments during Field-scale Biostimulation under Variable Bicarbonate Concentrations at the Rifle IFRC Site

NASA Astrophysics Data System (ADS)

Fox, P. M.; Davis, J. A.; Bargar, J.; Williams, K. H.; Singer, D. M.; Long, P.

2011-12-01

Bioremediation of uranium in subsurface environments is an approach that has been used at numerous field sites throughout the U.S in an attempt to lower dissolved U(VI) concentrations in groundwater. At the Rifle IFRC research site in Colorado, biostimulation of the native microbial population through acetate amendment for various periods of time has been tested in order to immobilize uranium through reduction U(VI) to U(IV). While this approach has successfully decreased U(VI) concentrations in the dissolved phase, often to levels below the EPA's maximum contaminant level of 0.13 μM, little work has examined the solid-phase accumulation of U during field-scale biostimulation. The lack of information on solid-phase U accumulation is due in large part to the difficulty of obtaining comparable pre- and post-biostimulation field sediment samples. In addition, the relatively low (<10 ppm) U concentrations present in most sediments preclude the use of spectroscopic techniques such as XAS for examining solid-phase U speciation. However, a recently developed technique of performing column experiments in situ has allowed us to overcome both of these problems, obtaining sediment samples which were exposed to the same biogeochemical conditions as subsurface sediments during the course of biostimulation. During the 2010 Rifle IFRC field experiment (dubbed "Super 8"), a number of in situ columns were deployed in various wells representing regions of the aquifer affected by acetate amendment (ambient bicarbonate) and concomitant acetate and bicarbonate amendment (elevated bicarbonate). Elevated levels of bicarbonate have been shown to cause desorption of U(VI) from the solid phase at the Rifle site under non-stimulated conditions, resulting in higher dissolved U(VI) concentrations in the aquifer. The Super 8 field experiment was designed in part to test the effect of elevated bicarbonate concentrations on U sequestration during biostimulation. Results from this experiment

Fabrication Processes to Generate Concentration Gradients in Polymer Solar Cell Active Layers

PubMed Central

Inaba, Shusei; Vohra, Varun

2017-01-01

Polymer solar cells (PSCs) are considered as one of the most promising low-cost alternatives for renewable energy production with devices now reaching power conversion efficiencies (PCEs) above the milestone value of 10%. These enhanced performances were achieved by developing new electron-donor (ED) and electron-acceptor (EA) materials as well as finding the adequate morphologies in either bulk heterojunction or sequentially deposited active layers. In particular, producing adequate vertical concentration gradients with higher concentrations of ED and EA close to the anode and cathode, respectively, results in an improved charge collection and consequently higher photovoltaic parameters such as the fill factor. In this review, we relate processes to generate active layers with ED–EA vertical concentration gradients. After summarizing the formation of such concentration gradients in single layer active layers through processes such as annealing or additives, we will verify that sequential deposition of multilayered active layers can be an efficient approach to remarkably increase the fill factor and PCE of PSCs. In fact, applying this challenging approach to fabricate inverted architecture PSCs has the potential to generate low-cost, high efficiency and stable devices, which may revolutionize worldwide energy demand and/or help develop next generation devices such as semi-transparent photovoltaic windows. PMID:28772878

Fabrication Processes to Generate Concentration Gradients in Polymer Solar Cell Active Layers.

PubMed

Inaba, Shusei; Vohra, Varun

2017-05-09

Polymer solar cells (PSCs) are considered as one of the most promising low-cost alternatives for renewable energy production with devices now reaching power conversion efficiencies (PCEs) above the milestone value of 10%. These enhanced performances were achieved by developing new electron-donor (ED) and electron-acceptor (EA) materials as well as finding the adequate morphologies in either bulk heterojunction or sequentially deposited active layers. In particular, producing adequate vertical concentration gradients with higher concentrations of ED and EA close to the anode and cathode, respectively, results in an improved charge collection and consequently higher photovoltaic parameters such as the fill factor. In this review, we relate processes to generate active layers with ED-EA vertical concentration gradients. After summarizing the formation of such concentration gradients in single layer active layers through processes such as annealing or additives, we will verify that sequential deposition of multilayered active layers can be an efficient approach to remarkably increase the fill factor and PCE of PSCs. In fact, applying this challenging approach to fabricate inverted architecture PSCs has the potential to generate low-cost, high efficiency and stable devices, which may revolutionize worldwide energy demand and/or help develop next generation devices such as semi-transparent photovoltaic windows.

Activity-Based Probes for Isoenzyme- and Site-Specific Functional Characterization of Glutathione S -Transferases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoddard, Ethan G.; Killinger, Bryan J.; Nair, Reji N.

Glutathione S-transferases (GSTs) comprise a highly diverse family of phase II drug metabolizing enzymes whose shared function is the conjugation of reduced glutathione to various endo- and xenobiotics. Although the conglomerate activity of these enzymes can be measured by colorimetric assays, measurement of the individual contribution from specific isoforms and their contribution to the detoxification of xenobiotics in complex biological samples has not been possible. For this reason, we have developed two activity-based probes that characterize active glutathione transferases in mammalian tissues. The GST active site is comprised of a glutathione binding “G site” and a distinct substrate binding “Hmore » site”. Therefore, we developed (1) a glutathione-based photoaffinity probe (GSH-ABP) to target the “G site”, and (2) a probe designed to mimic a substrate molecule and show “H site” activity (GST-ABP). The GSH-ABP features a photoreactive moiety for UV-induced covalent binding to GSTs and glutathione-binding enzymes. The GST-ABP is a derivative of a known mechanism-based GST inhibitor that binds within the active site and inhibits GST activity. Validation of probe targets and “G” and “H” site specificity was carried out using a series of competitors in liver homogenates. Herein, we present robust tools for the novel characterization of enzyme- and active site-specific GST activity in mammalian model systems.« less

Identification and characterization of the sodium-binding site of activated protein C.

PubMed

He, X; Rezaie, A R

1999-02-19

Activated protein C (APC) requires both Ca2+ and Na+ for its optimal catalytic function. In contrast to the Ca2+-binding sites, the Na+-binding site(s) of APC has not been identified. Based on a recent study with thrombin, the 221-225 loop is predicted to be a potential Na+-binding site in APC. The sequence of this loop is not conserved in trypsin. We engineered a Gla domainless form of protein C (GDPC) in which the 221-225 loop was replaced with the corresponding loop of trypsin. We found that activated GDPC (aGDPC) required Na+ (or other alkali cations) for its amidolytic activity with dissociation constant (Kd(app)) = 44.1 +/- 8.6 mM. In the presence of Ca2+, however, the requirement for Na+ by aGDPC was eliminated, and Na+ stimulated the cleavage rate 5-6-fold with Kd(app) = 2.3 +/- 0.3 mM. Both cations were required for efficient factor Va inactivation by aGDPC. In the presence of Ca2+, the catalytic function of the mutant was independent of Na+. Unlike aGDPC, the mutant did not discriminate among monovalent cations. We conclude that the 221-225 loop is a Na+-binding site in APC and that an allosteric link between the Na+ and Ca2+ binding loops modulates the structure and function of this anticoagulant enzyme.

Orthogonal use of a human tRNA synthetase active site to achieve multi-functionality

PubMed Central

Zhou, Quansheng; Kapoor, Mili; Guo, Min; Belani, Rajesh; Xu, Xiaoling; Kiosses, William B.; Hanan, Melanie; Park, Chulho; Armour, Eva; Do, Minh-Ha; Nangle, Leslie A.; Schimmel, Paul; Yang, Xiang-Lei

2011-01-01

Protein multi-functionality is an emerging explanation for the complexity of higher organisms. In this regard, while aminoacyl tRNA synthetases catalyze amino acid activation for protein synthesis, some also act in pathways for inflammation, angiogenesis, and apoptosis. How multiple functions evolved and their relationship to the active site is not clear. Here structural modeling analysis, mutagenesis, and cell-based functional studies show that the potent angiostatic, natural fragment of human TrpRS associates via Trp side chains that protrude from the cognate cellular receptor VE-cadherin. Modeling indicates that (I prefer the way it was because the conclusion was reached not only by modeling, but more so by experimental studies.)VE-cadherin Trp side chains fit into the Trp-specific active site of the synthetase. Thus, specific side chains of the receptor mimic (?) amino acid substrates and expand the functionality of the active site of the synthetase. We propose that orthogonal use of the same active site may be a general way to develop multi-functionality of human tRNA synthetases and other proteins. PMID:20010843

Active Layer and Moisture Measurements for Intensive Site 0 and 1, Barrow, Alaska

DOE Data Explorer

John Peterson

2015-04-17

These are measurements of Active Layer Thickness collected along several lines beginning in September, 2011 to the present. The data were collected at several time periods along the Site0 L2 Line, the Site1 AB Line, and an ERT Monitoring Line near Area A in Site1.

Stereospecific suppression of active site mutants by methylphosphonate substituted substrates reveals the stereochemical course of site-specific DNA recombination

PubMed Central

Rowley, Paul A.; Kachroo, Aashiq H.; Ma, Chien-Hui; Maciaszek, Anna D.; Guga, Piotr; Jayaram, Makkuni

2015-01-01

Tyrosine site-specific recombinases, which promote one class of biologically important phosphoryl transfer reactions in DNA, exemplify active site mechanisms for stabilizing the phosphate transition state. A highly conserved arginine duo (Arg-I; Arg-II) of the recombinase active site plays a crucial role in this function. Cre and Flp recombinase mutants lacking either arginine can be rescued by compensatory charge neutralization of the scissile phosphate via methylphosphonate (MeP) modification. The chemical chirality of MeP, in conjunction with mutant recombinases, reveals the stereochemical contributions of Arg-I and Arg-II. The SP preference of the native reaction is specified primarily by Arg-I. MeP reaction supported by Arg-II is nearly bias-free or RP-biased, depending on the Arg-I substituent. Positional conservation of the arginines does not translate into strict functional conservation. Charge reversal by glutamic acid substitution at Arg-I or Arg-II has opposite effects on Cre and Flp in MeP reactions. In Flp, the base immediately 5′ to the scissile MeP strongly influences the choice between the catalytic tyrosine and water as the nucleophile for strand scission, thus between productive recombination and futile hydrolysis. The recombinase active site embodies the evolutionary optimization of interactions that not only favor the normal reaction but also proscribe antithetical side reactions. PMID:25999343

Systematic Functional Analysis of Active-Site Residues in l-Threonine Dehydrogenase from Thermoplasma volcanium

DOE Office of Scientific and Technical Information (OSTI.GOV)

Desjardins, Morgan; Mak, Wai Shun; O’Brien, Terrence E.

Enzymes have been through millions of years of evolution during which their active-site microenvironments are fine-tuned. Active-site residues are commonly conserved within protein families, indicating their importance for substrate recognition and catalysis. In this work, we systematically mutated active-site residues of l-threonine dehydrogenase from Thermoplasma volcanium and characterized the mutants against a panel of substrate analogs. Our results demonstrate that only a subset of these residues plays an essential role in substrate recognition and catalysis and that the native enzyme activity can be further enhanced roughly 4.6-fold by a single point mutation. Kinetic characterization of mutants on substrate analogs showsmore » that l-threonine dehydrogenase possesses promiscuous activities toward other chemically similar compounds not previously observed. Quantum chemical calculations on the hydride-donating ability of these substrates also reveal that this enzyme did not evolve to harness the intrinsic substrate reactivity for enzyme catalysis. Our analysis provides insights into connections between the details of enzyme active-site structure and specific function. Finally, these results are directly applicable to rational enzyme design and engineering.« less

Systematic Functional Analysis of Active-Site Residues in l-Threonine Dehydrogenase from Thermoplasma volcanium

DOE PAGES

Desjardins, Morgan; Mak, Wai Shun; O’Brien, Terrence E.; ...

2017-07-07

Enzymes have been through millions of years of evolution during which their active-site microenvironments are fine-tuned. Active-site residues are commonly conserved within protein families, indicating their importance for substrate recognition and catalysis. In this work, we systematically mutated active-site residues of l-threonine dehydrogenase from Thermoplasma volcanium and characterized the mutants against a panel of substrate analogs. Our results demonstrate that only a subset of these residues plays an essential role in substrate recognition and catalysis and that the native enzyme activity can be further enhanced roughly 4.6-fold by a single point mutation. Kinetic characterization of mutants on substrate analogs showsmore » that l-threonine dehydrogenase possesses promiscuous activities toward other chemically similar compounds not previously observed. Quantum chemical calculations on the hydride-donating ability of these substrates also reveal that this enzyme did not evolve to harness the intrinsic substrate reactivity for enzyme catalysis. Our analysis provides insights into connections between the details of enzyme active-site structure and specific function. Finally, these results are directly applicable to rational enzyme design and engineering.« less

Active site of tripeptidyl peptidase II from human erythrocytes is of the subtilisin type

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tomkinson, B.; Wernstedt, C.; Hellman, U.

1987-11-01

The present report presents evidence that the amino acid sequence around the serine of the active site of human tripeptidyl peptidase II is of the subtilisin type. The enzyme from human erythrocytes was covalently labeled at its active site with (/sup 3/H)diisopropyl fluorophosphate, and the protein was subsequently reduced, alkylated, and digested with trypsin. The labeled tryptic peptides were purified by gel filtration and repeated reversed-phase HPLC, and their amino-terminal sequences were determined. Residue 9 contained the radioactive label and was, therefore, considered to be the active serine residue. The primary structure of the part of the active site (residuesmore » 1-10) containing this residue was concluded to be Xaa-Thr-Gln-Leu-Met-Asx-Gly-Thr-Ser-Met. This amino acid sequence is homologous to the sequence surrounding the active serine of the microbial peptidases subtilisin and thermitase. These data demonstrate that human tripeptidyl peptidase II represents a potentially distinct class of human peptidases and raise the question of an evolutionary relationship between the active site of a mammalian peptidase and that of the subtilisin family of serine peptidases.« less

Active site of tripeptidyl peptidase II from human erythrocytes is of the subtilisin type.

PubMed Central

Tomkinson, B; Wernstedt, C; Hellman, U; Zetterqvist, O

1987-01-01

The present report presents evidence that the amino acid sequence around the serine of the active site of human tripeptidyl peptidase II is of the subtilisin type. The enzyme from human erythrocytes was covalently labeled at its active site with [3H]diisopropyl fluorophosphate, and the protein was subsequently reduced, alkylated, and digested with trypsin. The labeled tryptic peptides were purified by gel filtration and repeated reversed-phase HPLC, and their amino-terminal sequences were determined. Residue 9 contained the radioactive label and was, therefore, considered to be the active serine residue. The primary structure of the part of the active site (residues 1-10) containing this residue was concluded to be Xaa-Thr-Gln-Leu-Met-Asx-Gly-Thr-Ser-Met. This amino acid sequence is homologous to the sequence surrounding the active serine of the microbial peptidases subtilisin and thermitase. These data demonstrate that human tripeptidyl peptidase II represents a potentially distinct class of human peptidases and raise the question of an evolutionary relationship between the active site of a mammalian peptidase and that of the subtilisin family of serine peptidases. PMID:3313395

Natural Radionuclide Activity Concentrations In Spas Of Argentina

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gnoni, G.; Czerniczyniec, M.; Canoba, A.

2008-08-07

Geothermal waters have been used on a large scale for bathing, drinking and medical purposes. These waters can contain natural radionuclides that may increase the exposure to people. In this work the most important natural radionuclide activity concentrations in different thermal spas of Argentina were measured to characterize waters and to evaluate the exposure of workers and members of the public.

Calorimetric studies of the interactions of metalloenzyme active site mimetics with zinc-binding inhibitors.

PubMed

Robinson, Sophia G; Burns, Philip T; Miceli, Amanda M; Grice, Kyle A; Karver, Caitlin E; Jin, Lihua

2016-07-19

The binding of drugs to metalloenzymes is an intricate process that involves several interactions, including binding of the drug to the enzyme active site metal, as well as multiple interactions between the drug and the enzyme residues. In order to determine the free energy contribution of Zn(2+) binding by known metalloenzyme inhibitors without the other interactions, valid active site zinc structural mimetics must be formed and binding studies need to be performed in biologically relevant conditions. The potential of each of five ligands to form a structural mimetic with Zn(2+) was investigated in buffer using Isothermal Titration Calorimetry (ITC). All five ligands formed strong 1 : 1 (ligand : Zn(2+)) binary complexes. The complexes were used in further ITC experiments to study their interaction with 8-hydroxyquinoline (8-HQ) and/or acetohydroxamic acid (AHA), two bidentate anionic zinc-chelating enzyme inhibitors. It was found that tetradentate ligands were not suitable for creating zinc structural mimetics for inhibitor binding in solution due to insufficient coordination sites remaining on Zn(2+). A stable binary complex, [Zn(BPA)](2+), which was formed by a tridentate ligand, bis(2-pyridylmethyl)amine (BPA), was found to bind one AHA in buffer or a methanol : buffer mixture (60 : 40 by volume) at pH 7.25 or one 8-HQ in the methanol : buffer mixture at pH 6.80, making it an effective structural mimetic for the active site of zinc metalloenzymes. These results are consistent with the observation that metalloenzyme active site zinc ions have three residues coordinated to them, leaving one or two sites open for inhibitors to bind. Our findings indicate that Zn(BPA)X2 can be used as an active site structural mimetic for zinc metalloenzymes for estimating the free energy contribution of zinc binding to the overall inhibitor active site interactions. Such use will help aid in the rational design of inhibitors to a variety of zinc metalloenzymes.

Regional specialization in human nuclei: visualization of discrete sites of transcription by RNA polymerase III.

PubMed Central

Pombo, A; Jackson, D A; Hollinshead, M; Wang, Z; Roeder, R G; Cook, P R

1999-01-01

Mammalian nuclei contain three different RNA polymerases defined by their characteristic locations and drug sensitivities; polymerase I is found in nucleoli, and polymerases II and III in the nucleoplasm. As nascent transcripts made by polymerases I and II are concentrated in discrete sites, the locations of those made by polymerase III were investigated. HeLa cells were lysed with saponin in an improved 'physiological' buffer that preserves transcriptional activity and nuclear ultrastructure; then, engaged polymerases were allowed to extend nascent transcripts in Br-UTP, before the resulting Br-RNA was immunolabelled indirectly with fluorochromes or gold particles. Biochemical analysis showed that approximately 10 000 transcripts were being made by polymerase III at the moment of lysis, while confocal and electron microscopy showed that these transcripts were concentrated in only approximately 2000 sites (diameter approximately 40 nm). Therefore, each site contains approximately five active polymerases. These sites contain specific subunits of polymerase III, but not the hyperphosphorylated form of the largest subunit of polymerase II. The results indicate that the active forms of all three nuclear polymerases are concentrated in their own dedicated transcription sites or 'factories', suggesting that different regions of the nucleus specialize in the transcription of different types of gene. PMID:10205177

Quantum mechanics study of the hydroxyethylamines-BACE-1 active site interaction energies

NASA Astrophysics Data System (ADS)

Gueto-Tettay, Carlos; Drosos, Juan Carlos; Vivas-Reyes, Ricardo

2011-06-01

The identification of BACE-1, a key enzyme in the production of Amyloid-β (Aβ) peptides, generated by the proteolytic processing of amyloid precursor protein, was a major advance in the field of Alzheimer's disease as this pathology is characterized by the presence of extracellular senile plaques, mainly comprised of Aβ peptides. Hydroxyethylamines have demonstrated a remarkable potential, like candidate drugs, for this disease using BACE-1 as target. Density Functional Theory calculations were employed to estimate interaction energies for the complexes formed between the hydroxyethylamine derivated inhibitors and 24 residues in the BACE-1 active site. The collected data offered not only a general but a particular quantitative description that gives a deep insight of the interactions in the active site, showing at the same time how ligand structural variations affect them. Polar interactions are the major energetic contributors for complex stabilization and those ones with charged aspartate residues are highlighted, as they contribute over 90% of the total attractive interaction energy. Ligand-ARG296 residue interaction reports the most repulsive value and decreasing of the magnitude of this repulsion can be a key feature for the design of novel and more potent BACE-1 inhibitors. Also it was explained why sultam derivated BACE-1 inhibitors are better ones than lactam based. Hydrophobic interactions concentrated at S1 zone and other relevant repulsions and attractions were also evaluated. The comparison of two different theory levels (X3LYP and M062X) allowed to confirm the relevance of the detected interactions as each theory level has its own strength to depict the forces involved, as is the case of M062X which is better describing the hydrophobic interactions. Those facts were also evaluated and confirmed by comparing the quantitative trend, of selected ligand-residue interactions, with MP2 theory level as reference standard method for electrostatic plus

Quantum mechanics study of the hydroxyethylamines-BACE-1 active site interaction energies.

PubMed

Gueto-Tettay, Carlos; Drosos, Juan Carlos; Vivas-Reyes, Ricardo

2011-06-01

The identification of BACE-1, a key enzyme in the production of Amyloid-β (Aβ) peptides, generated by the proteolytic processing of amyloid precursor protein, was a major advance in the field of Alzheimer's disease as this pathology is characterized by the presence of extracellular senile plaques, mainly comprised of Aβ peptides. Hydroxyethylamines have demonstrated a remarkable potential, like candidate drugs, for this disease using BACE-1 as target. Density Functional Theory calculations were employed to estimate interaction energies for the complexes formed between the hydroxyethylamine derivated inhibitors and 24 residues in the BACE-1 active site. The collected data offered not only a general but a particular quantitative description that gives a deep insight of the interactions in the active site, showing at the same time how ligand structural variations affect them. Polar interactions are the major energetic contributors for complex stabilization and those ones with charged aspartate residues are highlighted, as they contribute over 90% of the total attractive interaction energy. Ligand-ARG296 residue interaction reports the most repulsive value and decreasing of the magnitude of this repulsion can be a key feature for the design of novel and more potent BACE-1 inhibitors. Also it was explained why sultam derivated BACE-1 inhibitors are better ones than lactam based. Hydrophobic interactions concentrated at S1 zone and other relevant repulsions and attractions were also evaluated. The comparison of two different theory levels (X3LYP and M062X) allowed to confirm the relevance of the detected interactions as each theory level has its own strength to depict the forces involved, as is the case of M062X which is better describing the hydrophobic interactions. Those facts were also evaluated and confirmed by comparing the quantitative trend, of selected ligand-residue interactions, with MP2 theory level as reference standard method for electrostatic plus

Influences of aerosol physiochemical properties and new particle formation on CCN activity from observation at a suburban site of China

NASA Astrophysics Data System (ADS)

Li, Yanan; Zhang, Fang; Li, Zhanqing; Sun, Li; Wang, Zhenzhu; Li, Ping; Sun, Yele; Ren, Jingye; Wang, Yuying; Cribb, Maureen; Yuan, Cheng

2017-05-01

With the aim of understanding the impact of aerosol particle size and chemical composition on CCN activity, the size-resolved cloud condensation nuclei (CCN) number concentration (NCCN), particle number size distribution (PSD) (10-600 nm), and bulk chemical composition of particles with a diameter < 1.0 μm (PM1) were measured simultaneously at Xinzhou, a suburban site in northern China, from 22 July to 26 August 2014. The NCCN was measured at five different supersaturations (SS) ranging from 0.075%-0.76%. Diurnal variations in the aerosol number concentration (NCN), NCCN, the bulk aerosol activation ratio (AR), the hygroscopicity parameter (κchem), and the ratio of 44 mass to charge ration (m/z 44) to total organic signal in the component spectrum (f44), and the PSD were examined integrally to study the influence of particle size and chemical composition on CCN activation. We found that particle size was more related to the CCN activation ratios in the morning, whereas in the afternoon ( 1400 LST), κchem and f44 were more closely associated with the bulk AR. Assuming the internal mixing of aerosol particles, NCCN was estimated using the bulk chemical composition and real-time PSD. We found that the predicted CCN number concentrations were underestimated by 20-30% at SS < 0.2% probably due to the measurement uncertainties. Estimates were more accurate at higher SS levels, suggesting that the hygroscopicity parameter based on bulk chemical composition information can provide a good estimate of CCN number concentrations. We studied the impacts of new particle formation (NPF) events on size-resolved CCN activity at the ;growth; stage and ;leveling-off; stage during a typical NPF event by comparing with the case during non-NPF event. It has been found that CCN activation was restrained at the ;growth; stage during which larger particle diameters were needed to reach an activation diameter(Da), and the bulk AR decreased as well. However, during the ;leveling

Coarse particulate matter concentrations from residential outdoor sites associated with the North Carolina Asthma and Children's Environment Studies (NC-ACES)

NASA Astrophysics Data System (ADS)

Chen, Fu-Lin; Williams, Ronald; Svendsen, Erik; Yeatts, Karin; Creason, John; Scott, James; Terrell, Dock; Case, Martin

Coarse particulate matter (PM 10) concentration data from residential outdoor sites were collected using portable samplers as part of an exposure assessment for the North Carolina Asthma and Children's Environment Studies (NC-ACES). PM 10 values were estimated using the differential between independent PM 10 and PM 2.5 collocated MiniVol measurements. Repeated daily 24-h integrated PM 10 and PM 2.5 residential outdoor monitoring was performed at a total of 26 homes during September 2003-June 2004 in the Research Triangle Park, NC area. This effort resulted in the collection of 73 total daily measurements. This assessment was conducted to provide data needed to investigate the association of exposures to coarse particle PM mass concentrations with observed human health effects. Potential instrument bias between the differential MiniVol methodology and a dichotomous sampler were investigated. Results indicated that minimal bias of PM 10 mass concentration estimates (slope = 0.8, intercept =0.36μg m -3) existed between the dichotomous and differential MiniVol procedures. Residential outdoor PM 10 mass concentrations were observed to be highly variable across measurement days and ranged from 1.1 to 12.6μg m -3 (mean of 5.4μg m -3). An average correlation coefficient of r=0.75 existed between residential outdoor PM 10 mass concentrations and those obtained from the central ambient monitoring site. Temporal and spatial variability of PM 10 mass concentrations during the study were observed and are described in this report.

Active site electrostatics protect genome integrity by blocking abortive hydrolysis during DNA recombination

PubMed Central

Ma, Chien-Hui; Rowley, Paul A; Macieszak, Anna; Guga, Piotr; Jayaram, Makkuni

2009-01-01

Water, acting as a rogue nucleophile, can disrupt transesterification steps of important phosphoryl transfer reactions in DNA and RNA. We have unveiled this risk, and identified safeguards instituted against it, during strand cleavage and joining by the tyrosine site-specific recombinase Flp. Strand joining is threatened by a latent Flp endonuclease activity (type I) towards the 3′-phosphotyrosyl intermediate resulting from strand cleavage. This risk is not alleviated by phosphate electrostatics; neutralizing the negative charge on the scissile phosphate through methylphosphonate (MeP) substitution does not stimulate type I endonuclease. Rather, protection derives from the architecture of the recombination synapse and conformational dynamics within it. Strand cleavage is protected against water by active site electrostatics. Replacement of the catalytic Arg-308 of Flp by alanine, along with MeP substitution, elicits a second Flp endonuclease activity (type II) that directly targets the scissile phosphodiester bond in DNA. MeP substitution, combined with appropriate active site mutations, will be useful in revealing anti-hydrolytic mechanisms engendered by systems that mediate DNA relaxation, DNA transposition, site-specific recombination, telomere resolution, RNA splicing and retrohoming of mobile introns. PMID:19440204

Large-scale production and properties of human plasma-derived activated Factor VII concentrate.

PubMed

Tomokiyo, K; Yano, H; Imamura, M; Nakano, Y; Nakagaki, T; Ogata, Y; Terano, T; Miyamoto, S; Funatsu, A

2003-01-01

An activated Factor VII (FVIIa) concentrate, prepared from human plasma on a large scale, has to date not been available for clinical use for haemophiliacs with antibodies against FVIII and FIX. In the present study, we attempted to establish a large-scale manufacturing process to obtain plasma-derived FVIIa concentrate with high recovery and safety, and to characterize its biochemical and biological properties. FVII was purified from human cryoprecipitate-poor plasma, by a combination of anion exchange and immunoaffinity chromatography, using Ca2+-dependent anti-FVII monoclonal antibody. To activate FVII, a FVII preparation that was nanofiltered using a Bemberg Microporous Membrane-15 nm was partially converted to FVIIa by autoactivation on an anion-exchange resin. The residual FVII in the FVII and FVIIa mixture was completely activated by further incubating the mixture in the presence of Ca2+ for 18 h at 10 degrees C, without any additional activators. For preparation of the FVIIa concentrate, after dialysis of FVIIa against 20 mm citrate, pH 6.9, containing 13 mm glycine and 240 mm NaCl, the FVIIa preparation was supplemented with 2.5% human albumin (which was first pasteurized at 60 degrees C for 10 h) and lyophilized in vials. To inactivate viruses contaminating the FVIIa concentrate, the lyophilized product was further heated at 65 degrees C for 96 h in a water bath. Total recovery of FVII from 15 000 l of plasma was approximately 40%, and the FVII preparation was fully converted to FVIIa with trace amounts of degraded products (FVIIabeta and FVIIagamma). The specific activity of the FVIIa was approximately 40 U/ micro g. Furthermore, virus-spiking tests demonstrated that immunoaffinity chromatography, nanofiltration and dry-heating effectively removed and inactivated the spiked viruses in the FVIIa. These results indicated that the FVIIa concentrate had both high specific activity and safety. We established a large-scale manufacturing process of human plasma

Relationship among Phosphorus Circulation Activity, Bacterial Biomass, pH, and Mineral Concentration in Agricultural Soil.

PubMed

Adhikari, Dinesh; Jiang, Tianyi; Kawagoe, Taiki; Kai, Takamitsu; Kubota, Kenzo; Araki, Kiwako S; Kubo, Motoki

2017-12-04

Improvement of phosphorus circulation in the soil is necessary to enhance phosphorus availability to plants. Phosphorus circulation activity is an index of soil's ability to supply soluble phosphorus from organic phosphorus in the soil solution. To understand the relationship among phosphorus circulation activity; bacterial biomass; pH; and Fe, Al, and Ca concentrations (described as mineral concentration in this paper) in agricultural soil, 232 soil samples from various agricultural fields were collected and analyzed. A weak relationship between phosphorus circulation activity and bacterial biomass was observed in all soil samples ( R ² = 0.25), and this relationship became significantly stronger at near-neutral pH (6.0-7.3; R ² = 0.67). No relationship between phosphorus circulation activity and bacterial biomass was observed at acidic (pH < 6.0) or alkaline (pH > 7.3) pH. A negative correlation between Fe and Al concentrations and phosphorus circulation activity was observed at acidic pH ( R ² = 0.72 and 0.73, respectively), as well as for Ca at alkaline pH ( R ² = 0.64). Therefore, bacterial biomass, pH, and mineral concentration should be considered together for activation of phosphorus circulation activity in the soil. A relationship model was proposed based on the effects of bacterial biomass and mineral concentration on phosphorus circulation activity. The suitable conditions of bacterial biomass, pH, and mineral concentration for phosphorus circulation activity could be estimated from the relationship model.

Using Carbohydrate Interaction Assays to Reveal Novel Binding Sites in Carbohydrate Active Enzymes.

PubMed

Cockburn, Darrell; Wilkens, Casper; Dilokpimol, Adiphol; Nakai, Hiroyuki; Lewińska, Anna; Abou Hachem, Maher; Svensson, Birte

2016-01-01

Carbohydrate active enzymes often contain auxiliary binding sites located either on independent domains termed carbohydrate binding modules (CBMs) or as so-called surface binding sites (SBSs) on the catalytic module at a certain distance from the active site. The SBSs are usually critical for the activity of their cognate enzyme, though they are not readily detected in the sequence of a protein, but normally require a crystal structure of a complex for their identification. A variety of methods, including affinity electrophoresis (AE), insoluble polysaccharide pulldown (IPP) and surface plasmon resonance (SPR) have been used to study auxiliary binding sites. These techniques are complementary as AE allows monitoring of binding to soluble polysaccharides, IPP to insoluble polysaccharides and SPR to oligosaccharides. Here we show that these methods are useful not only for analyzing known binding sites, but also for identifying new ones, even without structural data available. We further verify the chosen assays discriminate between known SBS/CBM containing enzymes and negative controls. Altogether 35 enzymes are screened for the presence of SBSs or CBMs and several novel binding sites are identified, including the first SBS ever reported in a cellulase. This work demonstrates that combinations of these methods can be used as a part of routine enzyme characterization to identify new binding sites and advance the study of SBSs and CBMs, allowing them to be detected in the absence of structural data.

Using Carbohydrate Interaction Assays to Reveal Novel Binding Sites in Carbohydrate Active Enzymes

PubMed Central

Wilkens, Casper; Dilokpimol, Adiphol; Nakai, Hiroyuki; Lewińska, Anna; Abou Hachem, Maher; Svensson, Birte

2016-01-01

Carbohydrate active enzymes often contain auxiliary binding sites located either on independent domains termed carbohydrate binding modules (CBMs) or as so-called surface binding sites (SBSs) on the catalytic module at a certain distance from the active site. The SBSs are usually critical for the activity of their cognate enzyme, though they are not readily detected in the sequence of a protein, but normally require a crystal structure of a complex for their identification. A variety of methods, including affinity electrophoresis (AE), insoluble polysaccharide pulldown (IPP) and surface plasmon resonance (SPR) have been used to study auxiliary binding sites. These techniques are complementary as AE allows monitoring of binding to soluble polysaccharides, IPP to insoluble polysaccharides and SPR to oligosaccharides. Here we show that these methods are useful not only for analyzing known binding sites, but also for identifying new ones, even without structural data available. We further verify the chosen assays discriminate between known SBS/CBM containing enzymes and negative controls. Altogether 35 enzymes are screened for the presence of SBSs or CBMs and several novel binding sites are identified, including the first SBS ever reported in a cellulase. This work demonstrates that combinations of these methods can be used as a part of routine enzyme characterization to identify new binding sites and advance the study of SBSs and CBMs, allowing them to be detected in the absence of structural data. PMID:27504624

Active-site protein dynamics and solvent accessibility in native Achromobacter cycloclastes copper nitrite reductase.

PubMed

Sen, Kakali; Horrell, Sam; Kekilli, Demet; Yong, Chin W; Keal, Thomas W; Atakisi, Hakan; Moreau, David W; Thorne, Robert E; Hough, Michael A; Strange, Richard W

2017-07-01

Microbial nitrite reductases are denitrifying enzymes that are a major component of the global nitrogen cycle. Multiple structures measured from one crystal (MSOX data) of copper nitrite reductase at 240 K, together with molecular-dynamics simulations, have revealed protein dynamics at the type 2 copper site that are significant for its catalytic properties and for the entry and exit of solvent or ligands to and from the active site. Molecular-dynamics simulations were performed using different protonation states of the key catalytic residues (Asp CAT and His CAT ) involved in the nitrite-reduction mechanism of this enzyme. Taken together, the crystal structures and simulations show that the Asp CAT protonation state strongly influences the active-site solvent accessibility, while the dynamics of the active-site 'capping residue' (Ile CAT ), a determinant of ligand binding, are influenced both by temperature and by the protonation state of Asp CAT . A previously unobserved conformation of Ile CAT is seen in the elevated temperature series compared with 100 K structures. DFT calculations also show that the loss of a bound water ligand at the active site during the MSOX series is consistent with reduction of the type 2 Cu atom.

An active site-tail interaction in the structure of hexahistidine-tagged Thermoplasma acidophilum citrate synthase

DOE PAGES

Murphy, Jesse R.; Donini, Stefano; Kappock, T. Joseph

2015-10-01

Citrate synthase (CS) plays a central metabolic role in aerobes and many other organisms. The CS reaction comprises two half-reactions: a Claisen aldol condensation of acetyl-CoA (AcCoA) and oxaloacetate (OAA) that forms citryl-CoA (CitCoA), and CitCoA hydrolysis. Protein conformational changes that `close' the active site play an important role in the assembly of a catalytically competent condensation active site. CS from the thermoacidophile Thermoplasma acidophilum (TpCS) possesses an endogenous Trp fluorophore that can be used to monitor the condensation reaction. The 2.2 Å resolution crystal structure of TpCS fused to a C-terminal hexahistidine tag (TpCSH6) reported here is an `open'more » structure that, when compared with several liganded TpCS structures, helps to define a complete path for active-site closure. One active site in each dimer binds a neighboring His tag, the first nonsubstrate ligand known to occupy both the AcCoA and OAA binding sites. Solution data collectively suggest that this fortuitous interaction is stabilized by the crystalline lattice. In conclusion, as a polar but almost neutral ligand, the active site-tail interaction provides a new starting point for the design of bisubstrate-analog inhibitors of CS.« less

An active site-tail interaction in the structure of hexahistidine-tagged Thermoplasma acidophilum citrate synthase.

PubMed

Murphy, Jesse R; Donini, Stefano; Kappock, T Joseph

2015-10-01

Citrate synthase (CS) plays a central metabolic role in aerobes and many other organisms. The CS reaction comprises two half-reactions: a Claisen aldol condensation of acetyl-CoA (AcCoA) and oxaloacetate (OAA) that forms citryl-CoA (CitCoA), and CitCoA hydrolysis. Protein conformational changes that `close' the active site play an important role in the assembly of a catalytically competent condensation active site. CS from the thermoacidophile Thermoplasma acidophilum (TpCS) possesses an endogenous Trp fluorophore that can be used to monitor the condensation reaction. The 2.2 Å resolution crystal structure of TpCS fused to a C-terminal hexahistidine tag (TpCSH6) reported here is an `open' structure that, when compared with several liganded TpCS structures, helps to define a complete path for active-site closure. One active site in each dimer binds a neighboring His tag, the first nonsubstrate ligand known to occupy both the AcCoA and OAA binding sites. Solution data collectively suggest that this fortuitous interaction is stabilized by the crystalline lattice. As a polar but almost neutral ligand, the active site-tail interaction provides a new starting point for the design of bisubstrate-analog inhibitors of CS.

Assessment of elemental concentrations in streams of the New Lead Belt in southeastern Missouri, 2002-05

USGS Publications Warehouse

Brumbaugh, William G.; May, Thomas W.; Besser, John M.; Allert, Ann L.; Schmitt, Christopher J.

2007-01-01

Concerns about possible effects of lead-mining activities on the water quality of federally protected streams located in southeastern Missouri prompted a suite of multidisciplinary studies to be conducted by the U.S. Geological Survey. As part of this investigation, a series of biological studies were initiated in 2001 for streams in the current mining region and the prospecting area. In this report, results are examined for trace elements and other selected chemical measurements in sediment, surface water, and sediment interstitial (pore) water sampled between 2002 and 2005 in association with these biological studies. Compared to reference sites, fine sediments collected downstream from mining areas were enriched in metals by factors as large as 75 for cadmium, 62 for cobalt, 171 for nickel, 95 for lead, and 150 for zinc. Greatest metal concentrations in sediments collected in 2002 were from sites downstream from mines on Strother Creek, Courtois Creek, and the West Fork Black River. Sediments from sites on Bee Fork, Logan Creek, and Sweetwater Creek also were noticeably enriched in lead. Sediments in Clearwater Lake, at least 75 kilometers downstream from mining activity, had metal concentrations that were 1.5 to 2.1 times greater than sediments in an area of the lake with no upstream mining activity. Longitudinal sampling along three streams in 2004 indicated that sediment metal concentrations decreased considerably a few kilometers downstream from mining activities; however, in Strother Creek some metals were still enriched by a factor of five or more as far as 13 kilometers downstream from the Buick tailings impoundment. Compared with 2002 samples, metals concentrations were dramatically lower in sediments collected in 2004 at an upper West Fork Black River site, presumably because beneficiation operations at the West Fork mill ceased in 2000. Concentrations of metals and sulfate in sediment interstitial (pore) waters generally tracked closely with metal

Non-canonical active site architecture of the radical SAM thiamin pyrimidine synthase.

PubMed

Fenwick, Michael K; Mehta, Angad P; Zhang, Yang; Abdelwahed, Sameh H; Begley, Tadhg P; Ealick, Steven E

2015-03-27

Radical S-adenosylmethionine (SAM) enzymes use a [4Fe-4S] cluster to generate a 5'-deoxyadenosyl radical. Canonical radical SAM enzymes are characterized by a β-barrel-like fold and SAM anchors to the differentiated iron of the cluster, which is located near the amino terminus and within the β-barrel, through its amino and carboxylate groups. Here we show that ThiC, the thiamin pyrimidine synthase in plants and bacteria, contains a tethered cluster-binding domain at its carboxy terminus that moves in and out of the active site during catalysis. In contrast to canonical radical SAM enzymes, we predict that SAM anchors to an additional active site metal through its amino and carboxylate groups. Superimposition of the catalytic domains of ThiC and glutamate mutase shows that these two enzymes share similar active site architectures, thus providing strong evidence for an evolutionary link between the radical SAM and adenosylcobalamin-dependent enzyme superfamilies.

PCB concentrations and activity of sea lamprey Petromyzon marinus vary by sex

USGS Publications Warehouse

Madenjian, Charles P.; Johnson, Nicholas S.; Binder, Thomas R.; Rediske, Richard R.; O'Keefe, James P.

2013-01-01

We determined the polychlorinated biphenyl (PCB) concentrations of 40 male and 40 female adult sea lampreys Petromyzon marinus captured in the Cheboygan River, a tributary to Lake Huron, during May 2011. In addition, we performed a laboratory experiment using passive integrated transponder tags to determine whether male adult sea lampreys were more active than female adult sea lampreys. Sex had a significant effect on PCB concentration, and PCB concentration at a given level of sea lamprey condition was approximately 25 % greater in males than in females. Adjusting for the difference in condition between the sexes, males averaged a 17 % greater PCB concentration compared with females. Results from the laboratory experiment indicated that males were significantly more active than females. The observed sex difference in PCB concentrations was not due to female sea lampreys releasing eggs at spawning because the sea lamprey is semelparous, and we caught the sea lampreys before spawning. Rather, we attributed the sex difference in PCB concentrations to a greater rate of energy expenditure in males compared with females. We proposed that this greater rate of energy expenditure was likely due to greater activity. Our laboratory experiment results supported this hypothesis. A greater resting metabolic rate may also have contributed to a greater rate of energy expenditure. Our findings should eventually be applicable toward improving control of sea lamprey, a pest responsible for considerable damage to fisheries in lakes where it is not native.

[Nuclease activity of the recombinant plancitoxin-1-like proteins with mutations in the active site from Trichinella spiralis].

PubMed

Liao, Chengshui; Wang, Xiaoli; Tian, Wenjing; Zhang, Mengke; Zhang, Chunjie; Li, Yinju; Wu, Tingcai; Cheng, Xiangchao

2017-08-25

Although there are 125 predicted DNase Ⅱ-like family genes in the Trichinella spiralis genome, plancitoxin-1-like (Ts-Pt) contains the HKD motif, a typical conserved region of DNase Ⅱ, in N- and C-terminal. It is generally believed that histidine is the active site in DNase Ⅱ. To study the nuclease activity of recombinant Ts-Pt with mutations in the active site from T. spiralis, different fragments of the mutated Ts-Pt genes were cloned using overlap PCR technique and inserted into the expressing vector pET-28a(+), and transformed into Escherichia coli Rosseta (DE3). The fusion proteins were purified by Ni-NTA affinity chromatography and SDS-PAGE. Nuclease activity of the recombinant proteins was detected by agarose gel electrophoresis and nuclease-zymography. The recombinant plasmids harboring the mutated Ts-Pt genes were constructed and expressed as inclusive body in a prokaryotic expression system. After renaturation in vitro, the recombinant proteins had no nuclease activity according to agarose gel electrophoresis. However, the expressed proteins as inclusive body displayed the ability to degrade DNA after renaturation in gel. And the nuclease activity was not affected after subjected to mutation of active site in N- and C-termini of Ts-Pt. These results provide the basis to study the relationship between DNase Ⅱ-like protein family and infection of T. spiralis.

A Variable Active Site Residue Influences the Kinetics of Response Regulator Phosphorylation and Dephosphorylation.

PubMed

Immormino, Robert M; Silversmith, Ruth E; Bourret, Robert B

2016-10-04

Two-component regulatory systems, minimally composed of a sensor kinase and a response regulator protein, are common mediators of signal transduction in microorganisms. All response regulators contain a receiver domain with conserved active site residues that catalyze the signal activating and deactivating phosphorylation and dephosphorylation reactions. We explored the impact of variable active site position T+1 (one residue C-terminal to the conserved Thr/Ser) on reaction kinetics and signaling fidelity, using wild type and mutant Escherichia coli CheY, CheB, and NarL to represent the three major sequence classes observed across response regulators: Ala/Gly, Ser/Thr, and Val/Ile/Met, respectively, at T+1. Biochemical and structural data together suggested that different amino acids at T+1 impacted reaction kinetics by altering access to the active site while not perturbing overall protein structure. A given amino acid at position T+1 had similar effects on autodephosphorylation in each protein background tested, likely by modulating access of the attacking water molecule to the active site. Similarly, rate constants for CheY autophosphorylation with three different small molecule phosphodonors were consistent with the steric constraints on access to the phosphorylation site arising from combination of specific phosphodonors with particular amino acids at T+1. Because other variable active site residues also influence response regulator phosphorylation biochemistry, we began to explore how context (here, the amino acid at T+2) affected the influence of position T+1 on CheY autocatalytic reactions. Finally, position T+1 affected the fidelity and kinetics of phosphotransfer between sensor kinases and response regulators but was not a primary determinant of their interaction.

Insights into the glycyl radical enzyme active site of benzylsuccinate synthase: a computational study.

PubMed

Bharadwaj, Vivek S; Dean, Anthony M; Maupin, C Mark

2013-08-21

The fumarate addition reaction, catalyzed by the enzyme benzylsuccinate synthase (BSS), is considered to be one of the most intriguing and energetically challenging reactions in biology. BSS belongs to the glycyl radical enzyme family and catalyzes the fumarate addition reaction, which enables microorganisms to utilize hydrocarbons as an energy source under anaerobic conditions. Unfortunately, the extreme sensitivity of the glycyl radical to oxygen has hampered the structural and kinetic characterization of BSS, thereby limiting our knowledge on this enzyme. To enhance our molecular-level understanding of BSS, a computational approach involving homology modeling, docking studies, and molecular dynamics (MD) simulations has been used to deduce the structure of BSS's catalytic subunit (BSSα) and illuminate the molecular basis for the fumarate addition reaction. We have identified two conserved and distinct binding pockets at the BSSα active site: a hydrophobic pocket for toluene binding and a polar pocket for fumaric acid binding. Subsequent dynamical and energetic evaluations have identified Glu509, Ser827, Leu390, and Phe384 as active site residues critical for substrate binding. The orientation of substrates at the active site observed in MD simulations is consistent with experimental observations of the syn addition of toluene to fumaric acid. It is also found that substrate binding tightens the active site and restricts the conformational flexibility of the thiyl radical, leading to hydrogen transfer distances conducive to the proposed reaction mechanism. The stability of substrates at the active site and the occurrence of feasible radical transfer distances between the thiyl radical, substrates, and the active site glycine indicate a substrate-assisted radical transfer pathway governing fumarate addition.

C2-C6 background hydrocarbon concentrations monitored at a roof top and green park site, in Dublin City centre.

PubMed

O'Donoghue, R T; Broderick, B M

2007-09-01

A 5 week monitoring campaign was carried out in Dublin City centre, to establish which site gave a more accurate background city centre estimation: a roof-top or green field site. This background represented a conservative estimate of HC exposure in Dublin City centre, useful for quantifying health effects related to this form of pollution and also for establishing a local background relative to the four surrounding main roads when the wind direction is travelling towards each road with the background receptor upwind. Over the entire monitoring campaign, the lowest concentrations and relative standard deviations were observed at the green field site, regardless of time of day or meteorological effects.

Flexibility and Stability Trade-Off in Active Site of Cold-Adapted Pseudomonas mandelii Esterase EstK.

PubMed

Truongvan, Ngoc; Jang, Sei-Heon; Lee, ChangWoo

2016-06-28

Cold-adapted enzymes exhibit enhanced conformational flexibility, especially in their active sites, as compared with their warmer-temperature counterparts. However, the mechanism by which cold-adapted enzymes maintain their active site stability is largely unknown. In this study, we investigated the role of conserved D308-Y309 residues located in the same loop as the catalytic H307 residue in the cold-adapted esterase EstK from Pseudomonas mandelii. Mutation of D308 and/or Y309 to Ala or deletion resulted in increased conformational flexibility. Particularly, the D308A or Y309A mutant showed enhanced substrate affinity and catalytic rate, as compared with wild-type EstK, via enlargement of the active site. However, all mutant EstK enzymes exhibited reduced thermal stability. The effect of mutation was greater for D308 than Y309. These results indicate that D308 is not preferable for substrate selection and catalytic activity, whereas hydrogen bond formation involving D308 is critical for active site stabilization. Taken together, conformation of the EstK active site is constrained via flexibility-stability trade-off for enzyme catalysis and thermal stability. Our study provides further insights into active site stabilization of cold-adapted enzymes.

Trace element geochemistry in archaeological sites.

PubMed

Jenkins, D A

1989-06-01

Man can leave a geochemical imprint on an archaeological site in several ways. In common with other components of the biosphere, there is a selective enrichment of elements in his body tissues which, upon death and burial, may lead to detectable anomalies. Of elements concentrated in this way P is the most obvious, but Sn could be another possibility worth further investigation. There has been particular geochemical impact due to the progressive use of such metals as Ag, Au, Cu, Pb and Zn during successive cultural stages. Anomalies may thus arise due to recognisable transported ore, slags or artefacts, although there has also often been cryptic redispersion of the metals within a site. Charcoal is one of the commoner finds during excavations, and it has the ability to adsorb and concentrate metals progressively from percolating solutions since the time of its burial: with careful interpretation its analysis may thus provide a valuable historical record, as is illustrated by material from several sites in North Wales. Providing care is taken to interpret results in their particular geochemical and pedochemical context, trace element analysis may thus offer a useful insight into the history of man's activities in an archaeological site.

Study of the adsorption of Cd and Zn onto an activated carbon: Influence of pH, cation concentration, and adsorbent concentration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Seco, A.; Marzal, P.; Gabaldon, C.

1999-06-01

The single adsorption of Cd and Zn from aqueous solutions has been investigated on Scharlau Ca 346 granular activated carbon in a wide range of experimental conditions: pH, metal concentration, and carbon concentration. The results showed the efficiency of the activated carbon as sorbent for both metals. Metal removals increase on raising the pH and carbon concentration, and decrease on raising the initial metal concentration. The adsorption processes have been modeled using the surface complex formation (SCF) Triple Layer Model (TLM). The adsorbent TLM parameters were determined. Modeling has been performed assuming a single surface bidentate species or an overallmore » surface species with fractional stoichiometry. The bidentate stoichiometry successfully predicted cadmium and zinc removals in all the experimental conditions. The Freundlich isotherm has been also checked.« less

Well-Defined Metal-O6 in Metal-Catecholates as a Novel Active Site for Oxygen Electroreduction.