Mechanisms for increased soil C storage with increasing temporal and spatial plant diversity in Agroecosystems

NASA Astrophysics Data System (ADS)

Tiemann, L. K.; Grandy, S.; Marin-Spiotta, E.; Atkinson, E. E.

2012-12-01

Generally, there are positive relationships between plant species diversity and net primary production and other key ecosystem functions. However, the effects of aboveground diversity on soil microbial communities and ecosystem processes they mediate, such as soil C sequestration, remain unclear. In this study, we used an 11-y cropping diversity study where increases in diversity have increased crop yields. At the experimental site, temporal diversity is altered using combinations of annual crop rotations, while spatial diversity is altered using cover crop species. We used five treatments ranging in diversity from one to five species consisting of continuous corn with no cover crop or one cover crop and corn-soy-wheat rotations with no cover, one cover or two cover crop species. We collected soils from four replicate plots of each treatment and measured the distribution of mega- (>2 mm), macro- (0.25-2 mm), and micro- (0.053-0.25 mm) aggregates. Within each aggregate size class, we also measured total soil C and N, permanganate oxidizable C (POXC), extracellular enzyme activities (EEA), and microbial community structure with phospholipid fatty acid (PLFA) analysis. We use these data to address the impacts of both rotational and cover crop diversity on soil physical structure, associated microbial community structure and activity and soil C storage. As spatial diversity increased, we found concurrent increases in mega-aggregate abundance as well as increasing soil C in the mega- and micro-aggregates but not macro-aggregates. The proportion of total soil C in each aggregate size class that is relatively labile (POXC) was highest in the micro-aggregates, as was enzyme activity associated with labile C acquisition across all levels of diversity. Enzyme activity associated with more recalcitrant forms of soil C was highest in the mega-aggregate class, also across all diversity levels; however, the ratio of labile to recalcitrant EEA increased with increasing diversity in the mega- and micro-aggregates. In addition, soil N increased with diversity such that microbial C:N EEA simultaneously decreased in mega-aggregates. We also found that cropping diversity has created distinctive soil microbial communities, highlighted by variation in the abundance of gram positive bacteria and Actinomycetes. Further research will help us determine how these changes in community structure with increasing diversity are related to concomitant changes in aggregation and enzyme activities. We suggest that the additional organic matter inputs from cover crops in the high diversity treatments have increased aggregation processes and C pools. While microbial activity has also increased in association with this increased C availability, the activity of recalcitrant and N-acquiring enzymes has declined, suggesting an overall decrease in SOM mineralization with possible increased SOM stabilization. The addition of crop species in rotation (temporal diversity) had minimal influence on any of the measured parameters. We thus conclude that spatial diversity is a more important driver of soil structure and microbial activity, likely due to the high quality organic matter inputs derived from the leguminous cover crops; however, spatial diversity alone did not lead to the same level of C storage potential as mixtures of temporal and spatial diversity.

Mobility of tethering factor EEA1 on endosomes is decreased upon stimulation of EGF receptor endocytosis in HeLa cells

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

Kosheverova, Vera V., E-mail: kosheverova_vera@incras.ru; Kamentseva, Rimma S., E-mail: rkamentseva@yandex.ru; St. Petersburg State University, 7-9, Universitetskaya nab, St. Petersburg, 199034

Tethering factor EEA1, mediating homotypic fusion of early endosomes, was shown to be localized in membrane-bound state both in serum-deprived and stimulated for EGF receptor endocytosis cells. However, it is not known whether dynamics behavior of EEA1 is affected by EGF stimulation. We investigated EEA1 cytosol-to-membrane exchange rate in interphase HeLa cells by FRAP analysis. The data obtained fitted two-states binding model, with the bulk of membrane-associated EEA1 protein represented by the mobile fraction both in serum-starved and EGF-stimulated cells. Fast recovery state had similar half-times in the two cases: about 1.6 s and 2.8 s, respectively. However, the recovery half-time ofmore » slowly cycled EEA1 fraction significantly increased in EGF-stimulated comparing to serum-starved cells (from 21 to 99 s). We suppose that the retardation of EEA1 fluorescence recovery upon EGF-stimulation may be due to the increase of activated Rab5 on endosomal membranes, the growth of the number of tethering events between EEA1-positive vesicles and their clustering. - Highlights: • EEA1 mobility was compared in serum-starved and EGF-stimulated interphase HeLa cells. • FRAP analysis revealed fast and slow components of EEA1 recovery in both cases. • Stimulation of EGFR endocytosis did not affect fast EEA1 turnover. • EGF stimulation significantly increased half-time of slowly exchanged EEA1 fraction.« less

N enrichment decouples the relation between N and P cycling in the rhizoplane but not in the bulk soil: evidence from an N-manipulation field experiment in a Mediterranean ecosystem

NASA Astrophysics Data System (ADS)

Ulm, Florian; Gouveia, Catarina; Dias, Teresa; Cruz, Cristina

2014-05-01

Increasing deposition of reactive nitrogen (N) is a major threat to global ecosystems' stability. Due to the N cascade, the 'extra' N affects the air, water and soil, climate and ecosystems' stability and biodiversity, costing 70-320 billion € per year in Europe alone. Within the European Union, the Mediterranean Basin is a N-limited biodiversity hotspot where N deposition is expected to increase threefold by 2050. However, most of our knowledge of the impacts of increased N availability on ecosystems comes from northern Europe and America. Since Mediterranean-type ecosystems appear on the 'neglected ecosystems list', in 2007 a unique N-manipulation field experiment (N dose and form) was set up in southern Europe (Portugal). The study site is located in Arrábida Natural Park, Portugal (38°29'N, 9°01'W). Vegetation consists of a dense maquis with skeletal soil (15-20 cm deep). We applied 3 N-treatments: 40 and 80 kg N ha-1 yr-1 of ammonium nitrate (NH4NO3) and 40 kg N ha-1 yr-1 of ammonium (1:1 mix of NH4Cl and (NH4)2SO4). Each treatment had 3 replicates (400 m2 plots). Since Mediterranean ecosystems are co-limited by N and phosphorus (P), increasing N availability is expected to alter N and P cycling, i.e., the no longer limiting N is likely to be channelled into P acquisition. Under increased N availability, we expect that phosphatase production and maintenance is no longer tied to the degradation of N-containing macromolecules. To explore this hypothesis, the extracellular enzyme activity (EEA) patterns of the dominant plant species' roots and surrounding soils were compared. We used a novel technique for EEA profiling to measure the activity of acid phosphatase and N-related enzymes (N-Acetylglucosaminidase and Leucine aminopeptidase). The proportions of roots and soil were determined per 100 g soil-root agglomerate to extrapolate the total turnover of each compartment. The total EEAs expressed per agglomerate were similar in soil and root compartments, even though the latter only contributed 2% to the total. This highlights the importance of the root surface (rhizoplane) as a compartment for nutrient cycling within the ecosystem. In the rhizoplane, linear correlations between N-related enzymes and phosphatase activity observed under natural conditions were lost upon addition of N, while in the soil, and despite the N additions, the linear correlations between N-related enzymes and phosphatase activity were maintained. Since most of the applied N is temporarily stored in the biotic compartment, the plant probably has enough N available to invest in rhizoplane phosphatases or provide microorganisms with the nutrients needed to exhibit higher activities. In contrast, as the inorganic N has been taken up by the plants, the soil microorganisms have to rely on N-containing polymers. Our data suggest that the additions of N decoupled the relation between activities of phosphatase and N-related enzymes in the rhizoplane but not in the bulk soil.

Origin, enzymatic response and fate of dissolved organic matter during flood and non-flood conditions in a river-floodplain system of the Danube (Austria).

PubMed

Sieczko, Anna; Peduzzi, Peter

2014-01-01

Spectroscopic techniques and extracellular enzyme activity measurements were combined with assessments of bacterial secondary production (BSP) to elucidate flood-pulse-linked differences in carbon (C) sources and related microbial processes in a river-floodplain system near Vienna (Austria). Surface connection with the main channel significantly influenced the quantity and quality of dissolved organic matter (DOM) in floodplain backwaters. The highest values of dissolved organic carbon (DOC) and chromophoric DOM (CDOM) were observed during the peak of the flood, when DOC increased from 1.36 to 4.37 mg l -1 and CDOM from 2.94 to 14.32 m -1 . The flood introduced DOC which consisted of more allochthonously-derived, aromatic compounds. Bacterial enzymatic activity, as a proxy to track the response to changes in DOM, indicated elevated utilization of imported allochthonous material. Based on the enzyme measurements, new parameters were calculated: metabolic effort and enzymatic indices (EEA 1 and EEA 2). During connection, bacterial glucosidase and protease activity were dominant, whereas during disconnected phases a switch to lignin degradation (phenol oxidase) occurred. The enzymatic activity analysis revealed that flooding mobilized reactive DOM, which then supported bacterial metabolism. No significant differences in overall BSP between the two phases were detected, indicating that heterogeneous sources of C sufficiently support BSP. The study demonstrates that floods are important for delivering DOM, which, despite its allochthonous origin, is reactive and can be effectively utilized by aquatic bacteria in this river-floodplain systems. The presence of active floodplains, characterized by hydrological connectivity with the main channel, creates the opportunity to process allochthonous DOC. This has potential consequences for carbon flux, enhancing C sequestration and mineralization processes in this river-floodplain system.

Impact of heavy metal on activity of some microbial enzymes in the riverbed sediments: Ecotoxicological implications in the Ganga River (India).

PubMed

Jaiswal, Deepa; Pandey, Jitendra

2018-04-15

We studied the extracellular enzyme activity (EEA) in the riverbed sediment along a 518km gradient of the Ganga River receiving carbon and nutrient load from varied human sources. Also, we tested, together with substrate-driven stimulation, if the heavy metal accumulated in the sediment inhibits enzyme activities. Because pristine values are not available, we considered Dev Prayag, a least polluted site located 624km upstream to main study stretch, as a reference site. There were distinct increases in enzyme activities in the sediment along the study gradient from Dev Prayag, however, between-site differences were in concordance with sediment carbon(C), nitrogen (N) and phosphorus (P). Fluorescein diacetate hydrolysis (FDAase), β-glucosidase (Glu) and protease activities showed positive correlation with C, N and P while alkaline phosphatase was found negatively correlated with P. Enzyme activities were found negatively correlated with heavy metal, although ecological risk index (E R i ) varied with site and metal species. Dynamic fit curves showed significant positive correlation between heavy metal and microbial metabolic quotient (qCO 2 ) indicating a decrease in microbial activity in response to increasing heavy metal concentrations. This study forms the first report linking microbial enzyme activities to regional scale sediment heavy metal accumulation in the Ganga River, suggests that the microbial enzyme activities in the riverbed sediment were well associated with the proportion of C, N and P and appeared to be a sensitive indicator of C, N and P accumulation in the river. Heavy metal accumulated in the sediment inhibits enzyme activities, although C rich sediment showed relatively low toxicity due probably to reduced bioavailability of the metal. The study has relevance from ecotoxicological as well as from biomonitoring perspectives. Copyright © 2017 Elsevier Inc. All rights reserved.

Status, quality and specific needs of Zika virus (ZIKV) diagnostic capacity and capability in National Reference Laboratories for arboviruses in 30 EU/EEA countries, May 2016

PubMed Central

Mögling, Ramona; Zeller, Hervé; Revez, Joana; Koopmans, Marion; Reusken, Chantal

2017-01-01

With international travel, Zika virus (ZIKV) is introduced to Europe regularly. A country's ability to robustly detect ZIKV introduction and local transmission is important to minimise the risk for a ZIKV outbreak. Therefore, sufficient expertise and diagnostic capacity and capability are required in European laboratories. To assess the capacity, quality, operational specifics (guidelines and algorithms), technical and interpretation issues and other possible difficulties that were related to ZIKV diagnostics in European countries, a questionnaire was conducted among national reference laboratories in 30 countries in the European Union/European Economic Area (EU/EEA) in May 2016. While the coverage and capacity of ZIKV diagnostics in the EU/EEA national reference laboratories were found to be adequate, the assessment of the quality and needs indicated several crucial points of improvement that will need support at national and EU/EEA level to improve ZIKV preparedness, response and EU/EEA ZIKV surveillance activities. PMID:28920574

The effectiveness and cost-effectiveness of screening for active tuberculosis among migrants in the EU/EEA: a systematic review.

PubMed

Greenaway, Christina; Pareek, Manish; Abou Chakra, Claire-Nour; Walji, Moneeza; Makarenko, Iuliia; Alabdulkarim, Balqis; Hogan, Catherine; McConnell, Ted; Scarfo, Brittany; Christensen, Robin; Tran, Anh; Rowbotham, Nick; Noori, Teymur; van der Werf, Marieke J; Pottie, Kevin; Matteelli, Alberto; Zenner, Dominik; Morton, Rachael L

2018-04-01

The foreign-born population make up an increasing and large proportion of tuberculosis (TB) cases in European Union/European Economic Area (EU/EEA) low-incidence countries and challenge TB elimination efforts. Methods : We conducted a systematic review to determine effectiveness (yield and performance of chest radiography (CXR) to detect active TB, treatment outcomes and acceptance of screening) and a second systematic review on cost-effectiveness of screening for active TB among migrants living in the EU/EEA. Results : We identified six systematic reviews, one report and three individual studies that addressed our aims. CXR was highly sensitive (98%) but only moderately specific (75%). The yield of detecting active TB with CXR screening among migrants was 350 per 100,000 population overall but ranged widely by host country (110-2,340), migrant type (170-1,192), TB incidence in source country (19-336) and screening setting (220-1,720). The CXR yield was lower (19.6 vs 336/100,000) and the numbers needed to screen were higher (5,076 vs 298) among migrants from source countries with lower TB incidence (≤ 50 compared with ≥ 350/100,000). Cost-effectiveness was highest among migrants originating from high (> 120/100,000) TB incidence countries. The foreign-born had similar or better TB treatment outcomes than those born in the EU/EEA. Acceptance of CXR screening was high (85%) among migrants. Discussion : Screening programmes for active TB are most efficient when targeting migrants from higher TB incidence countries. The limited number of studies identified and the heterogeneous evidence highlight the need for further data to inform screening programmes for migrants in the EU/EEA.

HIV-1 stimulates nuclear entry of amyloid beta via dynamin dependent EEA1 and TGF-β/Smad signaling

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

András, Ibolya E., E-mail: iandras@med.miami; Toborek, Michal, E-mail: mtoborek@med.miami.edu

Clinical evidence indicates increased amyloid deposition in HIV-1-infected brains, which contributes to neurocognitive dysfunction in infected patients. Here we show that HIV-1 exposure stimulates amyloid beta (Aβ) nuclear entry in human brain endothelial cells (HBMEC), the main component of the blood–brain barrier (BBB). Treatment with HIV-1 and/or Aβ resulted in concurrent increase in early endosomal antigen-1 (EEA1), Smad, and phosphorylated Smad (pSmad) in nuclear fraction of HBMEC. A series of inhibition and silencing studies indicated that Smad and EEA1 closely interact by influencing their own nuclear entry; the effect that was attenuated by dynasore, a blocker of GTP-ase activity ofmore » dynamin. Importantly, inhibition of dynamin, EEA1, or TGF-β/Smad effectively attenuated HIV-1-induced Aβ accumulation in the nuclei of HBMEC. The present study indicates that nuclear uptake of Aβ involves the dynamin-dependent EEA1 and TGF-β/Smad signaling pathways. These results identify potential novel targets to protect against HIV-1-associated dysregulation of amyloid processes at the BBB level. - Highlights: • HIV-1 induces nuclear accumulation of amyloid beta (Aβ) in brain endothelial cells. • EEA-1 and TGF-Β/Smad act in concert to regulate nuclear entry of Aβ. • Dynamin appropriates the EEA-1 and TGF-Β/Smad signaling. • Dynamin serves as a master regulator of HIV-1-induced nuclear accumulation of Aβ.« less

Changes in dentist and dental hygienist numbers in the European Union and economic area.

PubMed

Widström, E; Eaton, K A; Luciak-Donsberger, C

2010-08-01

To investigate the extent to which changes in the numbers of dental hygienists and dentists have occurred in the Member States of the European Union and Economic Area (EU/EEA) during the last ten years and discuss the changes in relation to the possibilities of sharing tasks between the two groups. Numbers for active dentists, registered hygienists and EU/EEA member state populations in 2007 were taken from the website of the Council of European Chief Dental Officers (CECDO) (www.cecdo.org) and from CECDO records for the EU/EEA member states in 1998 and for the new EU member states (who joined in 2004 and 2007) in 2000. From these data, population: active dentists, population: registered dental hygienist and active dentists: registered dental hygienist ratios were calculated together with percentage changes in the number of dentists and dental hygienists by member state, between 1998 and 2007 for the old and between 2000 and 2007 for the new EU member states. In 2007, there were a total of 343,922 active dentists and 30,963 registered dental hygienists in the 30 EU/EEA member states plus Switzerland. The mean population to dentist ratio was about 1500:1 and the mean population to dental hygienist ratio (in the 25 states where dental hygienists were registered) was 13,454:1. During the study period, the population of the EU/EEA plus Switzerland increased by less that 3%, the number of dentists increased by 13% and the number dental hygienists by 42%. The overall ratio of active dentists: dental hygienists changed from 18:1 to 11:1. In six of the 30 member states plus Switzerland the population to dental hygienist ratio was between 2000:1 and 6000:1 and the dentist: dental hygienist ratio less than 1:3. Although, most member states educate dental hygienists and their numbers in the EU/EEA during the last 10 years have risen more than the dentist numbers, there are still only a handful countries where the hygienist numbers are great enough to make a significant difference to the delivery of oral health care.

A synthesis theory for the externally excited adaptive system /EEAS/

NASA Technical Reports Server (NTRS)

Horowitz, I. M.; Smay, J. W.; Shapiro, A.

1974-01-01

The externally excited adaptive system (EEAS) is a two-degree-of-freedom feedback system with a nonlinearity which is saturated hard by an external periodic signal. Under certain conditions, the EEAS responds quasi-linearly to command and plant disturbance signals, permitting the development of a quantitative synthesis theory for satisfying system tolerances despite large plant uncertainty. The great advantage of the EEAS is its zero sensitivity to plant gain variations, a property it shares with the self-oscillating adaptive system (SOAS). The EEAS is, however, more flexible than the SOAS in satisfying the quasi-linearity constraints. The essential difference is that in the EEAS the loop transmission bandwidth is not rigorously tied to the 'carrier' signal, as it is in the SOAS. There is a class of problems for which the EEAS is superior to the purely linear system, which in turn is superior to the SOAS. The superiority of the EEAS over the SOAS is especially marked in the case of significant plant disturbances, which generally vitiate a SOAS design.

IQGAP1 promotes CXCR4 chemokine receptor function and trafficking via EEA-1+ endosomes

PubMed Central

Bamidele, Adebowale O.; Kremer, Kimberly N.; Hirsova, Petra; Clift, Ian C.; Gores, Gregory J.; Billadeau, Daniel D.

2015-01-01

IQ motif–containing GTPase-activating protein 1 (IQGAP1) is a cytoskeleton-interacting scaffold protein. CXCR4 is a chemokine receptor that binds stromal cell–derived factor-1 (SDF-1; also known as CXCL12). Both IQGAP1 and CXCR4 are overexpressed in cancer cell types, yet it was unclear whether these molecules functionally interact. Here, we show that depleting IQGAP1 in Jurkat T leukemic cells reduced CXCR4 expression, disrupted trafficking of endocytosed CXCR4 via EEA-1+ endosomes, and decreased efficiency of CXCR4 recycling. SDF-1–induced cell migration and activation of extracellular signal-regulated kinases 1 and 2 (ERK) MAPK were strongly inhibited, even when forced overexpression restored CXCR4 levels. Similar results were seen in KMBC and HEK293 cells. Exploring the mechanism, we found that SDF-1 treatment induced IQGAP1 binding to α-tubulin and localization to CXCR4-containing endosomes and that CXCR4-containing EEA-1+ endosomes were abnormally located distal from the microtubule (MT)-organizing center (MTOC) in IQGAP1-deficient cells. Thus, IQGAP1 critically mediates CXCR4 cell surface expression and signaling, evidently by regulating EEA-1+ endosome interactions with MTs during CXCR4 trafficking and recycling. IQGAP1 may similarly promote CXCR4 functions in other cancer cell types. PMID:26195666

Vaccination policies of immigrants in the EU/EEA Member States-the measles immunization example.

PubMed

Bica, Mihai A; Clemens, Ralf

2018-06-01

In 2015-16, the European Union/European Economic Area Member States (EU/EEA MSs) experienced an unprecedented volume and rate of migration, posing serious challenges to existing national immunization systems and strategies and raising the questions of where, when and who to vaccinate. We assessed existing strategies for vaccinating immigrant populations in the EU/EEA using measles as an example of the most important vaccine-preventable diseases. In this cross-sectional study, conducted from March to May 2016, an electronic questionnaire was sent to the Heads of National Immunization Technical Advisory Groups (NITAGs) or equivalent policy-making bodies in each of the 31 EU/EEA Member States. Responses were entered into a structured database and validated by survey responders for final analysis. Validated responses from all 31 EU/EEA NITAGs or equivalents showed that there is no common measles immunization policy for European immigrants. Policies vary widely from no policy at all (9 of 31, 29%) to vaccination of all comers (2 of 31, 6%), or vaccination of selected cohorts based on vaccination history (17 of 31, 55%) or serum antibody analysis (2 of 31, 6%). Further, the operational responsibilities for immigrant vaccination and documentation methods are not unified within the EU/EEA region. With some notable exceptions immunization policies to contain spread of infectious diseases through migration are either non-existent or vary widely between countries in the EU/EEA. With freedom of movement within the EU/EEA there ought to be harmonization and a common EU/EEA vaccination strategy to replace national policies for immigrant populations.

Enzymatic activities and prokaryotic abundance in relation to organic matter along a West-East Mediterranean transect (TRANSMED cruise).

PubMed

Zaccone, R; Boldrin, A; Caruso, G; La Ferla, R; Maimone, G; Santinelli, C; Turchetto, M

2012-07-01

The distribution of extracellular enzymatic activities (EEA) [leucine aminopeptidase (LAP), ß-glucosidase (GLU), alkaline phosphatase (AP)], as well as that of prokaryotic abundance (PA) and biomass (PB), dissolved organic carbon (DOC), particulate organic carbon and particulate total nitrogen (POC, PTN), was determined in the epi-, meso-, and bathypelagic waters of the Mediterranean Sea along a West-East transect and at one Atlantic station located outside the Strait of Gibraltar. This study represents a synoptical evaluation of the microbial metabolism during early summer. Decreasing trends with depth were observed for most of the parameters (PA, PB, AP, DOC, POC, PTN). Significant differences between the western and eastern basins of the Mediterranean Sea were found, displaying higher rates of LAP and GLU and lower C/N ratios more in the eastern than in the western areas. Conversely, in the epipelagic layer, PA and PB were found to be higher in the western than in the eastern basins. PB was significantly related to DOC concentration (all data, n = 145, r = 0.53, P < 0.01), while significant correlations of EEA with POC and PTN were found in the epipelagic layer, indicating an active response of microbial metabolism to organic substrates. Specific enzyme activities normalized to cell abundance pointed out high values of LAP and GLU in the bathypelagic layer, especially in the eastern basin, while cell-specific AP was high in the epi- and bathypelagic zone of the eastern basin indicating a rapid regeneration of inorganic P for both prokaryotes and phytoplankton needs. Low activity and abundance characterized the Atlantic station, while opposite trends of these parameters were observed along the Mediterranean transect, showing the uncoupling between abundance and activity data. In the east Mediterranean Sea, decomposition processes increased probably in response to mesoscale structures which lead to organic matter downwelling.

Laxative effects of agarwood on low-fiber diet-induced constipation in rats.

PubMed

Kakino, Mamoru; Tazawa, Shigemi; Maruyama, Hiroe; Tsuruma, Kazuhiro; Araki, Yoko; Shimazawa, Masamitsu; Hara, Hideaki

2010-11-15

Agarwood (Aquilaria sinensis), well known as incense in Southeast Asia, has been used as a digestive in traditional medicine. We investigated the laxative effects of an ethanol extract of agarwood leaves (EEA) in a rat model of low-fiber diet-induced constipation. A set of rats was bred on a normal diet while another set was placed on a low-fiber diet to induce constipation. The laxative effect of agarwood was then investigated on both sets of rats. Pretreatment of normal rats with single dose of EEA (600 mg/kg, p.o.) significantly increased frequency and weight of stools. Also, treatments with EEA (300 and 600 mg/kg, p.o.) for 14 days caused a significant increase in stool frequency and weight. Feeding of the animals with a low-fiber diet resulted in a decrease in stool weight, frequency, and water content and also delayed carmine egestion. A single treatment with EEA (600 mg/kg) or senna (150 and 300 mg/kg) significantly increased stool frequency, weight, and water content and also accelerated carmine egestion in the model rats. Once daily administrations of EEA (150 mg/kg), for 14 days, caused a significant increase in water content of stools. The higher doses of EEA (300 and 600 mg/kg) significantly increased frequency, weight, and water content of the stools while accelerating carmine egestion in the constipated rats. Senna (150 and 300 mg/kg) produced similar effect as the higher doses of EEA but, in addition, induced severe diarrhea. These findings indicate that EEA has a laxative effect, without causing diarrhea, in a rat model of low-fiber diet-induced constipation. These findings suggest that EEA may be highly effective on constipation as a complementary medicine in humans suffering from life style-induced constipation.

Comparison of endoscopic endonasal and bifrontal craniotomy approaches for olfactory groove meningiomas: A matched pair analysis of outcomes and frontal lobe changes on MRI.

PubMed

de Almeida, John R; Carvalho, Felipe; Vaz Guimaraes Filho, Francisco; Kiehl, Tim-Rasmus; Koutourousiou, Maria; Su, Shirley; Vescan, Allan D; Witterick, Ian J; Zadeh, Gelareh; Wang, Eric W; Fernandez-Miranda, Juan C; Gardner, Paul A; Gentili, Fred; Snyderman, Carl H

2015-11-01

We compare the outcomes and postoperative MRI changes of endoscopic endonasal (EEA) and bifrontal craniotomy (BFC) approaches for olfactory groove meningiomas (OGM). All patients who underwent either BFC or EEA for OGM were eligible. Matched pairs were created by matching tumor volumes of an EEA patient with a BFC patient, and matching the timing of the postoperative scans. The tumor dimensions, peritumoral edema, resectability issues, and frontal lobe changes were recorded based on preoperative and postoperative MRI. Postoperative fluid-attenuated inversion recovery (FLAIR) hyperintensity and residual cystic cavity (porencephalic cave) volume were compared using univariable and multivariable analyses. From a total of 70 patients (46 EEA, 24 BFC), 10 matched pairs (20 patients) were created. Three patients (30%) in the EEA group and two (20%) in the BFC had postoperative cerebrospinal fluid leaks (p=0.61). Gross total resections were achieved in seven (70%) of the EEA group and nine (90%) of the BFC group (p=0.26), and one patient from each group developed a recurrence. On postoperative MRI, there was no significant difference in FLAIR signal volumes between EEA and BFC approaches (6.9 versus 13.3 cm(3); p=0.17) or in porencephalic cave volumes (1.7 versus 5.0 cm(3); p=0.11) in univariable analysis. However, in a multivariable analysis, EEA was associated with less postoperative FLAIR change (p=0.02) after adjusting for the volume of preoperative edema. This study provides preliminary evidence that EEA is associated with quantifiable improvements in postoperative frontal lobe imaging. Copyright © 2015 Elsevier Ltd. All rights reserved.

Changes in chlamydia control activities in Europe between 2007 and 2012: a cross-national survey

PubMed Central

Sfetcu, Otilia; van der Sande, Marianne A.; Andersen, Berit; Herrmann, Björn; Ward, Helen; Götz, Hannelore M.; Uusküla, Anneli; Woodhall, Sarah C.; Redmond, Shelagh M.; Amato-Gauci, Andrew J.; Low, Nicola; van Bergen, Jan E.

2016-01-01

Background: In 2012, the levels of chlamydia control activities including primary prevention, effective case management with partner management and surveillance were assessed in 2012 across countries in the European Union and European Economic Area (EU/EEA), on initiative of the European Centre for Disease Control (ECDC) survey, and the findings were compared with those from a similar survey in 2007. Methods: Experts in the 30 EU/EEA countries were invited to respond to an online questionnaire; 28 countries responded, of which 25 participated in both the 2007 and 2012 surveys. Analyses focused on 13 indicators of chlamydia prevention and control activities; countries were assigned to one of five categories of chlamydia control. Results: In 2012, more countries than in 2007 reported availability of national chlamydia case management guidelines (80% vs. 68%), opportunistic chlamydia testing (68% vs. 44%) and consistent use of nucleic acid amplification tests (64% vs. 36%). The number of countries reporting having a national sexually transmitted infection control strategy or a surveillance system for chlamydia did not change notably. In 2012, most countries (18/25, 72%) had implemented primary prevention activities and case management guidelines addressing partner management, compared with 44% (11/25) of countries in 2007. Conclusion: Overall, chlamydia control activities in EU/EEA countries strengthened between 2007 and 2012. Several countries still need to develop essential chlamydia control activities, whereas others may strengthen implementation and monitoring of existing activities. PMID:26498953

Value of free-run electromyographic monitoring of lower cranial nerves in endoscopic endonasal approach to skull base surgeries.

PubMed

Thirumala, Parthasarathy D; Mohanraj, Santhosh Kumar; Habeych, Miguel; Wichman, Kelley; Chang, Yue-Fang; Gardner, Paul; Snyderman, Carl; Crammond, Donald J; Balzer, Jeffrey

2012-08-01

Objective The main objective of this study was to evaluate the value of free-run electromyography (f-EMG) monitoring of cranial nerves (CNs) VII, IX, X, XI, and XII in skull base surgeries performed using endoscopic endonasal approach (EEA) to reduce iatrogenic CN deficits. Design We retrospectively identified 73 patients out of 990 patients who had EEA in our institution who had at least one CN monitored. In each CN group, we classified patients who had significant (SG) f-EMG activity as group I and those who did not as group II. Results We monitored a total of 342 CNs. A total of 62 nerves had SG f-EMG activity including CN VII = 18, CN IX = 16, CN X = 13, CN XI = 5, and CN XII = 10. No nerve deficit was found in the nerves that had significant activity during procedure. A total of five nerve deficits including (CN IX = 1, CN X = 2, CN XII = 2) were observed in the group that did not display SG f-EMG activity during surgery. Conclusions f-EMG seems highly sensitive to surgical manipulations and in locating CNs. It seems to have limited value in predicting postoperative neurological deficits. Future studies to evaluate the EMG of lower CNs during EEA procedures need to be done with both f-EMG and triggered EMG.

Value of Free-Run Electromyographic Monitoring of Lower Cranial Nerves in Endoscopic Endonasal Approach to Skull Base Surgeries

PubMed Central

Thirumala, Parthasarathy D.; Mohanraj, Santhosh Kumar; Habeych, Miguel; Wichman, Kelley; Chang, Yue-Fang; Gardner, Paul; Snyderman, Carl; Crammond, Donald J.; Balzer, Jeffrey

2012-01-01

Objective The main objective of this study was to evaluate the value of free-run electromyography (f-EMG) monitoring of cranial nerves (CNs) VII, IX, X, XI, and XII in skull base surgeries performed using endoscopic endonasal approach (EEA) to reduce iatrogenic CN deficits. Design We retrospectively identified 73 patients out of 990 patients who had EEA in our institution who had at least one CN monitored. In each CN group, we classified patients who had significant (SG) f-EMG activity as group I and those who did not as group II. Results We monitored a total of 342 CNs. A total of 62 nerves had SG f-EMG activity including CN VII = 18, CN IX = 16, CN X = 13, CN XI = 5, and CN XII = 10. No nerve deficit was found in the nerves that had significant activity during procedure. A total of five nerve deficits including (CN IX = 1, CN X = 2, CN XII = 2) were observed in the group that did not display SG f-EMG activity during surgery. Conclusions f-EMG seems highly sensitive to surgical manipulations and in locating CNs. It seems to have limited value in predicting postoperative neurological deficits. Future studies to evaluate the EMG of lower CNs during EEA procedures need to be done with both f-EMG and triggered EMG. PMID:23904999

Limitations of the endonasal endoscopic approach in treating olfactory groove meningiomas. A systematic review.

PubMed

Shetty, Sathwik Raviraj; Ruiz-Treviño, Armando S; Omay, Sacit Bulent; Almeida, Joao Paulo; Liang, Buqing; Chen, Yu-Ning; Singh, Harminder; Schwartz, Theodore H

2017-10-01

To review current management strategies for olfactory groove meningioma (OGM)s and the recent literature comparing endoscopic endonasal (EEA) with traditional transcranial (TCA) approaches. A PubMed search of the recent literature (2011-2016) was performed to examine outcomes following EEA and TCA for OGM. The extent of resection, visual outcome, postoperative complications and recurrence rates were analyzed using percentages and proportions, the Fischer exact test and the Student's t-test using Graphpad PRISM 7.0Aa (San Diego, CA) software. There were 444 patients in the TCA group with a mean diameter of 4.61 (±1.17) cm and 101 patients in the EEA group with a mean diameter of 3.55 (± 0.58) cm (p = 0.0589). GTR was achieved in 90.9% (404/444) in the TCA group and 70.2% (71/101) in the EEA group (p < 0.0001). Of the patients with preoperative visual disturbances, 80.7% (21/26) of patients in the EEA cohort had an improvement in vision compared to 12.83%(29/226) in the TCA group (p < 0.0001). Olfaction was lost in 61% of TCA and in 100% of EEA patients. CSF leaks and meningitis occurred in 25.7% and 4.95% of EEA patients and 6.3% and 1.12% of TCA patients, respectively (p < 0.0001; p = 0.023). Our updated literature review demonstrates that despite more experience with endoscopic resection and skull base reconstruction, the literature still supports TCA over EEA with respect to the extent of resection and complications. EEA may be an option in selected cases where visual improvement is the main goal of surgery and postoperative anosmia is acceptable to the patient or in medium-sized tumors with existing preoperative anosmia. Nevertheless, based on our results, it seems more prudent at this time to use TCA for the majority of OGMs.

Defining Stability, Security, Transition, and Reconstruction (SSTR) Operations Requirements for Future Department of the Navy Training and Analytical Models and Simulations

DTIC Science & Technology

2008-09-01

43 B. WELL-DEFINED MEASURES ........................................................... 43 C. ESSENTIAL ELEMENTS OF ANALYSIS ( EEA ...45 D. EEA PROCESS FOR RESTORATION OF ESSENTIAL SERVICE - WATER...FBCB2 Force XXI Battle Command, Brigade-and-Below FM Army Field Manual EEA Essential Elements of Analysis EPG Electronic Proving Ground ESS

Prospective experimental study of transrectal viscerotomy closure using transanal endoscopic suture vs. circular stapler: a step toward NOTES.

PubMed

Diana, M; Leroy, J; Wall, J; De Ruijter, V; Lindner, V; Dhumane, P; Mutter, D; Marescaux, J

2012-06-01

Endoluminal full-thickness closure of the rectal wall is critical in emerging procedures including endoscopic submucosal dissection and transrectal natural orifice transluminal endoscopic surgery (NOTES). This study aimed to compare manual suture using the transanal endoscopic operation platform (TEO; Karl Storz, Tüttlingen, Germany) with the end-to-end anastomosis hemorrhoid circular stapler (EEA; Covidien, Dublin, Ireland) for closure of the rectal viscerotomy during transrectal NOTES segmental colectomy. A total of 12 swine underwent transrectal hybrid NOTES partial colectomies. Animals were divided into two groups according to the viscerotomy closure technique: 1) TEO manual suture; 2) EEA circular stapler closure. Mean (± SD) viscerotomy closure time was 67.5 ± 59.5 minutes and 31.5 ± 19.6 minutes for TEO and EEA, respectively. There was one conversion to laparoscopy in the TEO group and a misfiring in the EEA group that required a TEO salvage suture. There was one positive air-leak test in each group. Peritoneal fluid collected at the end of the procedure tested positive for bacterial contamination in all cases. A mild stenosis was present in 4 /6 viscerotomies (67 %) in the TEO group and in 1/6 (17 %) in the EEA group on endoscopic control. Inflammatory changes were mild in 3/5 (60 %) and 4/5 (80 %) viscerotomies in the TEO and EEA groups, respectively, whereas severe inflammation was found in 2/5 (TEO) and 1 /5 (EEA). Transrectal viscerotomy closure using the EEA circular stapler technique is feasible, easy to perform, and histologically comparable to suture closure through a TEO platform. It may offer an attractive alternative for NOTES segmental colectomies and endoscopic resections. © Georg Thieme Verlag KG Stuttgart · New York.

Pulmonary and percutaneous absorption of 2-propoxyethyl acetate and 2-ethoxyethyl acetate in beagle dogs.

PubMed Central

Guest, D; Hamilton, M L; Deisinger, P J; DiVincenzo, G D

1984-01-01

A comparison was made of the absorption and elimination rates of 2-propoxyethyl acetate (PEA) and 2-ethoxyethyl acetate (EEA) following inhalation, dermal application or IV administration. Male beagle dogs were exposed to 50 ppm PEA or EEA for 5 hr, and breath samples were collected during the exposure and a 3-hr recovery period. Both compounds were rapidly absorbed through the lungs. After 10 min of exposure, the concentrations of the parent compounds in the expired breath were 5 to 10 ppm (80-90% absorption) and reached plateau values at about 3 hr of 13 ppm for PEA (74% absorption) and 16 ppm for EEA (68% absorption). Post-exposure breath samples declined exponentially to 0.5 ppm and 2 ppm after 3 hr for PEA and EEA, respectively. Expired concentrations of PEA were slightly, but significantly (p less than 0.025), lower than those of EEA at corresponding times during the exposure. After IV dosing with 1 mg/kg [ethyl-1,2-14C]PEA, the urine contained 61% and 88% of the dose in 4 and 24 hr, respectively. [14C]EEA was eliminated more slowly, with 20% and 61% of the dose appearing in the urine in 4 and 24 hr, respectively. Blood elimination half-lives were 1.6 hr for [14C]PEA and 7.9 hr for [14C]EEA. Only trace amounts of 14CO2 (less than 1%) or volatile materials (less than 0.1%) were detected in the expired air with either compound. For studies of percutaneous absorption, [14C]PEA or [14C]EEA was added to undiluted compound and applied in a glass cell to a shaved area on a dog's thorax for 30 or 60 min.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6499802

Composition and extracellular enzymatic function of pelagic, particle-associated, and benthic bacterial communities in the central Arctic Ocean

NASA Astrophysics Data System (ADS)

Balmonte, J. P.; Teske, A.; Arnosti, C.

2016-02-01

The structure and function of Arctic bacterial communities have rarely been studied in concert, but are crucial to our understanding of biogeochemical cycles. As the Arctic transitions to become seasonally-ice free, a critical priority is to elucidate the present ecological role and environmental dependence of Arctic bacterial communities. We investigated the depth and regional variations in Central Arctic bacterial community composition (BCC) and extracellular enzymatic activities (EEA)—the initial step in organic matter breakdown—to explore links between community structure and function. Samples were collected across a gradient of sea-ice cover (open ocean, first year ice, multi-year ice) from 79°N to 88°N and from surface to bottom waters ( 3.5 to 4.5 km). Pelagic BCC most strongly varies with hydrography and with particle-association, which likely selects for a specialized community of heterotrophic opportunists; benthic BCC show little regional variation. In contrast, EEA reveal significant depth and regional differences in hydrolysis rates as well as in the spectrum of substrates hydrolyzed. Particle-associated EEA reveal an equal or greater range of enzymatic capabilities than in bulk-seawater measurements, supporting previous findings that particles are hotspots of microbial heterotrophic activity. These patterns suggest a complex relationship between BCC, EEA, and the environment: while water mass characteristics consistently differentiate bacterial communities, additional local factors shape their capabilities to hydrolyze organic matter. Multivariate analyses will be used to further explore the relationships between composition and function as well as their correlations with environmental data. Our findings provide a baseline for future comparisons and initial insight into the functionality and biogeography of Arctic bacterial communities.

Al Basrah Oil Terminal. Basrah, Iraq

DTIC Science & Technology

2007-04-26

made of plastic Conclusion Emergency evacuation capabilities are very important for the protection of personnel working on ABOT. The work... EEA ) Program and Health, Safety, and Environmental (HSE) Program URIs 22671 and 22672 As mentioned earlier in the Lifeboat section of this...accounting of all personnel.” PIJV recommended the development of an EEA plan and HSE program manual for the ABOT personnel. The EEA plan was

Cognitive Agility Measurement in a Complex Environment

DTIC Science & Technology

2017-04-01

correlate with their corresponding historical psychology tests? EEA3.1: Does the variable for Make Goal cognitive flexibility correlate with the...Stroop Test cognitive flexibility variable? EEA3.2: Does the variable for Make Goal cognitive openness correlate with the AUT cognitive openness...variable? EEA3.3: Does the variable for Make Goal focused attention correlate with the Go, No Go Paradigm focused attention variable? 1.6

Leaf-litter leachate concentration promotes heterotrophy in freshwater biofilms: Understanding consequences of water scarcity.

PubMed

Martínez, Aingeru; Kominoski, John Stephen; Larrañaga, Aitor

2017-12-01

Climate change is increasing overall temporal variability in precipitation resulting in a seasonal water availability, both increasing periods of flooding and water scarcity. During low water availability periods, the concentration of leachates from riparian vegetation increases, subsequently increasing dissolved organic matter (DOM). Moreover, shifts in riparian vegetation by land use changes impact the quantity and quality of DOM. Our objective was to test effects of increasing DOM concentrations from Eucalyptus grandis (one of the most cultivated tree species in the world) leachates on the metabolism (respiration, R; gross primary productivity, GPP) and extracellular enzyme activities (EEAs) of freshwater biofilms. To test effects of DOM concentrations on freshwater biofilm functions, we incubated commercial cellulose sponges in a freshwater pond to allow biofilm colonization, and then exposed biofilms to five different concentrations of leaf-litter leachates of E. grandis for five days. To test if responses to DOM concentrations varied with colonization stage of biofilms, we measured treatment effects on biofilms colonizing standard substrates after one, two, three and four weeks of colonization. Increases in leachates concentrations enhanced biofilm heterotrophy, increasing R rates and decreasing GPP. Leachate concentrations did not affect biofilm EEAs, and changes in biofilm metabolism were not explained by treatment-induced changes in biofilm biomass or stoichiometry. We detected the lowest production:respiration ratios, i.e. more heterotrophic assemblages, with the most concentrated leachate solution and the most advanced biofilm colonization stages. Shifts in quantity of dissolved organic matter in freshwaters may further influence ecosystem metabolism and carbon processing. Copyright © 2017 Elsevier B.V. All rights reserved.

Mind the gap: the distributional effects of raising the early eligibility age and full retirement age.

PubMed

Olsen, Anya

2012-01-01

Policymakers have proposed increases to the early eligibility age (EEA) and/or full retirement age (FRA) to address increasing life expectancy and Social Security solvency issues. This analysis uses the Social Security Administration's Modeling Income in the Near Term, version 6 (MINT6) model to compare three retirement-age increases suggested by the Social Security Advisory Board: increase the gap between the EEA and FRA by raising only the FRA, increase both the EEA and FRA to maintain a 4-year gap between them, and increase both the EEA and FRA to maintain a 5-year gap between them. Although all three options would improve system solvency by similar proportions, their effect on individual beneficiaries in the future would vary. Benefit reductions are greater under the proposals with more months between the EEA and FRA, while the option that maintains a 4-year gap results in benefit increases for some beneficiaries compared with current law.

Tuberculosis treatment outcome monitoring in European Union countries: systematic review

PubMed Central

van Hest, Rob; Ködmön, Csaba; Verver, Suzanne; Erkens, Connie G.M.; Straetemans, Masja; Manissero, Davide; de Vries, Gerard

2013-01-01

Treatment success measured by treatment outcome monitoring (TOM) is a key programmatic output of tuberculosis (TB) control programmes. We performed a systematic literature review on national-level TOM in the 30 European Union (EU)/European Economic Areas (EEA) countries to summarise methods used to collect and report data on TOM. Online reference bibliographic databases PubMed/MEDLINE and EMBASE were searched to identify relevant indexed and non-indexed literature published between January 2000 and August 2010. The search strategy resulted in 615 potentially relevant indexed citations, of which 27 full-text national studies (79 data sets) were included for final analysis. The selected studies were performed in 10 EU/EEA countries and gave a fragmented impression of TOM in the EU/EEA. Publication year, study period, sample size, databases, definitions, variables, patient and outcome categories, and population subgroups varied widely, portraying a very heterogeneous picture. This review confirmed previous reports of considerable heterogeneity in publications of TOM results across EU/EEA countries. PubMed/MEDLINE and EMBASE indexed studies are not a suitable instrument to measure representative TOM results for the 30 EU/EEA countries. Uniform and complete reporting to the centralised European Surveillance System will produce the most timely and reliable results of TB treatment outcomes in the EU/EEA. PMID:22790913

Exciton exciton annihilation dynamics in chromophore complexes. II. Intensity dependent transient absorption of the LH2 antenna system.

PubMed

Bruggemann, B; May, V

2004-02-01

Using the multiexciton density matrix theory of excitation energy transfer in chromophore complexes developed in a foregoing paper [J. Chem. Phys. 118, 746 (2003)], the computation of ultrafast transient absorption spectra is presented. Beside static disorder and standard mechanisms of excitation energy dissipation the theory incorporates exciton exciton annihilation (EEA) processes. To elucidate signatures of EEA in intensity dependent transient absorption data the approach is applied to the B850 ring of the LH2 found in rhodobacter sphaeroides. As main indications for two-exciton population and resulting EEA we found (i) a weakening of the dominant single-exciton bleaching structure in the transient absorption, and (ii) an intermediate suppression of long-wavelength and short-wavelength shoulders around the bleaching structure. The suppression is caused by stimulated emission from the two-exciton to the one-exciton state and the return of the shoulders follows from a depletion of two-exciton population according to EEA. The EEA-signature survives as a short-wavelength shoulder in the transient absorption if orientational and energetic disorder are taken into account. Therefore, the observation of the EEA-signatures should be possible when doing frequency resolved transient absorption experiments with a sufficiently strongly varying pump-pulse intensity. Copyright 2004 American Institute of Physics

Seroprevalence and susceptibility to hepatitis A in the European Union and European Economic Area: a systematic review.

PubMed

Carrillo-Santisteve, Paloma; Tavoschi, Lara; Severi, Ettore; Bonfigli, Sandro; Edelstein, Michael; Byström, Emma; Lopalco, Pierluigi

2017-10-01

Most of the European Union (EU) and European Economic Area (EEA) is considered a region of very low hepatitis A virus (HAV) endemicity; however, geographical differences exist. We did a systematic review with the aim of describing seroprevalence and susceptibility in the general population or special groups in the EU and EEA. We searched databases and public health national institutes websites for HAV seroprevalence records published between Jan 1, 1975, and June 30, 2014, with no language restrictions. An updated search was done on Aug 10, 2016. We defined seroprevalence profiles (very low, low, and intermediate) as the proportion of the population with age-specific anti-HAV antibodies at age 15 and 30 years, and susceptibility profiles (low, moderate, high, and very high) as the proportion of susceptible individuals at age 30 and 50 years. We included 228 studies from 28 of 31 EU and EEA countries. For the period 2000-14, 24 countries had a very low seroprevalence profile, compared with five in 1975-89. The susceptibility among adults ranged between low and very high and had a geographical gradient, with three countries in the low susceptibility category. Since 1975, EU and EEA countries have shown decreasing seropositivity; however, considerable regional variability exists. The main limitations of this study are that the studies retrieved for analysis might not be representative of all EU and EEA publications about HAV and might have poor national representativeness. A large proportion of EU and EEA residents are now susceptible to HAV infection. Our Review supports the need to reconsider specific prevention and control measures, to further decrease HAV circulation while providing protection against the infection in the EU and EEA, and could be used to inform susceptible travellers visiting EU and EEA countries with different HAV endemicity levels. Copyright © 2017 Elsevier Ltd. All rights reserved.

Estimating the scale of chronic hepatitis C virus infection in the EU/EEA: a focus on migrants from anti-HCV endemic countries.

PubMed

Falla, A M; Ahmad, A A; Duffell, E; Noori, T; Veldhuijzen, I K

2018-01-16

Increasing the proportion diagnosed with and on treatment for chronic hepatitis C (CHC) is key to the elimination of hepatitis C in Europe. This study contributes to secondary prevention planning in the European Union/European Economic Area (EU/EEA) by estimating the number of CHC (anti-HCV positive and viraemic) cases among migrants living in the EU/EEA and born in endemic countries, defining the most affected migrant populations, and assessing whether country of birth prevalence is a reliable proxy for migrant prevalence. Migrant country of birth and population size extracted from statistical databases and anti-HCV prevalence in countries of birth and in EU/EEA countries derived from a systematic literature search were used to estimate caseload among and most affected migrants. Reliability of country of birth prevalence as a proxy for migrant prevalence was assessed via a systematic literature search. Approximately 11% of the EU/EEA adult population is foreign-born, 79% of whom were born in endemic (anti-HCV prevalence ≥1%) countries. Anti-HCV/CHC prevalence in migrants from endemic countries residing in the EU/EEA is estimated at 2.3%/1.6%, corresponding to ~580,000 CHC infections or 14% of the CHC disease burden in the EU/EEA. The highest number of cases is found among migrants from Romania and Russia (50-60,000 cases each) and migrants from Italy, Morocco, Pakistan, Poland and Ukraine (25-35,000 cases each). Ten studies reporting prevalence in migrants in Europe were identified; in seven of these estimates, prevalence was comparable with the country of birth prevalence and in three estimates it was lower. Migrants are disproportionately affected by CHC, account for a considerable number of CHC infections in EU/EEA countries, and are an important population for targeted case finding and treatment. Limited data suggest that country of birth prevalence can be used as a proxy for the prevalence in migrants.

A Case Study of a Combat Helicopter’s Single Unit Vulnerability.

DTIC Science & Technology

1987-03-01

22 2.6 Generic Fault Tree Diagram ----------------------- 24 2.7 Example Kill Diagram ----------------------------- 25 2.8 Example EEA Summary...that of the vulnerability program, a susceptibility program is subdivided into three major tasks. First is an essential elements analysis ( EEA ...which leads to the 27 i final undesired event in much the same manner as a FTA. An example EEA is provided in Figure 2.8. [Ref.l:p226] The

The Causality Study of External Environment Analysis (EEA), Internal Environment Analysis (IEA), Strategy Implementation on Study Program Performance at Vocational High School (VHS) in Nias Archipelago, Indonesia

ERIC Educational Resources Information Center

Waruwu, Binahati; Sitompul, Harun; Manullang, Belferik

2016-01-01

The purposes of this study are to find out the significant effect of: (1) EEA on strategy implementation, (2) IEA on strategy implementation, (3) EEA on study program performance, (4) IEA on study program performance, and (5) strategy implementation on study program performance of Vocational High School (VHS) in Nias Archipelago. The population of…

Migration-related tuberculosis: epidemiology and characteristics of tuberculosis cases originating outside the European Union and European Economic Area, 2007 to 2013.

PubMed

Ködmön, Csaba; Zucs, Phillip; van der Werf, Marieke J

2016-01-01

Migrants arriving from high tuberculosis (TB)-incidence countries may pose a significant challenge to TB control programmes in the host country. TB surveillance data for 2007-2013 submitted to the European Surveillance System were analysed. Notified TB cases were stratified by origin and reporting country. The contribution of migrant TB cases to the TB epidemiology in EU/EEA countries was analysed. Migrant TB cases accounted for 17.4% (n = 92,039) of all TB cases reported in the EU/EEA in 2007-2013, continuously increasing from 13.6% in 2007 to 21.8% in 2013. Of 91,925 migrant cases with known country of origin, 29.3% were from the Eastern Mediterranean, 23.0% from south-east Asia, 21.4% from Africa, 13.4% from the World Health Organization European Region (excluding EU/EEA), and 12.9% from other regions. Of 46,499 migrant cases with known drug-susceptibility test results, 2.9% had multidrug-resistant TB, mainly (51.7%) originating from the European Region. The increasing contribution of TB in migrants from outside the EU/EEA to the TB burden in the EU/EEA is mainly due to a decrease in native TB cases. Especially in countries with a high proportion of TB cases in non-EU/EEA migrants, targeted prevention and control initiatives may be needed to progress towards TB elimination.

EEA stapler and omental graft in esophagogastrectomy: experience with 30 intrathoracic anastomoses for cancer.

PubMed Central

Fekete, F; Breil, P; Ronsse, H; Tossen, J C; Langonnet, F

1981-01-01

Experience with the EEA stapler device used in 30 esophagogastric resections for cancer with intrathoracic anastomosis, is reported. The mortality rate was 13.3%. The anastomotic failure rate was 3.3% (1/30) with only one death; three asymptomatic blind fistulas were found on a routine contrast examination of the anastomosis. It is felt that esophagogastric EEA stapled anastomosis associated with an omental graft is a very safe technique. Images Fig. 4. Fig. 5. Fig. 6. PMID:7247526

Algal extracellular release in river-floodplain dissolved organic matter: response of extracellular enzymatic activity during a post-flood period

PubMed Central

Sieczko, Anna; Maschek, Maria; Peduzzi, Peter

2015-01-01

River-floodplain systems are susceptible to rapid hydrological events. Changing hydrological connectivity of the floodplain generates a broad range of conditions, from lentic to lotic. This creates a mixture of allochthonously and autochthonously derived dissolved organic matter (DOM). Autochthonous DOM, including photosynthetic extracellular release (PER), is an important source supporting bacterial secondary production (BSP). Nonetheless, no details are available regarding microbial extracellular enzymatic activity (EEA) as a response to PER under variable hydrological settings in river-floodplain systems. To investigate the relationship between bacterial and phytoplankton components, we therefore used EEA as a tool to track the microbial response to non-chromophoric, but reactive and ecologically important DOM. The study was conducted in three floodplain subsystems with distinct hydrological regimes (Danube Floodplain National Park, Austria). The focus was on the post-flood period. Enhanced %PER (up to 48% of primary production) in a hydrologically isolated subsystem was strongly correlated with β-glucosidase, which was related to BSP. This shows that—in disconnected floodplain backwaters with high terrestrial input—BSP can also be driven by autochthonous carbon sources (PER). In a semi-isolated section, in the presence of fresh labile material from primary producers, enhanced activity of phenol oxidase was observed. In frequently flooded river-floodplain systems, BSP was mainly driven by enzymatic degradation of particulate primary production. Our research demonstrates that EEA measurements are an excellent tool to describe the coupling between bacteria and phytoplankton, which cannot be deciphered when focusing solely on chromophoric DOM. PMID:25741326

Priming effects and enzymatic activity in Israeli soils under treated wastewater and freshwater irrigation

NASA Astrophysics Data System (ADS)

Anissimova, Marina; Heinze, Stefanie; Chen, Yona; Tarchitzky, Jorge; Marschner, Bernd

2014-05-01

Irrigation of soils with treated wastewater (TWW) directly influences microbial processes of soil. TWW contains easily decomposable organic material, which can stimulate the activity of soil microorganisms and, as a result, lead to the excessive consumption of soil organic carbon pool. We investigated the effects of irrigation with TWW relative to those of irrigation with freshwater (FW) on the microbial parameters in soils with low (7%) and medium (13%) clay content in a lysimeter experiment. The objectives of our study were to (i) determine the impact of water quality on soil respiration and enzymatic activity influenced by clay content and depth, and (ii) work out the changes in the turnover of soil organic matter (PE, priming effects). Samples were taken from three soil depths (0-10, 10-20, and 40-60 cm). Soil respiration and PE were determined in a 21-days incubation experiment after addition of uniformly 14C-labeled fructose. Activity of 10 extracellular enzymes (EEA, from C-, N-, P-, and S-cycle), phenol oxidase and peroxidase activity (PO+PE), and dehydrogenase activity (DHA) were assayed. Microbial Community-Level Physiological Profiles (CLPP) using four substrates, and microbial biomass were determined. The results showed that the clay content acted as the main determinative factor. In the soil with low clay content the water quality had a greater impact: the highest PE (56%) was observed in the upper layer (0-10cm) under FW irrigation; EEA of C-, P-, and S-cycles was significantly higher in the upper soil layer under TWW irrigation. Microbial biomass was higher in the soil under TWW irrigation and decreased with increasing of depth (50 μg/g soil in the upper layer, 15 μg/g soil in the lowest layer). This tendency was also observed for DHA. Contrary to the low clay content, in the soil with medium clay content both irrigation types caused the highest PE in the lowest layer (65% under FW irrigation, 48% under TWW irrigation); the higher substrate mineralization (10%) and the highest phosphatase activity (in the case of FW irrigation) was observed. The PO+PE activity was two to three times higher than in the soil with low clay content and increased clearly with increasing of soil depth. The last tendency was also valid generally for the enzymes of C-, N-, and P-cycles under both types of irrigation. The upper layer in the soil under TWW irrigation was characterized by the highest microbial biomass value (74 μg/g soil). DHA in all soil depths under both types of irrigation was significantly higher than in the corresponding depths of soil with low clay content. CLPP data showed the highest consumption of ascorbic acid and D-glucosamine hydrochloride in comparison to consumption of D-glucose and L-glutamine in both irrigation types.

Reverse Transrectal Stapling Technique Using the EEA Stapler: An Alternative Approach in Difficult Reversal of Hartmann’s Procedure

PubMed Central

Zachariah, Sanoop K.

2010-01-01

The introduction of circular end-to-end stapling devices (CEEA OR EEA stapler) into colorectal surgery have revolutionised anastomotic techniques. The EEA stapler is generally regarded as an instrument that is safe, reliable, and simple to operate. Despite it’s popularity, very little information is available regarding the technical difficulties encountered during surgery. The routine technique to perform an end-to-end circular colonic anastomosis is to introduce the instrument distally through the anus (transrectal/transanal approach) and attach it to the anvil which is purse stringed at the distal end of the proximal bowel to be anastomosed. Two cases of reversal of Hartmann’s procedure for perforated diverticulitis are described in the present study, where difficulty was experienced while using the EEA stapler in the routine method. Hence, an alternative reverse technique which was used is presented. PMID:22091338

European information on climate change impacts, vulnerability and adaptation

NASA Astrophysics Data System (ADS)

Jol, A.; Isoard, S.

2010-09-01

Vulnerability to natural and technological disasters is increasing due to a combination of intensifying land use, increasing industrial development, further urban expansion and expanding infrastructure and also climate change. At EU level the European Commission's White Paper on adaptation to climate change (published in 2009) highlights that adaptation actions should be focused on the most vulnerable areas and communities in Europe (e.g. mountains, coastal areas, river flood prone areas, Mediterranean, Arctic). Mainstreaming of climate change into existing EU policies will be a key policy, including within the Water Framework Directive, Marine Strategy Framework Directive, Nature protection and biodiversity policies, integrated coastal zone management, other (sectoral) policies (agriculture, forestry, energy, transport, health) and disaster risk prevention. 2010 is the international year on biodiversity and the Conference of Parties of the biodiversity convention will meet in autumn 2010 (Japan) to discuss amongst other post-2010 strategies, objectives and indicators. Both within the Biodiversity Convention (CBD) and the Climate Change Convention (UNFCCC) there is increasing recognition of the need for integration of biodiversity conservation into climate change mitigation and adaptation activities. Furthermore a number of European countries and also some regions have started to prepare and/or have adopted national adaptation plans or frameworks. Sharing of good practices on climate change vulnerability methods and adaptation actions is so far limited, but is essential to improve such plans, at national, sub national and local level where much of the adaptation action is already taking place and will be expanding in future, also involving increasingly the business community. The EU Clearinghouse on CC impacts, vulnerability and adaptation should address these needs and it is planned to be operational end of 2011. The EEA is expected to have a role in its development in 2010 and is likely to manage the system after 2011. The European Commission in its Communication in 2009 on disaster risk prevention also calls for improving and better sharing of data on disasters, disaster risk mapping and disaster risk management, in the context of the EU civil protection mechanism. Such information might also be linked to the planned EU Clearinghouse on climate change adaptation. The activities of EEA on climate change impacts, vulnerability and adaptation (including disaster risk reduction) include indicators of the impacts of climate change; a regularly updated overview of national assessments and adaptation plans on the EEA web site and specific focused reports, e.g. on adaptation to the challenges of changing water resources in the Alps (2009) and on analysis of past trends in natural disasters (due in 2010) and regular expert meetings and workshops with EEA member countries. The ECAC presentation will include the latest developments in the EU Clearinghouse on adaptation and progress in relevant EEA activities.

Tuberculosis among migrant populations in the European Union and the European Economic Area.

PubMed

Odone, Anna; Tillmann, Taavi; Sandgren, Andreas; Williams, Gemma; Rechel, Bernd; Ingleby, David; Noori, Teymur; Mladovsky, Philipa; McKee, Martin

2015-06-01

Although tuberculosis (TB) incidence has been decreasing in the European Union/European Economic Area (EU/EEA) in the last decades, specific subgroups of the population, such as migrants, remain at high risk of TB. This study is based on the report 'Key Infectious Diseases in Migrant Populations in the EU/EEA' commissioned by The European Centre for Disease Prevention and Control. We collected, critically appraised and summarized the available evidence on the TB burden in migrants in the EU/EEA. Data were collected through: (i) a comprehensive literature review; (ii) analysis of data from The European Surveillance System (TESSy) and (iii) evidence provided by TB experts during an infectious disease workshop in 2012. In 2010, of the 73,996 TB cases notified in the EU/EEA, 25% were of foreign origin. The overall decrease of TB cases observed in recent years has not been reflected in migrant populations. Foreign-born people with TB exhibit different socioeconomic and clinical characteristics than native sufferers. This is one of the first studies to use multiple data sources, including the largest available European database on infectious disease notifications, to assess the burden and provide a comprehensive description and analysis of specific TB features in migrants in the EU/EEA. Strengthened information about health determinants and factors for migrants' vulnerability is needed to plan, implement and evaluate targeted TB care and control interventions for migrants in the EU/EEA. © The Author 2014. Published by Oxford University Press on behalf of the European Public Health Association.

[Auto-suture stapler EEA in surgery of the colon and rectum [author's transl)].

PubMed

Thiede, A; Troidl, H; Poser, H; Jostarndt, L; Hamelmann, H

1980-01-01

The increasing use of auto-suture staplers for gastrointestinal anastomoses makes it necessary to test the value of this new method of suturing. In a "pilot study" the practicability, the tightness and permeability of the anastomosis and complications were tested and analysed in a total of 30 colon and rectal anastomoses using the EEA-suture gun. The results of 14 colon resections and 16 low anterior resections justify the further use of the EEA auto-suture apparatus and raise the question of a prospective controlled random study in which manual and mechanical machine sutured anastomoses are compared.

Climate effect on soil enzyme activities and dissolved organic carbon in mountain calcareous soils: a soil-transplant experiment

NASA Astrophysics Data System (ADS)

Puissant, Jérémy; Cécillon, Lauric; Mills, Robert T. E.; Gavazov, Konstantin; Robroek, Bjorn J. M.; Spiegelberger, Thomas; Buttler, Alexandre; Brun, Jean-Jacques

2013-04-01

Mountain soils store huge amounts of carbon as soil organic matter (SOM) which may be highly vulnerable to the strong climate changes that mountain areas currently experience worldwide. Climate modifications are expected to impact microbial activity which could change the rate of SOM decomposition/accumulation, thereby questioning the net C source/sink character of mountain soils. To simulate future climate change expected in the 21st century in the calcareous pre-Alps, 15 blocks (30 cm deep) of undisturbed soil were taken from a mountain pasture located at 1400 m a.s.l. (Marchairuz, Jura, Switzerland) and transplanted into lysimeters at the same site (control) and at two other sites located at 1000 m a.s.l. and 600 m a.s.l. (5 replicates per site). This transplantation experiment which started in 2009 simulates a climate warming with a temperature increase of 4° C and a decreased humidity of 40 % at the lowest site. In this study, we used soil extracellular enzyme activities (EEA) as functional indicators of SOM decomposition to evaluate the effect of climate change on microbial activity and SOM dynamics along the seasons. Dissolved organic carbon (DOC) was also measured to quantify the assimilable carbon for microorganism. In autumn 2012, a first sampling step out of four (winter, spring and summer 2013) has been realized. We extracted 15 cm deep soil cores from each transplant (x15) and measured (i) DOC and (ii) the activities of nine different enzymes. Enzymes were chosen to represent the degradation of the most common classes of biogeochemical compounds in SOM. β-glucosidase, β-D-cellubiosidase, β-Xylosidase, N-acetyl-β-glucosaminidase, leucine aminopeptidase, lipase, phenoloxidase respectively represented the degradation of sugar, cellulose, hemicellulose, chitin, protein, lipid and lignin. Moreover, the fluorescein diacetate (FDA) hydrolysis was used to provide an estimate of global microbial activity and phosphatase was used to estimate phosphorus mineralization. The autumn results showed no differences for global microbial activity along the climate gradient (0.37 nKatal g-1 dry soil), no differences and a very low activity for leucine aminopeptidase and β-glucosidase and β-Xylosidase (about 0.09 nKatal g-1 dry soil) and no differences for cellulose, chitin and phosphorus mineralization. Conversely, we measured a greater activity at the highest elevation site for lipase and phenoloxydase (ANOVA test, p

Stabilization of red blood cells by the plasticizer, diethylhexylphthalate.

PubMed

Horowitz, B; Stryker, M H; Waldman, A A; Woods, K R; Gass, J D; Drago, J

1985-01-01

The red blood cells of blood stored in containers made of polyvinylchloride (PVC) film are osmotically more stable and lose on average about 1/3 less hemoglobin than when blood is stored in another plastic [poly-(ethylene-co-ethyl acrylate); EEA]. The stability of uniform volumes of stored red blood cells varies directly with PVC surface area, whereas changes in EEA surface area have comparatively little or no effect. PVC contains high concentrations of the plasticizer, diethylhexylphthalate (DEHP), known to migrate into blood and to have a high potential for toxicity. To determine if DEHP could be the red cell stabilizing agent in PVC, whole blood was stored in containers made from EEA into which was incorporated varying amounts of DEHP. Incorporation of DEHP into EEA significantly reduced erythrocyte osmotic fragility (p = 0.01). The degree of reduced fragility correlated with the level of DEHP in the cell phase implicating DEHP in PVC containers as the stabilizing agent for red cells.

Towards a European Framework to Monitor Infectious Diseases among Migrant Populations: Design and Applicability

PubMed Central

Riccardo, Flavia; Dente, Maria Grazia; Kärki, Tommi; Fabiani, Massimo; Napoli, Christian; Chiarenza, Antonio; Giorgi Rossi, Paolo; Velasco Munoz, Cesar; Noori, Teymur; Declich, Silvia

2015-01-01

There are limitations in our capacity to interpret point estimates and trends of infectious diseases occurring among diverse migrant populations living in the European Union/European Economic Area (EU/EEA). The aim of this study was to design a data collection framework that could capture information on factors associated with increased risk to infectious diseases in migrant populations in the EU/EEA. The authors defined factors associated with increased risk according to a multi-dimensional framework and performed a systematic literature review in order to identify whether those factors well reflected the reported risk factors for infectious disease in these populations. Following this, the feasibility of applying this framework to relevant available EU/EEA data sources was assessed. The proposed multidimensional framework is well suited to capture the complexity and concurrence of these risk factors and in principle applicable in the EU/EEA. The authors conclude that adopting a multi-dimensional framework to monitor infectious diseases could favor the disaggregated collection and analysis of migrant health data. PMID:26393623

Towards a European Framework to Monitor Infectious Diseases among Migrant Populations: Design and Applicability.

PubMed

Riccardo, Flavia; Dente, Maria Grazia; Kärki, Tommi; Fabiani, Massimo; Napoli, Christian; Chiarenza, Antonio; Giorgi Rossi, Paolo; Munoz, Cesar Velasco; Noori, Teymur; Declich, Silvia

2015-09-17

There are limitations in our capacity to interpret point estimates and trends of infectious diseases occurring among diverse migrant populations living in the European Union/European Economic Area (EU/EEA). The aim of this study was to design a data collection framework that could capture information on factors associated with increased risk to infectious diseases in migrant populations in the EU/EEA. The authors defined factors associated with increased risk according to a multi-dimensional framework and performed a systematic literature review in order to identify whether those factors well reflected the reported risk factors for infectious disease in these populations. Following this, the feasibility of applying this framework to relevant available EU/EEA data sources was assessed. The proposed multidimensional framework is well suited to capture the complexity and concurrence of these risk factors and in principle applicable in the EU/EEA. The authors conclude that adopting a multi-dimensional framework to monitor infectious diseases could favor the disaggregated collection and analysis of migrant health data.

Transbasal versus endoscopic endonasal versus combined approaches for olfactory groove meningiomas: importance of approach selection.

PubMed

Liu, James K; Silva, Nicole A; Sevak, Ilesha A; Eloy, Jean Anderson

2018-04-01

OBJECTIVE There has been much debate regarding the optimal surgical approach for resecting olfactory groove meningiomas (OGMs). In this paper, the authors analyzed the factors involved in approach selection and reviewed the surgical outcomes in a series of OGMs. METHODS A retrospective review of 28 consecutive OGMs from a prospective database was conducted. Each tumor was treated via one of 3 approaches: transbasal approach (n = 15), pure endoscopic endonasal approach (EEA; n = 5), and combined (endoscope-assisted) transbasal-EEA (n = 8). RESULTS The mean tumor volume was greatest in the transbasal (92.02 cm 3 ) and combined (101.15 cm 3 ) groups. Both groups had significant lateral dural extension over the orbits (transbasal 73.3%, p < 0.001; combined 100%), while the transbasal group had the most cerebral edema (73.3%, p < 0.001) and vascular involvement (66.7%, p < 0.001), and the least presence of a cortical cuff (33.3%, p = 0.019). All tumors in the combined group were recurrent tumors that invaded into the sinonasal cavity. The purely EEA group had the smallest mean tumor volume (33.33 cm 3 ), all with a cortical cuff and no lateral dural extension. Gross-total resection was achieved in 80% of transbasal, 100% of EEA, and 62.5% of combined cases. Near-total resection (> 95%) was achieved in 20% of transbasal and 37.5% of combined cases, all due to tumor adherence to the critical neurovascular structures. The rate of CSF leakage was 0% in the transbasal and combined groups, and there was 1 leak in the EEA group (20%), resulting in an overall CSF leakage rate of 3.6%. Olfaction was preserved in 66.7% in the transbasal group. There was no significant difference in length of stay or 30-day readmission rate between the 3 groups. The mean modified Rankin Scale score was 0.79 after the transbasal approach, 2.0 after EEA, and 2.4 after the combined approach (p = 0.0604). The mean follow-up was 14.5 months (range 1-76 months). CONCLUSIONS The transbasal approach provided the best clinical outcomes with the lowest rate of complications for large tumors (> 40 mm) and for smaller tumors (< 40 mm) with intact olfaction. The role of EEA appears to be limited to smaller, appropriately selected tumors in which olfaction is already absent. EEA also plays an important adjunctive role when combined with the transbasal approach for recurrent OGMs invading the sinonasal cavity. Careful patient selection using an individualized, tailored strategy is important to optimize surgical outcomes.

Food intake and the risk of endometrial endometrioid adenocarcinoma in Japanese women.

PubMed

Takayama, Shin; Monma, Yasutake; Tsubota-Utsugi, Megumi; Nagase, Satoru; Tsubono, Yoshitaka; Numata, Takehiro; Toyoshima, Masafumi; Utsunomiya, Hiroki; Sugawara, Junichi; Yaegashi, Nobuo

2013-01-01

The present study examined the association between food intake and endometrial cancer restricted to endometrial endometrioid adenocarcinoma (EEA) using a case-control study in Japanese women. One hundred sixty-one cases and 380 controls who completed a questionnaire regarding demographic, lifestyle, and food frequency questionnaire were analyzed. Odds ratio (OR) between selected food intakes and EEA were calculated by logistic regression analysis. After adjustment putative confounding factors, the higher intakes of vegetables [odds ratio (OR) = 0.47, 95% confidence interval (CI) = 0.26-0.83], peanuts (OR = 0.48, CI = 0.27-0.86), fish (OR = 0.52, CI = 0.29-0.93), boiled egg (OR = 0.24, CI = 0.33-0.92), instant noodles (OR = 1.94, CI = 1.12-3.34), instant food items (OR = 2.21, CI = 1.31-3.74), and deep-fried foods (OR = 2.87, CI = 1.58-5.21) were associated with a risk for EEA. The inverse association with a risk of EEA was also seen in higher intakes (g/1000 kcal) for vegetables (0.45, CI = 0.25-0.81) and fish (0.53, CI = 0.30-0.94) as compare to lower intake. Higher intake of vegetables, peanuts, fish, and boiled egg was associated with a reduced risk for EEA, whereas instant noodles, instant food items, and deep-fried foods was associated with an increased risk for EAA as compared to lower levels of intake.

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

Peteanu, Linda A.; Chowdhury, Sanchari; Wildeman, Jurjen

One measure of exciton mobility in an aggregate is the efficiency of exciton–exciton annihilation (EEA). Both exciton mobilities and EEA are enhanced for aggregate morphologies in which the distances between chromophores and their relative orientations are favorable for Förster energy transfer. Here this principle is applied to gauge the strength of interchain interactions in aggregates of two substituted PPV oligomers of 7 (OPPV7) and 13 (OPPV13) phenylene rings. These are models of the semiconducting conjugated polymer MEH–PPV. The aggregates were formed by adding a poor solvent (methanol or water) to the oligomers dissolved in a good solvent. Aggregates formed frommore » the longer-chain oligomer and/or by addition of the more polar solvent showed the largest contribution of EEA in their emission decay dynamics. This was found to correlate with the degree to which the steady-state emission spectrum of the monomer is altered by aggregation. Furthermore, the wavelength dependence of the EEA signal was also shown to be useful in differentiating emission features due to monomeric and aggregated chains when their spectra overlap significantly.« less

Exciton–exciton annihilation as a probe of interchain interactions in PPV–oligomer aggregates

DOE PAGES

Peteanu, Linda A.; Chowdhury, Sanchari; Wildeman, Jurjen; ...

2017-01-20

One measure of exciton mobility in an aggregate is the efficiency of exciton–exciton annihilation (EEA). Both exciton mobilities and EEA are enhanced for aggregate morphologies in which the distances between chromophores and their relative orientations are favorable for Förster energy transfer. Here this principle is applied to gauge the strength of interchain interactions in aggregates of two substituted PPV oligomers of 7 (OPPV7) and 13 (OPPV13) phenylene rings. These are models of the semiconducting conjugated polymer MEH–PPV. The aggregates were formed by adding a poor solvent (methanol or water) to the oligomers dissolved in a good solvent. Aggregates formed frommore » the longer-chain oligomer and/or by addition of the more polar solvent showed the largest contribution of EEA in their emission decay dynamics. This was found to correlate with the degree to which the steady-state emission spectrum of the monomer is altered by aggregation. Furthermore, the wavelength dependence of the EEA signal was also shown to be useful in differentiating emission features due to monomeric and aggregated chains when their spectra overlap significantly.« less

Exciton-Exciton Annihilation as a Probe of Interchain Interactions in PPV-Oligomer Aggregates.

PubMed

Peteanu, Linda A; Chowdhury, Sanchari; Wildeman, Jurjen; Sfeir, Matthew Y

2017-02-23

One measure of exciton mobility in an aggregate is the efficiency of exciton-exciton annihilation (EEA). Both exciton mobilities and EEA are enhanced for aggregate morphologies in which the distances between chromophores and their relative orientations are favorable for Förster energy transfer. Here this principle is applied to gauge the strength of interchain interactions in aggregates of two substituted PPV oligomers of 7 (OPPV7) and 13 (OPPV13) phenylene rings. These are models of the semiconducting conjugated polymer MEH-PPV. The aggregates were formed by adding a poor solvent (methanol or water) to the oligomers dissolved in a good solvent. Aggregates formed from the longer-chain oligomer and/or by addition of the more polar solvent showed the largest contribution of EEA in their emission decay dynamics. This was found to correlate with the degree to which the steady-state emission spectrum of the monomer is altered by aggregation. The wavelength dependence of the EEA signal was also shown to be useful in differentiating emission features due to monomeric and aggregated chains when their spectra overlap significantly.

Hepatitis E virus infection in Europe: surveillance and descriptive epidemiology of confirmed cases, 2005 to 2015.

PubMed

Aspinall, Esther J; Couturier, Elisabeth; Faber, Mirko; Said, Bengü; Ijaz, Samreen; Tavoschi, Lara; Takkinen, Johanna; Adlhoch, Cornelia

2017-06-29

Hepatitis E virus (HEV) is an under-recognised cause of acute hepatitis in high-income countries. The purpose of this study was to provide an overview of testing, diagnosis, surveillance activities, and data on confirmed cases in the European Union/European Economic Area (EU/EEA). A semi-structured survey was developed and sent to 31 EU/EEA countries in February 2016, 30 responded. Twenty of these countries reported that they have specific surveillance systems for HEV infection. Applied specific case definition for HEV infection varied widely across countries. The number of reported cases has increased from 514 cases per year in 2005 to 5,617 in 2015, with most infections being locally acquired. This increase could not be explained by additional countries implementing surveillance for HEV infections over time. Hospitalisations increased from less than 100 in 2005 to more than 1,100 in 2015 and 28 fatal cases were reported over the study period. EU/EEA countries are at different stages in their surveillance, testing schemes and policy response to the emergence of HEV infection in humans. The available data demonstrated a Europe-wide increase in cases. Standardised case definitions and testing policies would allow a better understanding of the epidemiology of HEV as an emerging cause of liver-related morbidity. This article is copyright of The Authors, 2017.

The effectiveness and cost-effectiveness of screening for latent tuberculosis among migrants in the EU/EEA: a systematic review

PubMed Central

Greenaway, Christina; Pareek, Manish; Abou Chakra, Claire-Nour; Walji, Moneeza; Makarenko, Iuliia; Alabdulkarim, Balqis; Hogan, Catherine; McConnell, Ted; Scarfo, Brittany; Christensen, Robin; Tran, Anh; Rowbotham, Nick; van der Werf, Marieke J; Noori, Teymur; Pottie, Kevin; Matteelli, Alberto; Zenner, Dominik; Morton, Rachael L.

2018-01-01

Background Migrants account for a large and growing proportion of tuberculosis (TB) cases in low-incidence countries in the European Union/European Economic Area (EU/EEA) which are primarily due to reactivation of latent TB infection (LTBI). Addressing LTBI among migrants will be critical to achieve TB elimination. Methods: We conducted a systematic review to determine effectiveness (performance of diagnostic tests, efficacy of treatment, uptake and completion of screening and treatment) and a second systematic review on cost-effectiveness of LTBI screening programmes for migrants living in the EU/EEA. Results: We identified seven systematic reviews and 16 individual studies that addressed our aims. Tuberculin skin tests and interferon gamma release assays had high sensitivity (79%) but when positive, both tests poorly predicted the development of active TB (incidence rate ratio: 2.07 and 2.40, respectively). Different LTBI treatment regimens had low to moderate efficacy but were equivalent in preventing active TB. Rifampicin-based regimens may be preferred because of lower hepatotoxicity (risk ratio = 0.15) and higher completion rates (82% vs 69%) compared with isoniazid. Only 14.3% of migrants eligible for screening completed treatment because of losses along all steps of the LTBI care cascade. Limited economic analyses suggest that the most cost-effective approach may be targeting young migrants from high TB incidence countries. Discussion: The effectiveness of LTBI programmes is limited by the large pool of migrants with LTBI, poorly predictive tests, long treatments and a weak care cascade. Targeted LTBI programmes that ensure high screening uptake and treatment completion will have greatest individual and public health benefit. PMID:29637889

What do employers think their role is in ensuring language proficiency of internationally trained pharmacists.

PubMed

Ziaei, Zainab; Hassell, Karen; Schafheutle, Ellen I

2018-04-01

Until 2016, internationally trained pharmacists (ITPs) from the European Economic Area (EEA) did not need to prove sufficient language proficiency to the pharmacy regulator upon registration. Pharmacists themselves have a professional responsibility to ensure they can communicate and work effectively, but some responsibility also rests with employers, yet very little research has explored this. The aim of this study was to explore employer views of the communicative proficiency of ITPs whose first language was not English, their role as employers and potential implications for patient safety. Semi-structured, telephone interviews were conducted with seven community and two hospital employers with experience of employing EEA pharmacists, between May and July 2010. Verbatim interview transcripts were coded and analysed in NVivo using the framework approach. All participants mentioned the importance of having processes in place to assure EEA pharmacists' linguistic competency in the workplace. During recruitment, different strategies were used to assure language competency. Some employers only conducted interviews while others required candidates to pass English language assessments. Participants were most familiar with the International English Language Testing System, which was described as too general and not unique to pharmacy. Other pharmacy-specific tests such as the University of Bath English Language Test and Linguarama English Assessment Test for Pharmacists were alternatives. Currently, there is no one standard procedure in place to check the communicative competency of EEA pharmacists. The findings from this study suggest that there is need to establish a uniform assessment system so all the EEA pharmacists could be tested justly and consistently. © 2017 Royal Pharmaceutical Society.

Safety and short-term effectiveness of EEA stapler vs PPH stapler in the treatment of degree III haemorrhoids: prospective randomized controlled trial.

PubMed

Giuratrabocchetta, S; Pecorella, G; Stazi, A; Tegon, G; De Fazio, M; Altomare, D F

2013-03-01

Stapled haemorrhoidopexy has gained wide acceptance due to less postoperative pain although postoperative bleeding and prolapse recurrence are among the major drawbacks of this technique compared with the standard Milligan-Morgan hemorrhoidectomy. The aim was to investigate a new stapler device designed to overcome these side effects. In all, 135 patients (71 men, mean age 42 years) with degree III haemorrhoids were randomly allotted to stapled haemorrhoidopexy with PPH® staplers (Ethicon EndoSurgery) (63 patients) or with an EEA® stapler (Covidien) (72 patients) in four referral colorectal centres. The number of haemostatic overstitches apposed on the stapled suture, the area of the resected mucosa (in square centimetres) and any postoperative bleeding within 30 days were recorded. The mean area of the resected mucosa was significantly wider in EEA than PPH patients (35.75 ± 17.51 vs 28.05 ± 10.23 cm(2), P = 0.002). The median number of haemostatic stitches apposed in the EEA group was significantly lower than in the PPH groups (median value 1, vs 3, interquartile range 0-2, vs 2-5, P < 0.0001). Intraoperative haemostasis was better in the EEA group compared with the PPH01 and PPH03 groups. Postoperative bleeding occurred only in two PPH patients. Data suggest that the EEA stapler has better haemostatic properties than the PPH stapler and allows resection of a larger area of mucosal prolapse with potential benefits over the recurrence rate of haemorrhoid prolapse. © 2012 The Authors. Colorectal Disease © 2012 The Association of Coloproctology of Great Britain and Ireland.

Early harvesting of the vascularized pedicled nasoseptal flap during endoscopic skull base surgery.

PubMed

Eloy, Jean Anderson; Patel, Amit A; Shukla, Pratik A; Choudhry, Osamah J; Liu, James K

2013-01-01

The vascularized pedicled nasoseptal flap (PNSF) represents a successful option for reconstruction of large skull base defects after expanded endoscopic endonasal approaches (EEA). This vascularized flap can be harvested early or late in the operation depending on the anticipation of high-flow CSF leaks. Each harvesting technique (early vs. late) is associated with different advantages and disadvantages. In this study, we evaluate our experience with early harvesting of the PNSF for repair of large skull base defects after EEA. A retrospective review was performed at a tertiary care medical center on patients who underwent early PNSF harvesting during reconstruction of intraoperative high-flow CSF leaks after EEA between December 2008 and March 2012. Demographic data, repair materials, surgical approach, and incidence of PNSF usage were collected. Eighty-seven patients meeting the inclusion criteria were identified. In 86 procedures (98.9%), the PNSF harvested at the beginning of the operation was used. In 1 case (1.1%), the PNSF was not used because a high-flow intraoperative CSF leak was not encountered. This patient had recurrence of intradural disease 8months later, and the previously elevated PNSF was subsequent used after tumor resection. Based on our data, a high-flow CSF leak and need for a PNSF can be accurately anticipated in patients undergoing EEA for skull base lesions. Because of the advantages of early harvesting of the PNSF and the high preoperative predictive value of CSF leak anticipations, this technique represents a feasible harvesting practice for EEA surgeries. Copyright © 2013 Elsevier Inc. All rights reserved.

Microbial decomposition is highly sensitive to leaf litter emersion in a permanent temperate stream.

PubMed

Mora-Gómez, Juanita; Duarte, Sofia; Cássio, Fernanda; Pascoal, Cláudia; Romaní, Anna M

2018-04-15

Drought frequency and intensity in some temperate regions are forecasted to increase under the ongoing global change, which might expose permanent streams to intermittence and have severe repercussions on stream communities and ecosystem processes. In this study, we investigated the effect of drought duration on microbial decomposition of Populus nigra leaf litter in a temperate permanent stream (Oliveira, NW Portugal). Specifically, we measured the response of the structural (assemblage composition, bacterial and fungal biomass) and functional (leaf litter decomposition, extracellular enzyme activities (EEA), and fungal sporulation) parameters of fungal and bacterial communities on leaf litter exposed to emersion during different time periods (7, 14 and 21d). Emersion time affected microbial assemblages and litter decomposition, but the response differed among variables. Leaf decomposition rates and the activity of β-glucosidase, cellobiohydrolase and phosphatase were gradually reduced with increasing emersion time, while β-xylosidase reduction was similar when emersion last for 7 or more days, and the phenol oxidase reduction was similar at 14 and 21days of leaf emersion. Microbial biomass and fungal sporulation were reduced after 21days of emersion. The structure of microbial assemblages was affected by the duration of the emersion period. The shifts in fungal assemblages were correlated with a decreased microbial capacity to degrade lignin and hemicellulose in leaf litter exposed to emersion. Additionally, some resilience was observed in leaf litter mass loss, bacterial biomass, some enzyme activities and structure of fungal assemblages. Our study shows that drought can strongly alter structural and functional aspects of microbial decomposers. Therefore, the exposure of leaf litter to increasing emersion periods in temperate streams is expected to affect decomposer communities and overall decomposition of plant material by decelerating carbon cycling in streams. Copyright © 2017 Elsevier B.V. All rights reserved.

Suprachiasmatic translamina terminalis corridor used in endoscopic endonasal approach for resecting third ventricular craniopharyngioma.

PubMed

Gu, Ye; Zhang, Xiaobiao; Hu, Fan; Yu, Yong; Xie, Tao; Sun, Chongjing; Li, Wensheng

2015-05-01

The translamina terminalis corridor was used in the transcranial anterior route to treat third ventricular craniopharyngioma (TVC), which presents a challenge to neurosurgeons. The endoscopic endonasal approach (EEA) has recently been used to treat craniopharyngiomas. However, there are few reports of the EEA being used to treat TVC. The authors' novel surgical approach of treating selected TVC by the endoscopic endonasal route via the suprachiasmatic translamina terminalis (STLT) corridor is described. In this single-center study, the EEA via the STLT corridor was used to resect TVC with great upper and anterior extension causing bulged lamina terminalis, and TVC with a residual upper compartment, after routine infrachiasmatic transmetastalk corridor resection. The STLT corridor was used in 3 patients. Gross-total resection was achieved in all cases. One patient achieved visual improvement, and the other 2 patients showed partial visual improvement. Leakage of CSF occurred in 1 patient. Postoperative hormone replacement therapy was required in all patients. The STLT corridor is a complementary minimally invasive corridor used in the EEA for treating selected TVC. The STLT alone or combined with infrachiasmatic transmetastalk corridors should be selected depending on the size of suprachiasmatic and infrachiasmatic space.

Work experiences of internationally trained pharmacists in Great Britain.

PubMed

Ziaei, Zainab; Hassell, Karen; Schafheutle, Ellen I

2015-04-01

Internationally trained health professionals are an important part of the domestic workforce, but little is known about the working experiences of internationally trained pharmacists (ITPs) in Great Britain (GB). The purpose of this study is to explore the work experiences of ITPs practising in the community or hospital sector in GB. Twenty-five semi-structured, face-to-face interviews were conducted with a sample of European Economic Area (EEA) and non-EEA pharmacists who, at the time of the study, practised in the community (n = 20) or hospital sector (n = 5) in the North West England from March to May 2009. In general, ITPs complained about their heavy workload, long working hours and lack of support from their employers. Specifically, EEA pharmacists in most cases felt excluded from the professional network and sensed colleagues saw them as 'foreigners' while some non-EEA pharmacists had to deal with a level of hostility from patients. This novel research provides a foundation for future work on ITPs in GB and could assist employers to better target their efforts in development of standards to support the working experiences of ITPs in GB. © 2014 Royal Pharmaceutical Society.

Biochemistry of whole blood in poly(ethylene-co-ethylacrylate) experimental blood containers.

PubMed

Horowitz, B; Rowe, A W; Stryker, M H; Van der Sande, J; Waldman, A A; Wong, S C; Gass, J D; Woods, K R

1985-01-01

The biochemical status of whole blood stored in containers fabricated of ethylene ethylacrylate (EEA) film was monitored at several times during 4 weeks of storage at 4 degrees C. Fifteen biochemical indicators were studied to reflect on erythrocyte integrity, cellular metabolism, plasma protein stability, and microaggregate formation. Comparison to storage in polyvinyl chloride (PVC) containers was made by distributing aliquots from each unit of blood among the containers being compared. Whole blood in EEA developed significantly higher levels of plasma hemoglobin, erythrocyte osmotic fragility, and D-glycerate-2,3-diphosphate (2,3-DPG), and somewhat greater glucose utilization, lactate production, and pH. These biochemical differences were not of great magnitude and the data suggest that EEA containers are compatible with the storage of whole blood.

A refined method for the calculation of the Non-Methane Volatile Organic Compound emission estimate from Domestic Solvent Usage in Ireland from 1992 to 2014 - A case study for Ireland

NASA Astrophysics Data System (ADS)

Barry, Stephen; O'Regan, Bernadette

2016-08-01

This study describes a new methodology to calculate Non-Methane Volatile Organic Compounds from Domestic Solvent Use including Fungicides over the period 1992-2014. Improved emissions data compiled at a much more refined level can help policy-makers develop more effective policy's to address environmental issues. However, a number of problems were found when member states attempt to use national statistics for Domestic Solvent Use including Fungicides. For instance, EMEP/EEA (2013) provides no guidance regarding which activity data should be used, resulting in emission estimates being potentially inconsistent and un-comparable. Also, previous methods and emission factors described in the EMEP/EEA (2013) guidebook do not exactly match data collected by state agencies. This makes using national statistics difficult. In addition, EMEP/EEA (2013) use broader categories than necessary (e.g. Cosmetics Aerosol/Non Aerosol) to estimate emissions while activity data is available at a more refined level scale (e.g. Personal Cleaning Products, Hair Products, Cosmetics, Deodorants and Perfumes). This can make identifying the drivers of emissions unclear. This study builds upon Tzanidakis et al. (2012) whereby it provides a method for collecting activity data from state statistics, developed country specific emission factors based on a survey of 177 Irish products and importantly, used a new method to account for the volatility of organic compounds found in commonly available domestic solvent containing products. This is the first study to account for volatility based on the characteristics of organic compounds and therefore is considered a more accurate method of accounting for emissions from this emission source. The results of this study can also be used to provide a simple method for other member parties to account for the volatility of organic compounds using sectorial adjustment factors described here. For comparison purposes, emission estimates were calculated using the Tier 1 approach currently used in the emission inventory, using activity data and emission factors unadjusted for volatility and adjusted for volatility. The unadjusted estimate is useful, because it demonstrates the failure to properly account for volatility can produce significantly over-estimated emissions from the Domestic Solvent Usage sector. Unadjusted emissions were found to be 30% lower than the EMEP/EEA (2013) Tier 1 period in 2014. Emissions were found to reduce a further 20.9% when the volatility of the organic compounds was included. This new method shows that member parties may be significantly overestimating emissions from Domestic Solvent Use including pesticides and further work should include refining organic compound content and the sectorial adjustment factor of products.

Aspects of medical migration with particular reference to the United Kingdom and the Netherlands.

PubMed

Herfs, Paul G P

2014-10-14

In most countries of the European Economic Area (EEA), there is no large-scale migration of medical graduates with diplomas obtained outside the EEA, which are international medical graduates (IMGs). In the United Kingdom however, health care is in part dependent on the influx of IMGs. In 2005, of all the doctors practising in the UK, 31% were educated outside the country. In most EEA-countries, health care is not dependent on the influx of IMGs.The aim of this study is to present data relating to the changes in IMG migration in the UK since the extension of the European Union in May 2004. In addition, data are presented on IMG migration in the Netherlands. These migration flows show that migration patterns differ strongly within these two EU-countries. This study makes use of registration data on migrating doctors from the General Medical Council (GMC) in the UK and from the Dutch Department of Health. Moreover, data on the ratio of medical doctors in relation to a country's population were extracted from the World Health Organization (WHO). The influx of IMGs in the UK has changed in recent years due to the extension of the European Union in 2004, the expansion of UK medical schools and changes in the policy towards non-EEA doctors.The influx of IMGs in the Netherlands is described in detail. In the Netherlands, many IMGs come from Afghanistan, Iraq and Surinam. There are clear differences between IMG immigration in the UK and in the Netherlands. In the UK, the National Health Service continues to be very reliant on immigration to fill shortage posts, whereas the number of immigrant doctors working in the Netherlands is much smaller. Both the UK and the Netherlands' regulatory bodies have shared great concerns about the linguistic and communication skills of both EEA and non-EEA doctors seeking to work in these countries. IMG migration is a global and intricate problem. The source countries, not only those where English is the first or second language, experience massive IMG migration flows.

Surveillance and laboratory detection for non-polio enteroviruses in the European Union/European Economic Area, 2016

PubMed Central

Harvala, Heli; Jasir, Aftab; Penttinen, Pasi; Pastore Celentano, Lucia; Greco, Donato; Broberg, Eeva

2017-01-01

Enteroviruses (EVs) cause severe outbreaks of respiratory and neurological disease as illustrated by EV-D68 and EV-A71 outbreaks, respectively. We have mapped European laboratory capacity for identification and characterisation of non-polio EVs to improve preparedness to respond to (re)-emerging EVs linked to severe disease. An online questionnaire on non-polio EV surveillance and laboratory detection was submitted to all 30 European Union (EU)/European Economic Area (EEA) countries. Twenty-nine countries responded; 26 conducted laboratory-based non-polio EV surveillance, and 24 included neurological infections in their surveillance. Eleven countries have established specific surveillance for EV-D68 via sentinel influenza surveillance (n = 7), typing EV-positive respiratory samples (n = 10) and/or acute flaccid paralysis surveillance (n = 5). Of 26 countries performing non-polio EV characterisation/typing, 10 further characterised culture-positive EV isolates, whereas the remainder typed PCR-positive but culture-negative samples. Although 19 countries have introduced sequence-based EV typing, seven still rely entirely on virus isolation. Based on 2015 data, six countries typed over 300 specimens mostly by sequencing, whereas 11 countries characterised under 50 EV-positive samples. EV surveillance activity varied between EU/EEA countries, and did not always specifically target patients with neurological and/or respiratory infections. Introduction of sequence-based typing methods is needed throughout the EU/EEA to enhance laboratory capacity for the detection of EVs. PMID:29162204

Surveillance and laboratory detection for non-polio enteroviruses in the European Union/European Economic Area, 2016.

PubMed

Harvala, Heli; Jasir, Aftab; Penttinen, Pasi; Pastore Celentano, Lucia; Greco, Donato; Broberg, Eeva

2017-11-01

Enteroviruses (EVs) cause severe outbreaks of respiratory and neurological disease as illustrated by EV-D68 and EV-A71 outbreaks, respectively. We have mapped European laboratory capacity for identification and characterisation of non-polio EVs to improve preparedness to respond to (re)-emerging EVs linked to severe disease. An online questionnaire on non-polio EV surveillance and laboratory detection was submitted to all 30 European Union (EU)/European Economic Area (EEA) countries. Twenty-nine countries responded; 26 conducted laboratory-based non-polio EV surveillance, and 24 included neurological infections in their surveillance. Eleven countries have established specific surveillance for EV-D68 via sentinel influenza surveillance (n = 7), typing EV-positive respiratory samples (n = 10) and/or acute flaccid paralysis surveillance (n = 5). Of 26 countries performing non-polio EV characterisation/typing, 10 further characterised culture-positive EV isolates, whereas the remainder typed PCR-positive but culture-negative samples. Although 19 countries have introduced sequence-based EV typing, seven still rely entirely on virus isolation. Based on 2015 data, six countries typed over 300 specimens mostly by sequencing, whereas 11 countries characterised under 50 EV-positive samples. EV surveillance activity varied between EU/EEA countries, and did not always specifically target patients with neurological and/or respiratory infections. Introduction of sequence-based typing methods is needed throughout the EU/EEA to enhance laboratory capacity for the detection of EVs.

Ecoenzymatic stoichiometry and microbial processing of organic matter in northern bogs and fens reveals a common P limitation among peatland types

EPA Science Inventory

We compared C, N, and P concentrations in atmospheric deposition, runoff, and soil standing stocks with microbial respiration (DHA) and ecoenzyme activity (EEA) in an ombrotrophic bog (S2) and a minerotrophic fen (S3) to investigate the environmental drivers of biogeochemical cyc...

Usefulness of bedside tissue Doppler echocardiography and B-type natriuretic peptide (BNP) in differentiating congestive heart failure from noncardiac cause of acute dyspnea in elderly patients with a normal left ventricular ejection fraction and permanent, nonvalvular atrial fibrillation: insights from a prospective, monocenter study.

PubMed

Arques, Stephane; Roux, Emmanuel; Sbragia, Pascal; Pieri, Bertrand; Gelisse, Richard; Luccioni, Roger; Ambrosi, Pierre

2007-05-01

The incremental role of bedside tissue Doppler echocardiography and B-type natriuretic peptide (BNP) over the clinical judgment has been recently reported in the emergency diagnosis of congestive heart failure with a normal left ventricular ejection fraction (HFNEF). However, how well does this diagnostic strategy be applicable in the setting of atrial fibrillation is unknown. To investigate the usefulness of bedside tissue Doppler echocardiography and BNP in the emergency diagnosis of HFNEF in elderly patients with permanent, nonvalvular atrial fibrillation. Forty-one consecutive elderly patients with an ejection fraction > or =50% (mean age 84 years; 22 with HFNEF and 19 with noncardiac cause), hospitalized for acute dyspnea at rest, were prospectively enrolled; bedside septal E/Ea and BNP were obtained at admission. By multivariable logistic regression analysis including the clinical judgment of heart failure, E/Ea and BNP, E/Ea (P = 0.014) and BNP (P = 0.018) provided independent diagnostic information. Optimal cutoffs were 13 for E/Ea (area under the ROC curve of 0.846, P < 0.0001; sensitivity 81.8%, specificity 89.5%) and 253 pg/ml for BNP (area under the ROC curve of 0.928, P < 0.0001; sensitivity 86.4%, specificity 89.5%). The concordance between the clinical judgment and BNP concentration at the cutoff of 253 pg/ml correctly classified 24 of 25 patients; E/Ea at the cutoff of 13 correctly classified 14 of the 16 patients with discrepancy. Bedside tissue Doppler echocardiography and BNP provide useful additional diagnostic information over the clinical judgment for the emergency diagnosis of HFNEF in elderly patients with permanent, nonvalvular atrial fibrillation.

Genital Chlamydia Prevalence in Europe and Non-European High Income Countries: Systematic Review and Meta-Analysis

PubMed Central

Redmond, Shelagh M.; Alexander-Kisslig, Karin; Woodhall, Sarah C.; van den Broek, Ingrid V. F.; van Bergen, Jan; Ward, Helen; Uusküla, Anneli; Herrmann, Björn; Andersen, Berit; Götz, Hannelore M.; Sfetcu, Otilia; Low, Nicola

2015-01-01

Background Accurate information about the prevalence of Chlamydia trachomatis is needed to assess national prevention and control measures. Methods We systematically reviewed population-based cross-sectional studies that estimated chlamydia prevalence in European Union/European Economic Area (EU/EEA) Member States and non-European high income countries from January 1990 to August 2012. We examined results in forest plots, explored heterogeneity using the I2 statistic, and conducted random effects meta-analysis if appropriate. Meta-regression was used to examine the relationship between study characteristics and chlamydia prevalence estimates. Results We included 25 population-based studies from 11 EU/EEA countries and 14 studies from five other high income countries. Four EU/EEA Member States reported on nationally representative surveys of sexually experienced adults aged 18–26 years (response rates 52–71%). In women, chlamydia point prevalence estimates ranged from 3.0–5.3%; the pooled average of these estimates was 3.6% (95% CI 2.4, 4.8, I2 0%). In men, estimates ranged from 2.4–7.3% (pooled average 3.5%; 95% CI 1.9, 5.2, I2 27%). Estimates in EU/EEA Member States were statistically consistent with those in other high income countries (I2 0% for women, 6% for men). There was statistical evidence of an association between survey response rate and estimated chlamydia prevalence; estimates were higher in surveys with lower response rates, (p = 0.003 in women, 0.018 in men). Conclusions Population-based surveys that estimate chlamydia prevalence are at risk of participation bias owing to low response rates. Estimates obtained in nationally representative samples of the general population of EU/EEA Member States are similar to estimates from other high income countries. PMID:25615574

Force recovery and axonal regeneration of the sternomastoid muscle reinnervated with the end-to-end nerve anastomosis

PubMed Central

Sobotka, Stanislaw; Mu, Liancai

2012-01-01

Background End-to-end nerve anastomosis (EEA) is a commonly used nerve repair technique. However, this method generally results in poor functional recovery. This study was designed to determine the correlation of functional recovery to the extent of axonal reinnervation after EEA procedure in a rat model. Materials and Methods Seven adult rats were subjected to the immediate reinnervation of an experimentally paralyzed sternomastoid (SM) muscle. The SM nerve was transected and immediately repaired with EEA. The SM muscle at the opposite side, without nerve transection, served as a control. Three months after EEA nerve repair, the muscle force of the SM muscle was measured and the regenerated axons in the muscle were detected using neurofilament immunohistochemistry. Results Three months after surgery, the reinnervated SM muscle produced limited anatomical and functional recovery (calculated as the percentage of the control). Specifically, the wet weight of the operated SM muscle (a measure of muscle mass recovery) was 78.0% of the control. The maximal tetanic force (a measure of muscle functional recovery) was 56.7% of the control. The area fraction of the neurofilament stained intramuscular axons (a measure of axonal regeneration and muscle reinnervation) was measured to be only 13.4% of the control. A positive correlation was revealed between the extent of muscle reinnervation and maximal muscle force. Conclusions The EEA reinnervated SM muscle in the rat yielded unsatisfactory muscle force recovery as a result of mild to moderate nerve regeneration. Further work is needed to improve the surgical procedure, enhance axonal regeneration, and/or develop novel treatment strategies for better functional recovery. PMID:23207170

Supraorbital Versus Endoscopic Endonasal Approaches for Olfactory Groove Meningiomas: A Cost-Minimization Study.

PubMed

Gandhoke, Gurpreet S; Pease, Matthew; Smith, Kenneth J; Sekula, Raymond F

2017-09-01

To perform a cost-minimization study comparing the supraorbital and endoscopic endonasal (EEA) approach with or without craniotomy for the resection of olfactory groove meningiomas (OGMs). We built a decision tree using probabilities of gross total resection (GTR) and cerebrospinal fluid (CSF) leak rates with the supraorbital approach versus EEA with and without additional craniotomy. The cost (not charge or reimbursement) at each "stem" of this decision tree for both surgical options was obtained from our hospital's finance department. After a base case calculation, we applied plausible ranges to all parameters and carried out multiple 1-way sensitivity analyses. Probabilistic sensitivity analyses confirmed our results. The probabilities of GTR (0.8) and CSF leak (0.2) for the supraorbital craniotomy were obtained from our series of 5 patients who underwent a supraorbital approach for the resection of an OGM. The mean tumor volume was 54.6 cm 3 (range, 17-94.2 cm 3 ). Literature-reported rates of GTR (0.6) and CSF leak (0.3) with EEA were applied to our economic analysis. Supraorbital craniotomy was the preferred strategy, with an expected value of $29,423, compared with an EEA cost of $83,838. On multiple 1-way sensitivity analyses, supraorbital craniotomy remained the preferred strategy, with a minimum cost savings of $46,000 and a maximum savings of $64,000. Probabilistic sensitivity analysis found the lowest cost difference between the 2 surgical options to be $37,431. Compared with EEA, supraorbital craniotomy provides substantial cost savings in the treatment of OGMs. Given the potential differences in effectiveness between approaches, a cost-effectiveness analysis should be undertaken. Copyright © 2017 Elsevier Inc. All rights reserved.

Force recovery and axonal regeneration of the sternomastoid muscle reinnervated with the end-to-end nerve anastomosis.

PubMed

Sobotka, Stanislaw; Mu, Liancai

2013-06-15

End-to-end nerve anastomosis (EEA) is a commonly used nerve repair technique. However, this method generally results in poor functional recovery. This study was designed to determine the correlation of functional recovery to the extent of axonal reinnervation after EEA procedure in a rat model. Seven adult rats were subjected to the immediate reinnervation of an experimentally paralyzed sternomastoid (SM) muscle. The SM nerve was transected and immediately repaired with EEA. The SM muscle at the opposite side, without nerve transection, served as a control. Three months after EEA nerve repair, the muscle force of the SM muscle was measured and the regenerated axons in the muscle were detected using neurofilament immunohistochemistry. Three months after surgery, the reinnervated SM muscle produced limited anatomical and functional recovery (calculated as the percentage of the control). Specifically, the wet weight of the operated SM muscle (a measure of muscle mass recovery) was 78.0% of the control. The maximal tetanic force (a measure of muscle functional recovery) was 56.7% of the control. The area fraction of the neurofilament stained intramuscular axons (a measure of axonal regeneration and muscle reinnervation) was measured to be only 13.4% of the control. A positive correlation was revealed between the extent of muscle reinnervation and maximal muscle force. The EEA reinnervated SM muscle in the rat yielded unsatisfactory muscle force recovery as a result of mild to moderate nerve regeneration. Further work is needed to improve the surgical procedure, enhance axonal regeneration, and/or develop novel treatment strategies for better functional recovery. Copyright © 2013 Elsevier Inc. All rights reserved.

Organic matter quantity and source affects microbial community structure and function following volcanic eruption on Kasatochi Island, Alaska

USGS Publications Warehouse

Zeglin, Lydia H.; Wang, Bronwen; Waythomas, Christopher F.; Rainey, Frederick; Talbot, Sandra L.

2016-01-01

In August 2008, Kasatochi volcano erupted and buried a small island in pyroclastic deposits and fine ash; since then, microbes, plants and birds have begun to re-colonize the initially sterile surface. Five years post-eruption, bacterial 16S rRNA gene and fungal internal transcribed spacer (ITS) copy numbers and extracellular enzyme activity (EEA) potentials were one to two orders of magnitude greater in pyroclastic materials with organic matter (OM) inputs relative to those without, despite minimal accumulation of OM (< 0.2%C). When normalized by OM levels, post-eruptive surfaces with OM inputs had the highest β-glucosidase, phosphatase, NAGase and cellobiohydrolase activities, and had microbial population sizes approaching those in reference soils. In contrast, the strongest factor determining bacterial community composition was the dominance of plants versus birds as OM input vectors. Although soil pH ranged from 3.9 to 7.0, and %C ranged 100×, differentiation between plant- and bird-associated microbial communities suggested that cell dispersal or nutrient availability are more likely drivers of assembly than pH or OM content. This study exemplifies the complex relationship between microbial cell dispersal, soil geochemistry, and microbial structure and function; and illustrates the potential for soil microbiota to be resilient to disturbance.

Ecoenzymatic stoichiometry and microbial processing of organic matter in northern bogs and fens reveals a common P-limitation between peatland types

Treesearch

Brian H. Hill; Colleen M. Elonen; Terri M. Jicha; Randall K. Kolka; LaRae L.P. Lehto; Stephen D. Sebestyen; Lindsey R. Seifert-Monson

2014-01-01

We compared carbon (C), nitrogen (N), and phosphorus (P) concentrations in atmospheric deposition, runoff, and soils with microbial respiration [dehydrogenase (DHA)] and ecoenzyme activity (EEA) in an ombrotrophic bog and a minerotrophic fen to investigate the environmental drivers of biogeochemical cycling in peatlands at the Marcell Experimental Forest in northern...

Surveillance of Zika virus infection in the EU/EEA, June 2015 to January 2017.

PubMed

Spiteri, G; Sudre, B; Septfons, A; Beauté, J

2017-10-01

Surveillance of Zika virus (ZIKV) infection in the European Union/European Economic Area (EU/EEA) was implemented in 2016 in response to the large outbreak reported in the Americas in 2015 associated with an increased number of infants born with microcephaly. Between June 2015 and January 2017, 21 EU/EEA countries reported 2,133 confirmed cases of ZIKV infection, of whom 106 were pregnant women. Cases infected in the Caribbean constituted 71% of reported cases. Almost all cases (99%) were most probably infected by mosquito bite during travel outside continental Europe, while only 1% were transmitted sexually. Considering that 584 imported cases were reported between May and October 2016 among residents of areas with established presence of Aedes albopictus , the absence of autochthonous vector-borne cases suggests that Ae. albopictus is not an efficient vector for ZIKV infection.

Refining Trait Resilience: Identifying Engineering, Ecological, and Adaptive Facets from Extant Measures of Resilience

PubMed Central

Maltby, John; Day, Liz; Hall, Sophie

2015-01-01

The current paper presents a new measure of trait resilience derived from three common mechanisms identified in ecological theory: Engineering, Ecological and Adaptive (EEA) resilience. Exploratory and confirmatory factor analyses of five existing resilience scales suggest that the three trait resilience facets emerge, and can be reduced to a 12-item scale. The conceptualization and value of EEA resilience within the wider trait and well-being psychology is illustrated in terms of differing relationships with adaptive expressions of the traits of the five-factor personality model and the contribution to well-being after controlling for personality and coping, or over time. The current findings suggest that EEA resilience is a useful and parsimonious model and measure of trait resilience that can readily be placed within wider trait psychology and that is found to contribute to individual well-being. PMID:26132197

Is the chiasm-pituitary corridor size important for achieving gross-total resection during endonasal endoscopic resection of craniopharyngiomas?

PubMed

Omay, Sacit Bulent; Almeida, João Paulo; Chen, Yu-Ning; Shetty, Sathwik R; Liang, Buqing; Ni, Shilei; Anand, Vijay K; Schwartz, Theodore H

2017-11-24

OBJECTIVE Craniopharyngiomas arise from the pituitary stalk, and in adults they are generally located posterior to the chiasm extending up into the third ventricle. The extended endonasal approach (EEA) can provide an ideal corridor between the bottom of the optic chiasm and the top of the pituitary gland (chiasm-pituitary corridor [CPC]) for their removal. A narrow CPC in patients with a prefixed chiasm and a large tumor extending up and behind the chiasm has been considered a contraindication to EEA, with a high risk of visual deterioration and subtotal resection. METHODS A database of all patients treated in the authors' center (Weill Cornell Medical College, NewYork-Presbyterian Hospital) between July 2004 and August 2016 was reviewed. Patients with craniopharyngiomas who underwent EEA with the goal of gross-total resection (GTR) were included in the study. Patients with postfixed chiasm or limited available preoperative imaging were excluded. Using preoperative contrast-enhanced T1-weighted sagittal midline MR images, the authors calculated the CPC as well as the distance from the chiasm to the top of the tumor (CTOT). From these numbers, they calculated a ratio of the CPC to the CTOT as a measure of difficulty in removing the tumors through the EEA and called this ratio the corridor index (CI). The relationship between the CI and the ability to achieve GTR and visual outcome were measured. RESULTS Thirty-four patients were included in the study. The mean CPC was 10.1 mm (range 5.2-19.1 mm). The mean CTOT was 12.8 mm (range 0-28.3 mm). The median CI was 0.8; the CI ranged from 0.4 to infinity (for tumors with a CTOT of 0). Thirty-two patients had GTR (94.1%) and 2 had subtotal resection. The CPC value had no relationship with our ability to achieve GTR and no effect on visual or endocrine outcome. CONCLUSIONS EEA for craniopharyngioma is generally considered the first-line surgical approach. Although a narrow corridor between the top of the pituitary gland and the bottom of the chiasm may seem to be a relative contraindication to surgery for larger tumors, the authors' data do not bear this out. EEA appears to be a successful technique for the majority of midline craniopharyngiomas.

Perspectives for integrating human and environmental exposure assessments.

PubMed

Ciffroy, P; Péry, A R R; Roth, N

2016-10-15

Integrated Risk Assessment (IRA) has been defined by the EU FP7 HEROIC Coordination action as "the mutual exploitation of Environmental Risk Assessment for Human Health Risk Assessment and vice versa in order to coherently and more efficiently characterize an overall risk to humans and the environment for better informing the risk analysis process" (Wilks et al., 2015). Since exposure assessment and hazard characterization are the pillars of risk assessment, integrating Environmental Exposure assessment (EEA) and Human Exposure assessment (HEA) is a major component of an IRA framework. EEA and HEA typically pursue different targets, protection goals and timeframe. However, human and wildlife species also share the same environment and they similarly inhale air and ingest water and food through often similar overlapping pathways of exposure. Fate models used in EEA and HEA to predict the chemicals distribution among physical and biological media are essentially based on common properties of chemicals, and internal concentration estimations are largely based on inter-species (i.e. biota-to-human) extrapolations. Also, both EEA and HEA are challenged by increasing scientific complexity and resources constraints. Altogether, these points create the need for a better exploitation of all currently existing data, experimental approaches and modeling tools and it is assumed that a more integrated approach of both EEA and HEA may be part of the solution. Based on the outcome of an Expert Workshop on Extrapolations in Integrated Exposure Assessment organized by the HEROIC project in January 2014, this paper identifies perspectives and recommendations to better harmonize and extrapolate exposure assessment data, models and methods between Human Health and Environmental Risk Assessments to support the further development and promotion of the concept of IRA. Ultimately, these recommendations may feed into guidance showing when and how to apply IRA in the regulatory decision-making process for chemicals. Copyright © 2015 Elsevier B.V. All rights reserved.

New sulphiding method for steel and cast iron parts

NASA Astrophysics Data System (ADS)

Tarelnyk, V.; Martsynkovskyy, V.; Gaponova, O.; Konoplianchenko, Ie; Dovzyk, M.; Tarelnyk, N.; Gorovoy, S.

2017-08-01

A new method for sulphiding steel and cast iron part surfaces by electroerosion alloying (EEA) with the use of a special electrode is proposed, which method is characterized in that while manufacturing the electrode, on its surface, in any known manner (punching, threading, pulling, etc.), there is formed at least a recess to be filled with sulfur as a consistent material, and then there is produced EEA by the obtained electrode without waiting for the consistent material to become dried.

Copy number variations and genetic admixtures in three Xinjiang ethnic minority groups

PubMed Central

Lou, Haiyi; Li, Shilin; Jin, Wenfei; Fu, Ruiqing; Lu, Dongsheng; Pan, Xinwei; Zhou, Huaigu; Ping, Yuan; Jin, Li; Xu, Shuhua

2015-01-01

Xinjiang is geographically located in central Asia, and it has played an important historical role in connecting eastern Eurasian (EEA) and western Eurasian (WEA) people. However, human population genomic studies in this region have been largely underrepresented, especially with respect to studies of copy number variations (CNVs). Here we constructed the first CNV map of the three major ethnic minority groups, the Uyghur, Kazakh and Kirgiz, using Affymetrix Genome-Wide Human SNP Array 6.0. We systematically compared the properties of CNVs we identified in the three groups with the data from representatives of EEA and WEA. The analyses indicated a typical genetic admixture pattern in all three groups with ancestries from both EEA and WEA. We also identified several CNV regions showing significant deviation of allele frequency from the expected genome-wide distribution, which might be associated with population-specific phenotypes. Our study provides the first genome-wide perspective on the CNVs of three major Xinjiang ethnic minority groups and has implications for both evolutionary and medical studies. PMID:25026903

Copy number variations and genetic admixtures in three Xinjiang ethnic minority groups.

PubMed

Lou, Haiyi; Li, Shilin; Jin, Wenfei; Fu, Ruiqing; Lu, Dongsheng; Pan, Xinwei; Zhou, Huaigu; Ping, Yuan; Jin, Li; Xu, Shuhua

2015-04-01

Xinjiang is geographically located in central Asia, and it has played an important historical role in connecting eastern Eurasian (EEA) and western Eurasian (WEA) people. However, human population genomic studies in this region have been largely underrepresented, especially with respect to studies of copy number variations (CNVs). Here we constructed the first CNV map of the three major ethnic minority groups, the Uyghur, Kazakh and Kirgiz, using Affymetrix Genome-Wide Human SNP Array 6.0. We systematically compared the properties of CNVs we identified in the three groups with the data from representatives of EEA and WEA. The analyses indicated a typical genetic admixture pattern in all three groups with ancestries from both EEA and WEA. We also identified several CNV regions showing significant deviation of allele frequency from the expected genome-wide distribution, which might be associated with population-specific phenotypes. Our study provides the first genome-wide perspective on the CNVs of three major Xinjiang ethnic minority groups and has implications for both evolutionary and medical studies.

The double stapling technique for low anterior resection. Results, modifications, and observations.

PubMed Central

Griffen, F D; Knight, C D; Whitaker, J M; Knight, C D

1990-01-01

Since the introduction of the end-to-end anastomosis (EEA) stapler for rectal reconstruction, we have used a modification of the conventional technique in which the lower rectal segment is closed with the linear stapler (TA-55) and the anastomosis is performed using the EEA instrument across the linear staple line (double stapling technique). Our experience with this procedure includes stapled colorectal anastomoses in 75 patients and is the basis for the report. This review presents the details and advantages of the technique and the results. Complications include two patients with anastomotic leak (2.7%), and two with stenosis that required treatment (2.7%). Protective colostomy was not done in this series. There were no deaths. Our experience and that of others suggests that this modification of the EEA technique can allow a lower anastomosis in some patients, and that it can be done with greater safety and facility. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. Fig. 6. PMID:2357137

Influence of Precipitation Regime on Microbial Decomposition Patterns in Semi-Arid Ecosystems

NASA Astrophysics Data System (ADS)

Feris, K. P.; Jilek, C.; Huber, D. P.; Reinhardt, K.; deGraaff, M.; Lohse, K.; Germino, M.

2011-12-01

In water-limited semi-arid sagebrush steppe ecosystems predicted changes in climate may manifest as a shift from historically winter/snow-dominated precipitation regimes to one dominated by spring rains. In these ecosystems soil microorganisms play a vital role in linking the effects of water availability and plant productivity to biogeochemical cycling. Patterns of soil microbial catalyzed organic matter decomposition patters (i.e. patterns of extracellular enzyme activity (EEA)) are thought to depend upon the quantity and quality of soil organic matter (SOM), pH, and mean annual precipitation (Sinsabaugh, 2008), and less on the timing and magnitude of precipitation. However, sagebrush-steppe plant communities respond strongly to changes in the timing and magnitude of precipitation, and preliminary findings by our group suggest that corresponding changes in SOM quantity, quality, N-cycle dynamics, and soil structure are occurring. Therefore, we hypothesized: 1) Shifts in the timing and magnitude of precipitation would indirectly affect soil microbial decomposition patterns via responses in the plant community structure; and 2) Changes in precipitation patterns can directly affect soil microbial community structure and function, in effect uncoupling the interaction between plant community structure and soil community structure. We tested our hypotheses by determining the influence of experimentally manipulated timing and magnitude of precipitation on soil microbial EEA using standard flourometric assays in soils sampled under plant canopies and plant interspaces. We assessed this response in a mature (18 + years) ecohydrologic field experiment in eastern Idaho that annually imitates three possible post climatic-shift precipitation regimes (Ambient (AMB): no additional precipitation, ~200mm annually; Summer (SUMM): 200mm provisioned at 50mm bi-weekly starting in June; and Fall/Spring (F/S): 200mm provisioned over 1-2 weeks in October or April) (n=3). Within plant interspaces Beta glucosaminide activity increased by 18% in treatments receiving additional F/S precipitation, whereas alpha glucopyranoside activity was lower in the F/S and SUMM plots. Conversely, underplant canopies alpha glucopyranoside activity increased by 15% in the SUMM and F/S precipitation treatments. Across treatments and sampling types (i.e. plant canopy vs. interspace), cellobioside activity levels are consistently elevated in response to additional precipitation compared to those of the control plots. When coupled with recent preliminary findings by our group regarding changes in plant and microbial community structure and SOM, C-storage, and soil structural responses, these preliminary findings suggest that 1) microbial community structure and function respond both directly and indirectly to changes in climate, and 2) thus provide a mechanism for changes in plant community structure to feed-forward to affect soil carbon decomposition patterns and ultimately soil carbon storage potential.

A twin study of specific bulimia nervosa symptoms.

PubMed

Mazzeo, S E; Mitchell, K S; Bulik, C M; Aggen, S H; Kendler, K S; Neale, M C

2010-07-01

Twin studies have suggested that additive genetic factors significantly contribute to liability to bulimia nervosa (BN). However, the diagnostic criteria for BN remain controversial. In this study, an item-factor model was used to examine the BN diagnostic criteria and the genetic and environmental contributions to BN in a population-based twin sample. The validity of the equal environment assumption (EEA) for BN was also tested. Participants were 1024 female twins (MZ n=614, DZ n=410) from the population-based Mid-Atlantic Twin Registry. BN was assessed using symptom-level (self-report) items consistent with DSM-IV and ICD-10 diagnostic criteria. Items assessing BN were included in an item-factor model. The EEA was measured by items assessing similarity of childhood and adolescent environment, which have demonstrated construct validity. Scores on the EEA factor were used to specify the degree to which twins shared environmental experiences in this model. The EEA was not violated for BN. Modeling results indicated that the majority of the variance in BN was due to additive genetic factors. There was substantial variability in additive genetic and environmental contributions to specific BN symptoms. Most notably, vomiting was very strongly influenced by additive genetic factors, while other symptoms were much less heritable, including the influence of weight on self-evaluation. These results highlight the importance of assessing eating disorders at the symptom level. Refinement of eating disorder phenotypes could ultimately lead to improvements in treatment and targeted prevention, by clarifying sources of variation for specific components of symptomatology.

Organic matter quantity and source affects microbial community structure and function following volcanic eruption on Kasatochi Island, Alaska.

PubMed

Zeglin, Lydia H; Wang, Bronwen; Waythomas, Christopher; Rainey, Frederick; Talbot, Sandra L

2016-01-01

In August 2008, Kasatochi volcano erupted and buried a small island in pyroclastic deposits and fine ash; since then, microbes, plants and birds have begun to re-colonize the initially sterile surface. Five years post-eruption, bacterial 16S rRNA gene and fungal internal transcribed spacer (ITS) copy numbers and extracellular enzyme activity (EEA) potentials were one to two orders of magnitude greater in pyroclastic materials with organic matter (OM) inputs relative to those without, despite minimal accumulation of OM (< 0.2%C). When normalized by OM levels, post-eruptive surfaces with OM inputs had the highest β-glucosidase, phosphatase, NAGase and cellobiohydrolase activities, and had microbial population sizes approaching those in reference soils. In contrast, the strongest factor determining bacterial community composition was the dominance of plants versus birds as OM input vectors. Although soil pH ranged from 3.9 to 7.0, and %C ranged 100×, differentiation between plant- and bird-associated microbial communities suggested that cell dispersal or nutrient availability are more likely drivers of assembly than pH or OM content. This study exemplifies the complex relationship between microbial cell dispersal, soil geochemistry, and microbial structure and function; and illustrates the potential for soil microbiota to be resilient to disturbance. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

Reduction in undiagnosed HIV infection in the European Union/European Economic Area, 2012 to 2016.

PubMed

van Sighem, Ard; Pharris, Anastasia; Quinten, Chantal; Noori, Teymur; Amato-Gauci, Andrew J

2017-11-01

It is well-documented that early HIV diagnosis and linkage to care reduces morbidity and mortality as well as HIV transmission. We estimated the median time from HIV infection to diagnosis in the European Union/European Economic Area (EU/EEA) at 2.9 years in 2016, with regional variation. Despite evidence of a decline in the number of people living with undiagnosed HIV in the EU/EEA, many remain undiagnosed, including 33% with more advanced HIV infection (CD4 < 350 cells/mm3).

Reduction in undiagnosed HIV infection in the European Union/European Economic Area, 2012 to 2016

PubMed Central

van Sighem, Ard; Pharris, Anastasia; Quinten, Chantal; Noori, Teymur; Amato-Gauci, Andrew J

2017-01-01

It is well-documented that early HIV diagnosis and linkage to care reduces morbidity and mortality as well as HIV transmission. We estimated the median time from HIV infection to diagnosis in the European Union/European Economic Area (EU/EEA) at 2.9 years in 2016, with regional variation. Despite evidence of a decline in the number of people living with undiagnosed HIV in the EU/EEA, many remain undiagnosed, including 33% with more advanced HIV infection (CD4 < 350 cells/mm3). PMID:29208159

Vulnerability and adaptation to water scarcity in the European Alps

NASA Astrophysics Data System (ADS)

Isoard, S.; McCallum, S.; Prutsch, A.; Benno Hain, B.; Schauser, I.

2009-04-01

The European Environment Agency (EEA) has recently undertaken a project addressing vulnerability and adaptation to water availability in the European Alps. Mountains are indeed one of the most vulnerable regions to climate change in Europe (EEA 2008, IPCC 2007).The Alps, in particular, can be presented as the ‘water towers' of Europe (the amount of water delivered by the Alps allocates 40% of EU consumption) where changes in water availability affect all socio-economical sectors. This therefore makes adaptation actions a regional topic with an outstanding European dimension. The specific objectives of the study were to highlight the importance of the Alps in their function as ‘water towers' for Europe and analyse the vulnerability of the Alpine Region with regard to impacts of climate change (but also to global change as a whole) focussing on water availability. Given the EU and regional policy background with regard to adaptation and water issues, the study assessed the possible needs, constraints and opportunities for adaptation to the adverse impacts for various sectors pending on water resources. Findings of this activity expanded the knowledge base, fed into the preparation of European Commission's 2009 White Paper and the Alpine Convention 2009 Report on the State of the Alps, and complemented other recent studies (e.g. OECD 2007, European Parliament Committee on Agriculture & Rural Development 2008). The method used for the study relied on the one hand on findings from recent key publications on climate change impacts (EEA 2008, IPCC 2007) and EU research projects (e.g. ClimChAlp, ProClim); on the other side it was based on selected case studies chosen within the four climatic regions of the eight Alpine countries for which an extensive series of interviews with local and regional stakeholders and decision makers has been undertaken. The interviewees had been directly involved in designing and implementing water availability-related adaptation measures relating to the respective case studies. The analysis of the case studies highlighted key drivers of adaptation measures, the multi-level governance surrounding this type of policy actions, success factors and barriers (e.g. lack of knowledge, governance shortcomings, coordination issues, spatial and temporal distribution, and competition for resources) that prevent more extensive adaptation actions. Examples of good practices were given as well as further adaptation needs in view of future climate change. Since the study relied mainly on stakeholders' interviews, the major focus lies on building local/regional adaptive capacity that effectively supports adaptation as a social and institutional process. It also provided a synthesis of key vulnerabilities and adaptation options across Europe/Alpine river catchments and recommendations on how to transfer experiences and knowledge in practical implementation of adaptation measures, in particular to other mountain regions (e.g. Carpathian). In this context the study supports regional and local administrations in making informed decisions to better develop and implement adaptation strategies. This EEA study was conducted by the European Topic Centre on Air and Climate Change (UBA Germany, UBA Austria, Potsdam Institute for Climate Impact Research) in cooperation with the Permanent Secretariat of the Alpine Convention (Innsbruck), the European Academy (Bolzano) and the Environment Protection Agency of Slovenia. The study will be published in the first half of 2009 as an EEA Technical Report and will be actively disseminated to stakeholders and decision-makers in the Alpine region through best suited events in collaboration with the Alpine Convention.

Restricted access to antiretroviral treatment for undocumented migrants: a bottle neck to control the HIV epidemic in the EU/EEA.

PubMed

Deblonde, Jessika; Sasse, André; Del Amo, Julia; Burns, Fiona; Delpech, Valerie; Cowan, Susan; Levoy, Michele; Keith, Lilana; Pharris, Anastasia; Amato-Gauci, Andrew; Noori, Teymur

2015-12-10

In the European Union/European Economic Area (EU/EEA), migrants from high-endemic countries are disproportionately affected by HIV. Between 2007 and 2012, migrants represented 39 % of reported HIV cases. There is growing evidence that a significant proportion of HIV acquisition among migrant populations occurs after their arrival in Europe. Migrants are confronted with multiple risk factors that shape patterns of population HIV susceptibility and vulnerability, which simultaneously affect HIV transmission. Undocumented migrants incur additional risks for contracting HIV due to limited access to adequate health care services, protection and justice, alongside insecure housing and employment conditions. All EU/EEA countries have ratified a number of international and regional human rights instruments that enshrine access to health care as a human right that should be available to everyone without discrimination. From a clinical and public health perspective, early HIV care and treatment is associated with viral suppression, improved health outcomes and reductions in transmission risks. A current challenge of the HIV epidemic is to reach the highest proportion of overall viral suppression among people living with HIV in order to impact on HIV transmission. Although the majority of EU/EEA countries regard migrants as an important sub-population for their national responses to HIV, and despite the overwhelming evidence of the individual and public health benefits associated with HIV care and treatment, a significant number of EU/EEA countries do not provide antiretroviral treatment to undocumented migrants. HIV transmission dynamics in migrant populations depend on the respective weight of all risk and vulnerability factors to which they are exposed, which act together in a synergistic way. People who are not linked to HIV care will continue to unwillingly contribute to the on-going transmission of HIV. Following the recommendations of the European Union Agency for Fundamental Rights, ensuring access to HIV-care for all sub-populations, including undocumented migrants, would fulfil the human rights of those populations and also strengthen the control of HIV incidence among those not currently able to access HIV care.

Impact of hepatitis B vaccination on acute hepatitis B epidemiology in European Union/European Economic Area countries, 2006 to 2014

PubMed Central

Miglietta, Alessandro; Quinten, Chantal; Lopalco, Pier Luigi; Duffell, Erika

2018-01-01

Hepatitis B prevention in European Union/European Economic Area (EU/EEA) countries relies on vaccination programmes. We describe the epidemiology of acute hepatitis B virus (HBV) at country and EU/EEA level during 2006–2014. Using a multi-level mixed-effects Poisson regression model we assessed differences in the acute HBV infection notification rates between groups of countries that started universal HBV vaccination before/in vs after 1995; implemented or not a catch-up strategy; reached a vaccine coverage ≥ 95% vs < 95% and had a hepatitis B surface antigen prevalence ≥ 1% vs < 1%. Joinpoint regression analysis was used to assess trends by groups of countries, and additional Poisson regression models to evaluate the association between three-dose HBV vaccine coverage and acute HBV infection notification rates at country and EU/EEA level. The EU/EEA acute HBV infection notification rate decreased from 1.6 per 100,000 population in 2006 to 0.7 in 2014. No differences (p > 0.05) were found in the acute HBV infection notification rates between groups of countries, while as vaccine coverage increased, such rates decreased (p < 0.01). Countries with universal HBV vaccination before 1995, a catch-up strategy, and a vaccine coverage ≥ 95% had significant decreasing trends (p < 0.01). Ending HBV transmission in Europe by 2030 will require high vaccine coverage delivered through universal programmes, supported, where appropriate, by catch-up vaccination campaigns. PMID:29439751

Gonorrhoea and gonococcal antimicrobial resistance surveillance networks in the WHO European Region, including the independent countries of the former Soviet Union.

PubMed

Unemo, Magnus; Ison, Catherine A; Cole, Michelle; Spiteri, Gianfranco; van de Laar, Marita; Khotenashvili, Lali

2013-12-01

Antimicrobial resistance (AMR) in Neisseria gonorrhoeae has emerged for essentially all antimicrobials following their introduction into clinical practice. During the latest decade, susceptibility to the last remaining options for antimicrobial monotherapy, the extended-spectrum cephalosporins (ESC), has markedly decreased internationally and treatment failures with these ESCs have been verified. In response to this developing situation, WHO and the European Centre for Disease Prevention and Control (ECDC) have published global and region-specific response plans, respectively. One main component of these action/response plans is to enhance the surveillance of AMR and treatment failures. This paper describes the perspectives from the diverse WHO European Region (53 countries), including the independent countries of the former Soviet Union, regarding gonococcal AMR surveillance networks. The WHO European Region has a high prevalence of resistance to all previously recommended antimicrobials, and most of the first strictly verified treatment failures with cefixime and ceftriaxone were also reported from Europe. In the European Union/European Economic Area (EU/EEA), the European gonococcal antimicrobial surveillance programme (Euro-GASP) funded by the ECDC is running. In 2011, the Euro-GASP included 21/31 (68%) EU/EEA countries, and the programme is further strengthened annually. However, in the non-EU/EEA countries, internationally reported and quality assured gonococcal AMR data are lacking in 87% of the countries and, worryingly, appropriate support for establishment of a GASP is still lacking. Accordingly, national and international support, including political and financial commitment, for gonococcal AMR surveillance in the non-EU/EEA countries of the WHO European Region is essential.

Echocardiographic estimation of left atrial pressure in beagle dogs with experimentally-induced mitral valve regurgitation.

PubMed

Ishikawa, Taisuke; Fukushima, Ryuji; Suzuki, Shuji; Miyaishi, Yuka; Nishimura, Taiki; Hira, Satoshi; Hamabe, Lina; Tanaka, Ryou

2011-08-01

Non-invasive and immediate estimation of left atrial pressure (LAP) is very useful for the management of mitral regurgitation (MR), and many reports have assessed echocardiographic estimations of LAP to date. However, it has been unclear of which examination and evaluate article possess the best accuracy for the MR severity. The present research aims to establish the echocardiographic estimation equation of LAP that is well applicable for clinical MR dogs. After the chordae tendineae rupture was experimentally induced via left atriotomy in six healthy beagle dogs (three males and three females, two years old, weighing between 9.8 to 12.8 kg), a radio telemetry transmitter catheter was inserted, which allows the continuous recordings of LAP without the use of sedation. Approximately 5 weeks after the surgery, echocardiographic examination, indirect blood pressure measurement, measurement of plasma atrial natriuretic peptide (ANP) and LAP measurement by way of the radio telemetry system was performed simultaneously. Subsequently, simple linear regression equations between LAP and each variable were obtained, and the equations were evaluated whether to be applicable for clinical MR dogs. As a result, the ratio of early diastolic mitral flow to early diastolic lateral mitral annulus velocity (E/Ea) had the strongest correlation as maximum LAP=7.03*(E/Ea)-54.86 (r=0.74), and as mean LAP=4.94*(E/Ea)-40.37 (r=0.70) among the all variables. Therefore, these two equations associated with E/Ea should bring more precise and instant estimations of maximum and mean LAP in clinical MR dogs.

National Health Service Principles as Experienced by Vulnerable London Migrants in "Austerity Britain": A Qualitative Study of Rights, Entitlements, and Civil-Society Advocacy

PubMed Central

Rafighi, Elham; Poduval, Shoba; Legido-Quigley, Helena; Howard, Natasha

2016-01-01

Background: Recent British National Health Service (NHS) reforms, in response to austerity and alleged ‘health tourism,’ could impose additional barriers to healthcare access for non-European Economic Area (EEA) migrants. This study explores policy reform challenges and implications, using excerpts from the perspectives of non-EEA migrants and health advocates in London. Methods: A qualitative study design was selected. Data were collected through document review and 22 in-depth interviews with non-EEA migrants and civil-society organisation representatives. Data were analysed thematically using the NHS principles. Results: The experiences of those ‘vulnerable migrants’ (ie, defined as adult non-EEA asylum-seekers, refugees, undocumented, low-skilled, and trafficked migrants susceptible to marginalised healthcare access) able to access health services were positive, with healthcare professionals generally demonstrating caring attitudes. However, general confusion existed about entitlements due to recent NHS changes, controversy over ‘health tourism,’ and challenges registering for health services or accessing secondary facilities. Factors requiring greater clarity or improvement included accessibility, communication, and clarity on general practitioner (GP) responsibilities and migrant entitlements. Conclusion: Legislation to restrict access to healthcare based on immigration status could further compromise the health of vulnerable individuals in Britain. This study highlights current challenges in health services policy and practice and the role of non-governmental organizations (NGOs) in healthcare advocacy (eg, helping the voices of the most vulnerable reach policy-makers). Thus, it contributes to broadening national discussions and enabling more nuanced interpretation of ongoing global debates on immigration and health. PMID:27694650

National Health Service Principles as Experienced by Vulnerable London Migrants in "Austerity Britain": A Qualitative Study of Rights, Entitlements, and Civil-Society Advocacy.

PubMed

Rafighi, Elham; Poduval, Shoba; Legido-Quigley, Helena; Howard, Natasha

2016-05-08

Recent British National Health Service (NHS) reforms, in response to austerity and alleged 'health tourism,' could impose additional barriers to healthcare access for non-European Economic Area (EEA) migrants. This study explores policy reform challenges and implications, using excerpts from the perspectives of non-EEA migrants and health advocates in London. A qualitative study design was selected. Data were collected through document review and 22 in-depth interviews with non-EEA migrants and civil-society organisation representatives. Data were analysed thematically using the NHS principles. The experiences of those 'vulnerable migrants' (ie, defined as adult non-EEA asylum-seekers, refugees, undocumented, low-skilled, and trafficked migrants susceptible to marginalised healthcare access) able to access health services were positive, with healthcare professionals generally demonstrating caring attitudes. However, general confusion existed about entitlements due to recent NHS changes, controversy over 'health tourism,' and challenges registering for health services or accessing secondary facilities. Factors requiring greater clarity or improvement included accessibility, communication, and clarity on general practitioner (GP) responsibilities and migrant entitlements. Legislation to restrict access to healthcare based on immigration status could further compromise the health of vulnerable individuals in Britain. This study highlights current challenges in health services policy and practice and the role of non-governmental organizations (NGOs) in healthcare advocacy (eg, helping the voices of the most vulnerable reach policy-makers). Thus, it contributes to broadening national discussions and enabling more nuanced interpretation of ongoing global debates on immigration and health. © 2016 by Kerman University of Medical Sciences

Endoscopic Endonasal Optic Nerve Decompression for Fibrous Dysplasia

PubMed Central

DeKlotz, Timothy R.; Stefko, S. Tonya; Fernandez-Miranda, Juan C.; Gardner, Paul A.; Snyderman, Carl H.; Wang, Eric W.

2016-01-01

Objective To evaluate visual outcomes and potential complications for optic nerve decompression using an endoscopic endonasal approach (EEA) for fibrous dysplasia. Design Retrospective chart review of patients with fibrous dysplasia causing extrinsic compression of the canalicular segment of the optic nerve that underwent an endoscopic endonasal optic nerve decompression at the University of Pittsburgh Medical Center from 2010 to 2013. Main Outcome Measures The primary outcome measure assessed was best-corrected visual acuity (BCVA) with secondary outcomes, including visual field testing, color vision, and complications associated with the intervention. Results A total of four patients and five optic nerves were decompressed via an EEA. All patients were symptomatic preoperatively and had objective findings compatible with compressive optic neuropathy: decreased visual acuity was noted preoperatively in three patients while the remaining patient demonstrated an afferent pupillary defect. BCVA improved in all patients postoperatively. No major complications were identified. Conclusion EEA for optic nerve decompression appears to be a safe and effective treatment for patients with compressive optic neuropathy secondary to fibrous dysplasia. Further studies are required to identify selection criteria for an open versus an endoscopic approach. PMID:28180039

APPL endosomes are not obligatory endocytic intermediates but act as stable cargo-sorting compartments

PubMed Central

Kalaidzidis, Inna; Miaczynska, Marta; Brewińska-Olchowik, Marta; Hupalowska, Anna; Ferguson, Charles; Parton, Robert G.; Kalaidzidis, Yannis

2015-01-01

Endocytosis allows cargo to enter a series of specialized endosomal compartments, beginning with early endosomes harboring Rab5 and its effector EEA1. There are, however, additional structures labeled by the Rab5 effector APPL1 whose role in endocytic transport remains unclear. It has been proposed that APPL1 vesicles are transport intermediates that convert into EEA1 endosomes. Here, we tested this model by analyzing the ultrastructural morphology, kinetics of cargo transport, and stability of the APPL1 compartment over time. We found that APPL1 resides on a tubulo-vesicular compartment that is capable of sorting cargo for recycling or degradation and that displays long lifetimes, all features typical of early endosomes. Fitting mathematical models to experimental data rules out maturation of APPL1 vesicles into EEA1 endosomes as a primary mechanism for cargo transport. Our data suggest instead that APPL1 endosomes represent a distinct population of Rab5-positive sorting endosomes, thus providing important insights into the compartmental organization of the early endocytic pathway. PMID:26459602

Endoscopic Endonasal Approach for Resection of a Pediatric Craniopharyngioma: Operative Video and Technical Nuances.

PubMed

Liu, James K; Eloy, Jean Anderson

2018-04-01

We present a pediatric case of a retrochiasmatic craniopharyngioma in the suprasellar region with third ventricular extension that was resected through a purely endoscopic endonasal approach (EEA) via the transplanum transtuberculum corridor. The patient is a 12-year-old boy who presented with progressive visual loss and panhypopituitarism. The EEA allows direct visualization of the undersurface of the optic chiasm and hypothalamus so that safe and meticulous tumor dissection can be performed to preserve these critical neurovascular structures. This video atlas demonstrates the operative technique and surgical nuances of the endoscopic skull base approach, microdissection of the tumor from the critical neurovascular structures, and multilayered reconstruction of the skull base defect with a nasoseptal flap. A gross total resection was achieved, and the patient was neurologically intact with improved visual acuity and visual fields. In summary, the EEA via the transplanum transtuberculum corridor is an important strategy in the armamentarium for surgical management of pediatric craniopharyngiomas. The link to the video can be found at: https://youtu.be/bmgO_PMRHPk .

Improved infrared-sensing running wheel systems with an effective exercise activity indicator.

PubMed

Chen, Chi-Chun; Chang, Ming-Wen; Chang, Ching-Ping; Chang, Wen-Ying; Chang, Shin-Chieh; Lin, Mao-Tsun; Yang, Chin-Lung

2015-01-01

This paper describes an infrared-sensing running wheel (ISRW) system for the quantitative measurement of effective exercise activity in rats. The ISRW system provides superior exercise training compared with commercially available traditional animal running platforms. Four infrared (IR) light-emitting diode/detector pairs embedded around the rim of the wheel detect the rat's real-time position; the acrylic wheel has a diameter of 55 cm and a thickness of 15 cm, that is, it is larger and thicker than traditional exercise wheels, and it is equipped with a rubber track. The acrylic wheel hangs virtually frictionless, and a DC motor with an axially mounted rubber wheel, which has a diameter of 10 cm, drives the acrylic wheel from the outer edge. The system can automatically train rats to run persistently. The proposed system can determine effective exercise activity (EEA), with the IR sensors (which are connected to a conventional PC) recording the rat exercise behavior. A prototype of the system was verified by a hospital research group performing ischemic stroke experiments on rats by considering middle cerebral artery occlusion. The experimental data demonstrated that the proposed system provides greater neuroprotection in an animal stroke model compared with a conventional treadmill and a motorized running wheel for a given exercise intensity. The quantitative exercise effectiveness indicator showed a 92% correlation between an increase in the EEA and a decrease in the infarct volume. This indicator can be used as a noninvasive and objective reference in clinical animal exercise experiments.

Soil Microbial Activity Responses to Fire in a Semi-arid Savannah Ecosystem Pre- and Post-Monsoon Season

NASA Astrophysics Data System (ADS)

Jimenez, J. R.; Raub, H. D.; Jong, E. L.; Muscarella, C. R.; Smith, W. K.; Gallery, R. E.

2017-12-01

Extracellular enzyme activities (EEA) of soil microorganisms can act as important proxies for nutrient limitation and turnover in soil and provide insight into the biochemical requirements of microbes in terrestrial ecosystems. In semi-arid ecosystems, microbial activity is influenced by topography, disturbances such as fire, and seasonality from monsoon rains. Previous studies from forest ecosystems show that microbial communities shift to similar compositions after severe fires despite different initial conditions. In semi-arid ecosystems with high spatial heterogeniety, we ask does fire lead to patch intensification or patch homogenization and how do monsoon rains influence the successional trajectories of microbial responses? We analyzed microbial activity and soil biogeochemistry throughout the monsoon season in paired burned and unburned sites in the Santa Rita Experimental Range, AZ. Surface soil (5cm) from bare-ground patches, bole, canopy drip line, and nearby grass patches for 5 mesquite trees per site allowed tests of spatiotemporal responses to fire and monsoon rain. Microbial activity was low during the pre-monsoon season and did not differ between the burned and unburned sites. We found greater activity near mesquite trees that reflects soil water and nutrient availability. Fire increased soil alkalinity, though soils near mesquite trees were less affected. Soil water content was significantly higher in the burned sites post-monsoon, potentially reflecting greater hydrophobicity of burned soils. Considering the effects of fire in these semi-arid ecosystems is especially important in the context of the projected changing climate regime in this region. Assessing microbial community recovery pre-, during, and post-monsoon is important for testing predictions about whether successional pathways post-fire lead to recovery or novel trajectories of communities and ecosystem function.

Modelling Levels of Collaboration Among Health and Social Care Professionals in the Management of a Mental Health Plan.

PubMed

Luzi, Daniela; Pecoraro, Fabrizio; Tamburis, Oscar

2018-01-01

Professional collaboration among health and social care providers is considered an essential pattern to improve the integration of care. This is particularly important considering the planning activities for children with complex conditions. In this paper the level of collaboration among professionals in the development and implementation of the personalized plan in the mental health domain is analysed across 30 EU/EEA countries within the MOCHA project.

Contralateral transmaxillary corridor: an augmented endoscopic approach to the petrous apex.

PubMed

Patel, Chirag R; Wang, Eric W; Fernandez-Miranda, Juan C; Gardner, Paul A; Snyderman, Carl H

2017-10-20

OBJECTIVE The endoscopic endonasal approach (EEA) has been shown to be an effective means of accessing lesions of the petrous apex. Lesions that are lateral to the paraclival segment of the internal carotid artery (ICA) require lateralization of the paraclival segment of the ICA or a transpterygoid infrapetrous approach. In this study the authors studied the feasibility of adding a contralateral transmaxillary (CTM) corridor to provide greater access to the petrous apex with decreased need for manipulation of the ICA. METHODS Using image guidance, EEA and CTM extension were performed bilaterally on 5 cadavers. The anterior wall of the sphenoid sinus and rostrum were removed. The angle of the surgical approach from the axis of the petrous segment of the ICA was measured. Five illustrative clinical cases are presented. RESULTS The CTM corridor required a partial medial maxillectomy. When measured from the axis of the petrous ICA, the CTM corridor decreased the angle from 44.8° ± 2.78° to 20.1° ± 4.31°, a decrease of 24.7° ± 2.58°. Drilling through the CTM corridor allowed the drill to reach lateral aspects of the petrous apex that would have required lateralization of the ICA or would not have been accessible via EEA. The CTM corridor allowed us to achieve gross-total resection of the petrous apex region in 5 clinical cases with significant paraclival extension. CONCLUSIONS The CTM corridor is a feasible extension to the standard EEA to the petrous apex that offers a more lateral trajectory with improved access. This approach may reduce the risk and morbidity associated with manipulation of the paraclival ICA.

Altered TGF-β endocytic trafficking contributes to the increased signaling in Marfan syndrome.

PubMed

Siegert, Anna-Maria; Serra-Peinado, Carla; Gutiérrez-Martínez, Enric; Rodríguez-Pascual, Fernando; Fabregat, Isabel; Egea, Gustavo

2018-02-01

The main cardiovascular alteration in Marfan syndrome (MFS) is the formation of aortic aneurysms in which augmented TGF-β signaling is reported. However, the primary role of TGF-β signaling as a molecular link between the genetic mutation of fibrillin-1 and disease onset is controversial. The compartmentalization of TGF-β endocytic trafficking has been shown to determine a signaling response in which clathrin-dependent internalization leads to TGF-β signal propagation, and caveolin-1 (CAV-1) associated internalization leads to signal abrogation. We here studied the contribution of endocytic trafficking compartmentalization to increased TGF-β signaling in vascular smooth muscle cells (VSMC) from MFS patients. We examined molecular components involved in clathrin- (SARA, SMAD2) and caveolin-1- (SMAD7, SMURF2) dependent endocytosis. Marfan VSMC showed higher recruitment of SARA and SMAD2 to membranes and their increased interaction with TGF-β receptor II, as well as higher colocalization of SARA with the early endosome marker EEA1. We assessed TGF-β internalization using a biotinylated ligand (b-TGF-β), which colocalized equally with either EEA1 or CAV-1 in VSMC from Marfan patients and controls. However, in Marfan cells, colocalization of b-TGF-β with SARA and EEA1 was increased and accompanied by decreased colocalization with CAV-1 at EEA1-positive endosomes. Moreover, Marfan VSMC showed higher transcriptional levels and membrane enrichment of RAB5. Our results indicate that increased RAB5-associated SARA localization to early endosomes facilitates its TGF-β receptor binding and phosphorylation of signaling mediator SMAD2 in Marfan VSMC. This is accompanied by a reduction of TGF-β sorting into multifunctional vesicles containing cargo from both internalization pathways. Copyright © 2017 Elsevier B.V. All rights reserved.

Elastic Pitch Beam Tail Rotor for LOH

DTIC Science & Technology

1976-07-01

properties at Station 14.5 are EEA = 2.63 x 106 lb (Figure 7) El yy 3.12 x 106 lb-in. 2 (Figure 8) EEIxx .13 x 106 lb-in. 2 (Figure 9) Cy .52 in. (Figure...3M-1002-1014 "S" glass, f CF E 4250(7.0 x 106) = EEA 3.307 x 106 ft = 8996 psi Ftu = 220,000 psi MSt = Ftu - 220,000 1 ft 8996 MSt = 23.46 Ample 60 2...Beam truss members were centered with respect to the hub and formed seals were positioned at the outboard edges of the hub, Furane Plastics 8615 Uralane

Surveillance perspective on Lyme borreliosis across the European Union and European Economic Area

PubMed Central

van den Wijngaard, Cees C; Hofhuis, Agnetha; Simões, Mariana; Rood, Ente; van Pelt, Wilfrid; Zeller, Herve; Van Bortel, Wim

2017-01-01

Lyme borreliosis (LB) is the most prevalent tick-borne disease in Europe. Erythema migrans (EM), an early, localised skin rash, is its most common presentation. Dissemination of the bacteria can lead to more severe manifestations including skin, neurological, cardiac, musculoskeletal and ocular manifestations. Comparison of LB incidence rates in the European Union (EU)/European Economic Area (EEA) and Balkan countries are difficult in the absence of standardised surveillance and reporting procedures. We explored six surveillance scenarios for LB surveillance in the EU/EEA, based on the following key indicators: (i) erythema migrans, (ii) neuroborreliosis, (iii) all human LB manifestations, (iv) seroprevalence, (v) tick bites, and (vi) infected ticks and reservoir hosts. In our opinion, neuroborreliosis seems most feasible and useful as the standard key indicator, being one of the most frequent severe LB manifestations, with the possibility of a specific case definition. Additional surveillance with erythema migrans as key indicator would add value to the surveillance of neuroborreliosis and lead to a more complete picture of LB epidemiology in the EU/EEA. The other scenarios have less value as a basis for EU-level surveillance, but can be considered periodically and locally, as they could supply complementary insights. PMID:28703098

The endonasal endoscopic harvest and anatomy of the buccal fat pad flap for closure of skull base defects.

PubMed

Markey, Jeff; Benet, Arnau; El-Sayed, Ivan H

2015-10-01

Extirpation via expanded endonasal approaches (EEA) to the skull base can result in defects requiring vascularized rotational flap reconstruction. The buccal fat pad (BFP) is a vascularized graft described in open skull base resections, but its harvest and adequacy of vascular supply have not been examined for use with EEA. A transfacial cadaveric dissection was carried forth in a latex-injected specimen to characterize the BFP blood supply. Then a cadaveric dissection series was performed involving the endoscopic harvest and rotation of 10 buccal fat pads in five cadaveric specimens to assess defect coverage. An endoscopic medial maxillectomy combined with an anterior maxillotomy was performed prior to endoscopic harvest in cadaveric specimens. The BFP was rotated to assess its capability to reconstruct seven possible ventral skull base defects. Finally, the BFP vascular anatomy was further characterized following harvest and transposition. The BFP reconstructed defects at the greater sphenoid wing, inferior and superior clivus, sella, planum, and bilateral ethmoids in all cadaveric specimens. In some cases it covered two sites concurrently. The BFP pedicled rotational flap is a potential alternate flap following EEA in select cases. NA © 2015 The American Laryngological, Rhinological and Otological Society, Inc.

Potomac horse fever.

PubMed

Palmer, J E

1993-08-01

E. risticii, the cause of classic Potomac horse fever, is now known to produce two disease syndromes: EEC and EEA. The pathogen appears to commonly infect horses based on seroepidemiologic studies; however, the method of transmission remains unknown. The most common clinical disease is EEC, commonly called Potomac horse fever, which presents a wide spectrum of clinical signs. Diagnosis is currently dependent on serology, which frequently does not lead to a definitive diagnosis and at best results in a retrospective diagnosis. A new diagnostic approach, polymerase chain reaction, may offer a rapid and accurate diagnostic test but is yet to be proven. Antimicrobials found to be highly effective in treating EEC include intravenous oxytetracycline and the oral combination of erythromycin and rifampin. Without an understanding of the mode of transmission, prevention of EEC will remain impossible. Although two commercial bacterins are available, vaccination does not result in complete protection. EEA is a recently recognized clinical syndrome caused by E. risticii. After recovery from EEC, pregnant mares may abort. Based on histologic examination, the aborted fetus appears to be a victim of an ehrlichial colitis as well as placentitis. The effect of antimicrobials and vaccination on the occurrence of EEA has yet to be investigated.

Is EU/EEA population protected from polio?

PubMed Central

Nijsten, DRE; Carrillo-Santisteve, P; Miglietta, A; Ruitenberg, J; Lopalco, PL

2015-01-01

The WHO European Region has been declared polio-free since 2002. By 2010, inactivated polio vaccine (IPV) was the only polio vaccine in use in the EU/EEA for the primary vaccination of children. A systematic review of the literature on polio seroprevalence studies, complemented by the analysis of available vaccine coverage data, has been carried out with the aim of assessing the level of protection against polio in the European population. A total of 52 studies, with data from 14 out of the 31 EU/EEA countries, were included in the analysis. This systematic review shows that, overall, seroprevalence for PV1 and PV3 is high in most countries, although seroimmunity gaps have been detected in several birth cohorts. In particular, relatively low immunity status was found in some countries for individuals born in the 60's and 70's. Discrepancies between reported vaccination coverage and immunity levels have been also highlighted. Countries should make sure that their population is being vaccinated for polio to reduce the risk of local poliovirus transmission in case of importation. Moreover, assessing immunity status should be priority for those traveling to areas where wild polioviruses are still circulating. PMID:25898095

Is EU/EEA population protected from polio?

PubMed

Nijsten, Dre; Carrillo-Santisteve, P; Miglietta, A; Ruitenberg, J; Lopalco, P L

2015-01-01

The WHO European Region has been declared polio-free since 2002. By 2010, inactivated polio vaccine (IPV) was the only polio vaccine in use in the EU/EEA for the primary vaccination of children. A systematic review of the literature on polio seroprevalence studies, complemented by the analysis of available vaccine coverage data, has been carried out with the aim of assessing the level of protection against polio in the European population. A total of 52 studies, with data from 14 out of the 31 EU/EEA countries, were included in the analysis. This systematic review shows that, overall, seroprevalence for PV1 and PV3 is high in most countries, although seroimmunity gaps have been detected in several birth cohorts. In particular, relatively low immunity status was found in some countries for individuals born in the 60's and 70's. Discrepancies between reported vaccination coverage and immunity levels have been also highlighted. Countries should make sure that their population is being vaccinated for polio to reduce the risk of local poliovirus transmission in case of importation. Moreover, assessing immunity status should be priority for those traveling to areas where wild polioviruses are still circulating.

Variations in the sales and sales patterns of veterinary antimicrobial agents in 25 European countries.

PubMed

Grave, Kari; Torren-Edo, Jordi; Muller, Arno; Greko, Christina; Moulin, Gerard; Mackay, David

2014-08-01

To describe sales and sales patterns of veterinary antimicrobial agents in 25 European Union (EU)/European Economic Area (EEA) countries for 2011. Data on the sales of veterinary antimicrobial agents from 25 EU member states and EEA countries for 2011 were collected at package level (name, formulation, strength, pack size, number of packages sold) according to a standardized protocol and template and presented in a harmonized manner. These data were calculated to express amounts sold, in metric tonnes, of active ingredient of each package. A population correction unit (PCU) was applied as a proxy for the animal biomass potentially treated with antimicrobial agents. The indicator used to express sales was milligrams of active substance per PCU. Substantial variations in the sales patterns and in the magnitude of sales of veterinary antimicrobial agents, expressed as mg/PCU, between the countries were observed. The proportion of sales, in mg/PCU, of products applicable for treatment of groups or herds of animals (premixes, oral powders and oral solution) varied considerably between the countries. Some countries reported much lower sales of veterinary antimicrobial agents than others, when expressed as mg/PCU. Sales patterns varied between countries, particularly with respect to pharmaceutical forms. Further studies are needed to understand the factors that explain the observed differences. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Energy Engineering Analysis (EEA) program for Red River Army Depot, Texas. Executive summary. Final report

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

NONE

The objectives of this Energy Engineering Analysis (EEA) for RRAD were three fold: Develop a systematic plan of projects which will result in reducing energy consumption. Consider renewable energy sources with the objective of establishing an orderly procedure for reducing use of non-renewable energy sources. Determine the feasibility of Total Energy (TE), Selective Energy (SE), and Central Heating Plant (CHP) concepts using alternative fuels. In essence, an assessment of the entire energy picture at RRAD was undertaken. This report is a summary of that effort. RRAD was originally built during 1941 and 1942 as a reserve ordnance depot for themore » Army. Since then, the missions and activities at RRAD have broadened to the point that it is now one of the largest Army depots in the continental United States. Located just west of Texarkana, Texas, RRAD encompasses an area of approximately 19,886 acres. The primary missions at RRAD are general supply and maintenance of vital Army equipment and ordnance material. The supply activities constitute the stocking, distribution, storage and supply of general Army supplies for the central region of the United States. Supply of vehicles, ammunition and guided missiles from RRAD encompasses a much larger region. The maintenance and repair functions at RRAD consists of the overhaul, modification, conversion and repair of automotive equipment and combat vehicles, missile systems and components, armament, and ammunition. Approximately 5500 people are currently employed at RRAD to carry out its mission.« less

Surveillance perspective on Lyme borreliosis across the European Union and European Economic Area.

PubMed

van den Wijngaard, Cees C; Hofhuis, Agnetha; Simões, Mariana; Rood, Ente; van Pelt, Wilfrid; Zeller, Herve; Van Bortel, Wim

2017-07-06

Lyme borreliosis (LB) is the most prevalent tick-borne disease in Europe. Erythema migrans (EM), an early, localised skin rash, is its most common presentation. Dissemination of the bacteria can lead to more severe manifestations including skin, neurological, cardiac, musculoskeletal and ocular manifestations. Comparison of LB incidence rates in the European Union (EU)/European Economic Area (EEA) and Balkan countries are difficult in the absence of standardised surveillance and reporting procedures. We explored six surveillance scenarios for LB surveillance in the EU/EEA, based on the following key indicators: (i) erythema migrans, (ii) neuroborreliosis, (iii) all human LB manifestations, (iv) seroprevalence, (v) tick bites, and (vi) infected ticks and reservoir hosts. In our opinion, neuroborreliosis seems most feasible and useful as the standard key indicator, being one of the most frequent severe LB manifestations, with the possibility of a specific case definition. Additional surveillance with erythema migrans as key indicator would add value to the surveillance of neuroborreliosis and lead to a more complete picture of LB epidemiology in the EU/EEA. The other scenarios have less value as a basis for EU-level surveillance, but can be considered periodically and locally, as they could supply complementary insights. This article is copyright of The Authors, 2017.

Assessment of HIV molecular surveillance capacity in the European Union, 2016.

PubMed

Keating, Patrick; Pharris, Anastasia; Leitmeyer, Katrin; De Angelis, Stefania; Wensing, Annemarie; Amato-Gauci, Andrew J; Broberg, Eeva

2017-12-01

IntroductionExpanding access to HIV antiretroviral treatment is expected to decrease HIV incidence and acquired immunodeficiency syndrome (AIDS) mortality. However, this may also result in increased HIV drug resistance (DR). Better monitoring and surveillance of HIV DR is required to inform treatment regimens and maintain the long term effectiveness of antiretroviral drugs. As there is currently no formal European Union (EU)-wide collection of HIV DR data, this study aimed to assess the current HIV molecular surveillance capacity in EU/European Economic Area (EEA) countries in order to inform the planning of HIV DR monitoring at EU level. Methods: Thirty EU/EEA countries were invited to participate in a survey on HIV molecular surveillance capacity, which also included laboratory aspects. Results: Among 21 responding countries, 13 reported using HIV sequence data (subtype and/or DR) for surveillance purposes at national level. Of those, nine stated that clinical, epidemiological and sequence data were routinely linked for analysis. Discussion/conclusion : We identified similarities between existing HIV molecular surveillance systems, but also found important challenges including human resources, data ownership and legal issues that would need to be addressed.Information on capacities should allow better planning of the phased introduction of HIV DR surveillance at EU/EEA level.

Deglacial Tropical Atlantic subsurface warming links ocean circulation variability to the West African Monsoon.

PubMed

Schmidt, Matthew W; Chang, Ping; Parker, Andrew O; Ji, Link; He, Feng

2017-11-13

Multiple lines of evidence show that cold stadials in the North Atlantic were accompanied by both reductions in Atlantic Meridional Overturning Circulation (AMOC) and collapses of the West African Monsoon (WAM). Although records of terrestrial change identify abrupt WAM variability across the deglaciation, few studies show how ocean temperatures evolved across the deglaciation. To identify the mechanism linking AMOC to the WAM, we generated a new record of subsurface temperature variability over the last 21 kyr based on Mg/Ca ratios in a sub-thermocline dwelling planktonic foraminifera in an Eastern Equatorial Atlantic (EEA) sediment core from the Niger Delta. Our subsurface temperature record shows abrupt subsurface warming during both the Younger Dryas (YD) and Heinrich Event 1. We also conducted a new transient coupled ocean-atmosphere model simulation across the YD that better resolves the western boundary current dynamics and find a strong negative correlation between AMOC strength and EEA subsurface temperatures caused by changes in ocean circulation and rainfall responses that are consistent with the observed WAM change. Our combined proxy and modeling results provide the first evidence that an oceanic teleconnection between AMOC strength and subsurface temperature in the EEA impacted the intensity of the WAM on millennial time scales.

Screening for Infectious Diseases among Newly Arrived Migrants in EU/EEA Countries—Varying Practices but Consensus on the Utility of Screening

PubMed Central

Kärki, Tommi; Napoli, Christian; Riccardo, Flavia; Fabiani, Massimo; Dente, Maria Grazia; Carballo, Manuel; Noori, Teymur; Declich, Silvia

2014-01-01

Screening is one possible tool for monitoring infectious diseases among migrants. However, there is limited information on screening programmes targeted for newly arrived migrants in EU/EEA countries. Our aim was to investigate the implementation, practices and usefulness of these programmes. We conducted a survey among country experts from EU/EEA countries and Switzerland, asking whether their countries had implemented screening programmes. We also estimated the association between the implementation of these programmes and the rate of asylum-seekers in the population. Of the countries, 16 (59%) had implemented screening programmes and 15 (56%) had national guidelines. The rate of asylum-seekers was associated with implementation of screening programmes (p = 0.014). Screening was performed most often for tuberculosis; most commonly on holding level, and was targeted to specific migrant groups in over half of the countries performing screening. Twenty-five of all the country experts (96%) considered screening among migrants useful, and 24 (92%) would welcome EU level guidelines for screening. The implementation of screening programmes varied, and the practices were different among countries. Our survey suggests, that establishing EU level guidelines for screening would be useful, although they would have to take into account differences between individual countries. PMID:25337945

The combined endonasal and transoral approach for the management of skull base and nasopharyngeal pathology: a case series.

PubMed

Sreenath, Satyan B; Rawal, Rounak B; Zanation, Adam M

2014-01-01

The posterior skull base and the nasopharynx have historically represented technically difficult regions to approach surgically given their central anatomical locations. Through continued improvements in endoscopic instrumentation and technology, the expanded endonasal approach (EEA) has introduced a new array of surgical options in the management of pathology involving these anatomically complex areas. Similarly, the transoral robotic surgical (TORS) approach was introduced as a minimally invasive surgical option to approach tongue base, nasopharyngeal, parapharyngeal, and laryngeal lesions. Although both the EEA and the TORS approach have been extensively described as viable surgical options in managing nasopharyngeal and centrally located head and neck pathology, both endonasal and transoral techniques have inherent limitations. Given these limitations, several institutions have published feasibility studies with the combined EEA and TORS approaches for a variety of skull base and nasopharyngeal pathologies. In this article, the authors present their clinical experience with the combined endonasal and transoral approach through a case series presentation, and discuss advantages and limitations of this approach for surgical management of the middle and posterior skull base and nasopharynx. In addition, a presentation is included of a unique, simultaneous endonasal and transoral dissection of the nasopharynx through an innovative intraoperative setup.

Clival chordomas: considerations after 16 years of endoscopic endonasal surgery.

PubMed

Zoli, Matteo; Milanese, Laura; Bonfatti, Rocco; Faustini-Fustini, Marco; Marucci, Gianluca; Tallini, Giovanni; Zenesini, Corrado; Sturiale, Carmelo; Frank, Giorgio; Pasquini, Ernesto; Mazzatenta, Diego

2018-02-01

OBJECTIVE In the past decade, the role of the endoscopic endonasal approach (EEA) has relevantly evolved for skull base tumors. In this study, the authors review their surgical experience with using an EEA in the treatment of clival chordomas, which are deep and infiltrative skull base lesions, and they highlight the advantages and limitations of this ventral approach. METHODS All consecutive cases of chordoma treated with an EEA between 1998 and 2015 at a single institution are included in this study. Preoperative assessment consisted of neuroimaging (MRI and CT with angiography sequences) and endocrinological, neurological, and ophthalmological evaluations, which were repeated 3 months after surgery and annually thereafter. Postoperative adjuvant therapies were considered. RESULTS Sixty-five patients (male/female ratio 1:0.9) were included in this study. The median age was 48 years (range 9-80 years). Gross-total resection (GTR) was achieved in 47 cases (58.7%). On univariate analysis, primary procedures (p = 0.001), location in the superior or middle third of the clivus (p = 0.043), extradural location (p = 0.035), and histology of conventional chordomas (p = 0.013) were associated with a higher rate of GTR. The complication rate was 15.1%, and there were no perioperative deaths. Most complications did not result in permanent sequelae and included 2 CSF leaks (2.5%), 5 transient cranial nerve VI palsies (6.2%), and 2 internal carotid artery injuries (2.5%), which were treated with coil occlusion of the internal carotid artery without neurological deficits. Three patients (3.8%) presented with complications resulting in permanent neurological deficits due to a postoperative hematoma (1.2%) causing a hemiparesis, and 2 permanent ophthalmoplegias (2.5%). Seventeen patients (26.2%) have died of tumor progression over the course of follow-up (median 52 months, range 7-159 months). Based on Kaplan-Meier analysis, the survival rate was 77% at 5 years and 57% at 10 years. On multivariate analysis, the extent of tumor removal (p = 0.001) and the absence of previous treatments (p = 0.001) proved to be correlated with a longer survival rate. CONCLUSIONS The EEA was associated with a high rate of tumor removal and symptom control, with low morbidity and preservation of a good quality of life. These results allow for a satisfactory overall survival rate, particularly after GTR and for primary surgery. Considering these results, the authors believe that an EEA can be a helpful tool in chordoma surgery, achieving a good balance between as much tumor removal as possible and the preservation of an acceptable patient quality of life.

Cleveland Harbor, Ohio. Section 3. Study. Termination Report.

DTIC Science & Technology

1983-04-01

dot’ t-I p ModC’ 00 b Y^AIefs V S~jx7 4 rI e I2L wt 9?,j 100 y~Ii eea -Ss c sww.../5v- rLI r /. d V 7 c P...pJAf~ 1 ,, Alr~’ Wd4b IS~4 (0s~e A0 (4- . 4", . 1w ",o 6/4 c. ~* h ’.JS 4E. /0, 0,C.4 6-sr-- t9CeA. /11 x to f. C-,. o iv, jt, Is~T~- eea ~r H - tIT...of the clamshell grab were placed in a clean, plastic bucket. After thorough homogenization, approximately one quart of material from each ;ite was

Eating when depressed, anxious, bored, or happy: Are emotional eating types associated with unique psychological and physical health correlates?

PubMed

Braden, Abby; Musher-Eizenman, Dara; Watford, Tanya; Emley, Elizabeth

2018-06-01

The majority of research on emotional eating has examined general emotional eating, to the exclusion of more distinct emotions such as boredom and positive emotions. The current study aimed to examine whether specific types of emotional eating (i.e., eating in response to depression (EE-D), anxiety/anger (EE-A), boredom (EE-B), and positive emotions (EE-P)) were related to a range of psychological (i.e., global psychological well-being, eating disorder symptoms, emotion regulation) and physical health variables. A sample of adults (n = 189) with overweight/obesity were recruited via Amazon Mechanical Turk. Participants self-reported height and weight and completed a battery of questionnaires. Correlational analyses showed that more frequent EE-D, EE-A, and EE-B were related to poorer psychological well-being, greater eating disorder symptoms, and more difficulties with emotion regulation. EE-P was not significantly related to outcome variables. In regression analyses, eating in response to depression (EE-D) was the type of emotional eating most closely related to psychological well-being, eating disorder symptoms, and emotion regulation difficulties. Exploratory analyses revealed associations between EE-D, EE-A, and EE-B and facets of emotion regulation and specific disordered eating symptoms. Findings suggest that unique patterns exist between specific types of emotional eating and psychological outcomes. Copyright © 2018 Elsevier Ltd. All rights reserved.

Determination of Rab5 activity in the cell by effector pull-down assay.

PubMed

Qi, Yaoyao; Liang, Zhimin; Wang, Zonghua; Lu, Guodong; Li, Guangpu

2015-01-01

Rab5 targets to early endosomes and is a master regulator of early endosome fusion and endocytosis in all eukaryotic cells. Like other GTPases, Rab5 functions as a molecular switch by alternating between GTP-bound and GDP-bound forms, with the former being biologically active via interactions with multiple effector proteins. Thus the Rab5-GTP level in the cell reflects Rab5 activity in promoting endosome fusion and endocytosis and is indicative of cellular endocytic activity. In this chapter, we describe a Rab5 activity assay by using GST fusion proteins with the Rab5 effectors such as Rabaptin-5, Rabenosyn-5, and EEA1 that specifically bind to GTP-bound Rab5. We compare the efficiencies of the three GST fusion proteins in the pull-down of mammalian and fungal Rab5 proteins.

[Application of extended exergy method in driving mechanism and efficiency of regional eco-economy].

PubMed

Fan, Xin Gang; Mi, Wen Bao; Hou, Jing Wei

2017-01-01

To analyze social-economic causes of the regional ecological degradation, and avoid such problems as the complex circulation network and difficulty to identify laws caused by extended exergy analysis (EEA) previously applied at the national scale, this paper reduced spatial scale to the county scale and took Pengyang County in Ningxia as an example. Eco-economic system in Peng-yang County was divided into seven interrelated sectors. The exergy value of circulations in the eco-economic system including materials, labor and capital were calculated respectively to analyze the extended exergy characteristics of the driving sectors, factors and paths and evaluate their ecological efficiency. The results showed that agriculture and households were the main driving sectors of the eco-economic system in Pengyang County. The average exergy value of 31 flow paths among the sectors was 0.80 PJ. There were only 8 flow paths whose exergy values were higher than the average value. Eco-economic system in Pengyang County development was driven by two continuous flow paths, labor output of the households sector and demands of the households sector supported by other sectors. The mineral resources were massively exploited, and then directly exported to the outside, which could not promote the local development from the inside, but, on the contrary, increase the ecological environment pressure because of the over-exploitation. The eco-efficiency of Pengyang County in 2014 was 68.1%, almost equivalent to the by-level of the national scale at home and abroad ten years ago, mainly because of the lower eco-efficiencies of the service sector and households sector. EEA had the advantage of networking and structuring, could specify the sectors, factors and driven paths, and break through the bottleneck of driving mechanism research of the eco-economic system. EEA had certain adaptability to explore the operational principle and optimal pattern of the regional eco-economic system. Compared with the national scale, EEA at the regional scale could more easily identify the driving mechanism of eco-economic system, and could clearly guide the regional administrative department to reduce the ecological environment pressure.

[Esophagojejunal anastomoses with the EEA stapler after total gastrectomy. Technic and immediate results apropos of a recent series of 32 cases].

PubMed

Le Treut, Y P; Delpero, J R; Sabiani, P; Berthet, B; Bozon-Verduraz, E; Pol, B; Bricot, R

1987-12-01

32 consecutive total gastrectomies for cancer (20 extended total gastrectomies) were carried out through an abdominal approach. Reconstruction using a Rouxen-Y loop (28 cases) or an interposed jejunal loop (four cases) was done with an EEA stapled oesophago jejunostomy: 25 end to side and 5 end to end anastomosis were performed and two technical failures led to complete manual anastomosis. All patients have had post-operative X-ray examination: there was no anastomotic leakage in this series, even in the 3 patients (10%) who have died during the post-operative course. A critical study of this series and eight others one from elsewhere is presented: it is concluded that the stapling device allows an easier and safer oesophagojejunostomy than manual procedure, when great care is taken of technical details.

Variation in adult vaccination policies across Europe: an overview from VENICE network on vaccine recommendations, funding and coverage.

PubMed

Kanitz, Elisabeth E; Wu, Lauren A; Giambi, Cristina; Strikas, Raymond A; Levy-Bruhl, Daniel; Stefanoff, Pawel; Mereckiene, Jolita; Appelgren, Eva; D'Ancona, Fortunato

2012-07-27

In 2010-2011, in the framework of the VENICE project, we surveyed European Union (EU) and Economic Area (EEA) countries to fill the gap of information regarding vaccination policies in adults. This project was carried out in collaboration with the United States National Vaccine Program Office, who conducted a similar survey in all developed countries. VENICE representatives of all 29 EU/EEA-countries received an online questionnaire including vaccination schedule, recommendations, funding and coverage in adults for 17 vaccine-preventable diseases. The response rate was 100%. The definition of age threshold for adulthood for the purpose of vaccination ranged from 15 to 19 years (median=18 years). EU/EEA-countries recommend between 4 and 16 vaccines for adults (median=11 vaccines). Tetanus and diphtheria vaccines are recommended to all adults in 22 and 21 countries respectively. The other vaccines are mostly recommended to specific risk groups; recommendations for seasonal influenza and hepatitis B exist in all surveyed countries. Six countries have a comprehensive summary document or schedule describing all vaccines which are recommended for adults. None of the surveyed countries was able to provide coverage estimates for all the recommended adult vaccines. Vaccination policies for adults are not consistent across Europe, including the meaning of "recommended vaccine" which is not comparable among countries. Coverage data for adults should be collected routinely like for children vaccination. Copyright © 2012 Elsevier Ltd. All rights reserved.

Dissolved Organic Carbon Degradation in Response to Nutrient Amendments in Southwest Greenland Lakes

NASA Astrophysics Data System (ADS)

Burpee, B. T.; Northington, R.; Simon, K. S.; Saros, J. E.

2014-12-01

Aquatic ecosystems across the Arctic are currently experiencing rapid shifts in biotic, chemical, and physical factors in response to climate change. Preliminary data from multiple lakes in southwestern Greenland indicate decreasing dissolved organic carbon (DOC) concentrations over the past decade. Though several factors may be contributing to this phenomenon, this study attempts to elucidate the potential of heterotrophic bacteria to degrade DOC in the presence of increasing nutrient concentrations. In certain Arctic regions, nutrient subsidies have been released into lakes due to permafrost thaw. If this is occurring in southwestern Greenland, we hypothesized that increased nutrient concentrations will relieve nutrient limitation, thereby allowing heterotrophic bacteria to utilize DOC as an energy source. This prediction was tested using experimental DOC degradation assays from four sample lakes. Four nutrient amendment treatments (control, N, P, and N + P) were used to simulate in situ subsidies. Five time points were sampled during the incubation: days 0, 3, 6, 14, and 60. Total organic carbon (TOC) and parallel factor (PARAFAC) analysis were used to monitor the relative concentrations of different DOC fractions over time. In addition, samples for extracellular enzyme activity (EEA) analysis were collected at every time point. Early analysis of fulvic and humic pools of DOC do not indicate any significant change from days 0 to 14. This could be due to the fact that these DOC fractions are relatively recalcitrant. This study will be important in determining whether bacterial degradation could be a contributing factor to DOC decline in arctic lakes.

Selection of interest and inflation rates for infrastructure investment analyses.

DOT National Transportation Integrated Search

2014-12-01

The South Dakota Department of Transportation (SDDOT) uses engineering economic analyses (EEA) to : support planning, design, and construction decision-making such as project programming and planning, : pavement type selection, and the occasional val...

Changes of lysosomes in the earliest stages of the development of atherosclerosis.

PubMed

Bobryshev, Yuri V; Shchelkunova, Tatyana A; Morozov, Ivan A; Rubtsov, Petr M; Sobenin, Igor A; Orekhov, Alexander N; Smirnov, Alexander N

2013-05-01

One of hypotheses of atherosclerosis is based on a presumption that the zones prone to the development of atherosclerosis contain lysosomes which are characterized by enzyme deficiency and thus, are unable to dispose of lipoproteins. The present study was undertaken to investigate the characteristics and changes of lysosomes in the earliest stages of the development of atherosclerosis. Electron microscopic immunocytochemistry revealed that there were certain changes in the distribution of CD68 antigen in lysosomes along the 'normal intima-initial lesion-fatty streak' sequence. There were no significant changes found in the key mRNAs encoding for the components of endosome/lysosome compartment in initial atherosclerotic lesions, but in fatty streaks, the contents of EEA1 and Rab5a mRNAs were found to be diminished while the contents of CD68 and p62 mRNAs were increased, compared with the intact tissue. The study reinforces a view that changes occurring in lysosomes play a role in atherogenesis from the very earlier stages of the disease. © 2013 The Authors. Published by Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

Identification of effective exciton-exciton annihilation in squaraine-squaraine copolymers.

PubMed

Hader, Kilian; May, Volkhard; Lambert, Christoph; Engel, Volker

2016-05-11

Ultrafast time-resolved transient absorption spectroscopy is able to monitor the fate of the excited state population in molecular aggregates or polymers. Due to many competing decay processes, the identification of exciton-exciton annihilation (EEA) is difficult. Here, we use a microscopic model to describe exciton annihilation processes in squaraine-squaraine copolymers. Transient absorption time traces measured at different laser powers exhibit an unusual time-dependence. The analysis points towards dynamics taking place on three time-scales. Immediately after laser-excitation a localization of excitons takes place within the femtosecond time-regime. This is followed by exciton-exciton annihilation which is responsible for a fast decay of the exciton population. At later times, excitations being localized on units which are not directly connected remain so that diffusion dominates the dynamics and leads to a slower decay. We thus provide evidence for EEA tracked by time-resolved spectroscopy which has not been reported that clearly before.

A Role for EHD4 in the Regulation of Early Endosomal Transport

PubMed Central

Sharma, Mahak; Naslavsky, Naava; Caplan, Steve

2009-01-01

All four of the C-terminal Eps15 homology domain (EHD) proteins have been implicated in the regulation of endocytic trafficking. However, the high level of amino acid sequence identity among these proteins has made it challenging to elucidate the precise function of individual EHD proteins. We demonstrate here with specific peptide antibodies that endogenous EHD4 localizes to Rab5-, early embryonic antigen 1 (EEA1)- and Arf6-containing endosomes and colocalizes with internalized transferrin in the cell periphery. Knock-down of EHD4 expression by both small interfering RNA and short hairpin RNA leads to the generation of enlarged early endosomal structures that contain Rab5 and EEA1 as well as internalized transferrin or major histocompatibility complex class I molecules. In addition, cargo destined for degradation, such as internalized low-density lipoprotein, also accumulates in the enlarged early endosomes in EHD4-depleted cells. Moreover, we have demonstrated that these enlarged early endosomes are enriched in levels of the activated GTP-bound Rab5. Finally, we show that endogenous EHD4 and EHD1 interact in cells, suggesting coordinated involvement in the regulation of receptor transport along the early endosome to endocytic recycling compartment axis. The results presented herein provide evidence that EHD4 is involved in the control of trafficking at the early endosome and regulates exit of cargo toward both the recycling compartment and the late endocytic pathway. PMID:18331452

Solo/Trio8, a membrane-associated short isoform of Trio, modulates endosome dynamics and neurite elongation.

PubMed

Sun, Ying-Jie; Nishikawa, Kaori; Yuda, Hideki; Wang, Yu-Lai; Osaka, Hitoshi; Fukazawa, Nobuna; Naito, Akira; Kudo, Yoshihisa; Wada, Keiji; Aoki, Shunsuke

2006-09-01

With DNA microarrays, we identified a gene, termed Solo, that is downregulated in the cerebellum of Purkinje cell degeneration mutant mice. Solo is a mouse homologue of rat Trio8-one of multiple Trio isoforms recently identified in rat brain. Solo/Trio8 contains N-terminal sec14-like and spectrin-like repeat domains followed by a single guanine nucleotide exchange factor 1 (GEF1) domain, but it lacks the C-terminal GEF2, immunoglobulin-like, and kinase domains that are typical of Trio. Solo/Trio8 is predominantly expressed in Purkinje neurons of the mouse brain, and expression begins following birth and increases during Purkinje neuron maturation. We identified a novel C-terminal membrane-anchoring domain in Solo/Trio8 that is required for enhanced green fluorescent protein-Solo/Trio8 localization to early endosomes (positive for both early-endosome antigen 1 [EEA1] and Rab5) in COS-7 cells and primary cultured neurons. Solo/Trio8 overexpression in COS-7 cells augmented the EEA1-positive early-endosome pool, and this effect was abolished via mutation and inactivation of the GEF domain or deletion of the C-terminal membrane-anchoring domain. Moreover, primary cultured neurons transfected with Solo/Trio8 showed increased neurite elongation that was dependent on these domains. These results suggest that Solo/Trio8 acts as an early-endosome-specific upstream activator of Rho family GTPases for neurite elongation of developing Purkinje neurons.

Good practices and challenges in addressing poliomyelitis and measles in the European Union.

PubMed

Kinsman, John; Stöven, Svenja; Elgh, Fredrik; Murillo, Pilar; Sulzner, Michael

2018-04-06

All European Union (EU) and European Economic Area (EEA) Member States have pledged to ensure political commitment towards sustaining the region's poliomyelitis-free status and eliminating measles. However, there remain significant gaps between policy and practice in many countries. This article reports on an assessment conducted for the European Commission that aimed to support improvements in preparedness and response to poliomyelitis and measles in Europe. A documentary review was complemented by qualitative interviews with professionals working in International and EU agencies, and in at-risk or recently affected EU/EEA Member States (six each for poliomyelitis and measles). Twenty-six interviews were conducted on poliomyelitis and 24 on measles; the data were subjected to thematic analysis. Preliminary findings were then discussed at a Consensus Workshop with 22 of the interviewees and eight other experts. Generic or disease-specific plans exist in the participating countries and cross-border communications during outbreaks were generally reported as satisfactory. However, surveillance systems are of uneven quality, and clinical expertise for the two diseases is limited by a lack of experience. Serious breaches of protocol have recently been reported from companies producing poliomyelitis vaccines, and vaccine coverage rates for both diseases were also sub-optimal. A set of suggested good practices to address these and other challenges is presented. Poliomyelitis and measles should be brought fully onto the policy agendas of all EU/EEA Member States, and adequate resources provided to address them. Each country must abide by the relevant commitments that they have already made.

Extended Endoscopic Endonasal Approach for Craniopharyngioma Removal.

PubMed

Messerer, Mahmoud; Maduri, Rodolfo; Daniel, Roy Thomas

2018-02-01

Objective  Endoscopic transsphenoidal extended endoscopic approach (EEA) represents a valid alternative to microsurgery for craniopharyngiomas removal, especially for retrochiasmatic lesions without large parasellar extension. The present video illustrates the salient surgical steps of the EEA for craniopahryngioma removal. Patient  A 52-year-old man presented with a bitemporal hemianopia and a bilateral decreased visual acuity. MRI showed a Kassam type III cystic craniopharyngioma with a solid component ( Fig. 1 , panels A and B). Surgical Procedure  The head is rotated 10 degrees toward the surgeons. The nasal step is started through the left nostril with a middle turbinectomy. A nasoseptal flap is harvested and positioned in the left choana. The binostril approach allows a large sphenoidotomy to expose the key anatomic landmarks. The craniotomy boundaries are the planum sphenoidale superiorly, the median opticocarotid recesses, the internal carotid artery laterally and the clival recess inferiorly. After dural opening and superior intercavernous sinus coagulation, the tumor is entirely removed ( Fig. 2 , panels A and B). Skull base reconstruction is ensured by fascia lata grafting and nasoseptal flap positioning. Results  Postoperative MRI showed the complete tumor resection ( Fig. 1 , panels C and D). At 3 months postoperatively, the bitemporal hemianopia regressed and the visual acuity improved. A novel left homonymous hemianopia developed secondary to optic tract manipulation. Conclusions  The extended EEA is a valid surgical approach for craniopharyngioma resection. A comprehensive knowledge of the sellar and parasellar anatomy is mandatory for safe tumor removal with decreased morbidity and satisfactory oncologic results. The link to the video can be found at: https://youtu.be/NrCPPnVK2qA .

Immunisation Information Systems – useful tools for monitoring vaccination programmes in EU/EEA countries, 2016

PubMed Central

Derrough, Tarik; Olsson, Kate; Gianfredi, Vincenza; Simondon, Francois; Heijbel, Harald; Danielsson, Niklas; Kramarz, Piotr; Pastore-Celentano, Lucia

2017-01-01

Immunisation Information Systems (IIS) are computerised confidential population based-systems containing individual-level information on vaccines received in a given area. They benefit individuals directly by ensuring vaccination according to the schedule and they provide information to vaccine providers and public health authorities responsible for the delivery and monitoring of an immunisation programme. In 2016, the European Centre for Disease Prevention and Control (ECDC) conducted a survey on the level of implementation and functionalities of IIS in 30 European Union/European Economic Area (EU/EEA) countries. It explored the governance and financial support for the systems, IIS software, system characteristics in terms of population, identification of immunisation recipients, vaccinations received, and integration with other health record systems, the use of the systems for surveillance and programme management as well as the challenges involved with implementation. The survey was answered by 27 of the 30 EU/EEA countries having either a system in production at national or subnational levels (n = 16), or being piloted (n = 5) or with plans for setting up a system in the future (n = 6). The results demonstrate the added-value of IIS in a number of areas of vaccination programme monitoring such as monitoring vaccine coverage at local geographical levels, linking individual immunisation history with health outcome data for safety investigations, monitoring vaccine effectiveness and failures and as an educational tool for both vaccine providers and vaccine recipients. IIS represent a significant way forward for life-long vaccination programme monitoring. PMID:28488999

Immunisation Information Systems - useful tools for monitoring vaccination programmes in EU/EEA countries, 2016.

PubMed

Derrough, Tarik; Olsson, Kate; Gianfredi, Vincenza; Simondon, Francois; Heijbel, Harald; Danielsson, Niklas; Kramarz, Piotr; Pastore-Celentano, Lucia

2017-04-27

Immunisation Information Systems (IIS) are computerised confidential population based-systems containing individual-level information on vaccines received in a given area. They benefit individuals directly by ensuring vaccination according to the schedule and they provide information to vaccine providers and public health authorities responsible for the delivery and monitoring of an immunisation programme. In 2016, the European Centre for Disease Prevention and Control (ECDC) conducted a survey on the level of implementation and functionalities of IIS in 30 European Union/European Economic Area (EU/EEA) countries. It explored the governance and financial support for the systems, IIS software, system characteristics in terms of population, identification of immunisation recipients, vaccinations received, and integration with other health record systems, the use of the systems for surveillance and programme management as well as the challenges involved with implementation. The survey was answered by 27 of the 30 EU/EEA countries having either a system in production at national or subnational levels (n = 16), or being piloted (n = 5) or with plans for setting up a system in the future (n = 6). The results demonstrate the added-value of IIS in a number of areas of vaccination programme monitoring such as monitoring vaccine coverage at local geographical levels, linking individual immunisation history with health outcome data for safety investigations, monitoring vaccine effectiveness and failures and as an educational tool for both vaccine providers and vaccine recipients. IIS represent a significant way forward for life-long vaccination programme monitoring. This article is copyright of The Authors, 2017.

Visualization of self-delivering hydrophobically modified siRNA cellular internalization

PubMed Central

Ly, Socheata; Navaroli, Deanna M.; Didiot, Marie-Cécile; Cardia, James; Pandarinathan, Lakshmipathi; Alterman, Julia F.; Fogarty, Kevin; Standley, Clive; Lifshitz, Lawrence M.; Bellve, Karl D.; Prot, Matthieu; Echeverria, Dimas; Corvera, Silvia; Khvorova, Anastasia

2017-01-01

siRNAs are a new class of therapeutic modalities with promising clinical efficacy that requires modification or formulation for delivery to the tissue and cell of interest. Conjugation of siRNAs to lipophilic groups supports efficient cellular uptake by a mechanism that is not well characterized. Here we study the mechanism of internalization of asymmetric, chemically stabilized, cholesterol-modified siRNAs (sd-rxRNAs®) that efficiently enter cells and tissues without the need for formulation. We demonstrate that uptake is rapid with significant membrane association within minutes of exposure followed by the formation of vesicular structures and internalization. Furthermore, sd-rxRNAs are internalized by a specific class of early endosomes and show preferential association with epidermal growth factor (EGF) but not transferrin (Tf) trafficking pathways as shown by live cell TIRF and structured illumination microscopy (SIM). In fixed cells, we observe ∼25% of sd-rxRNA co-localizing with EGF and <5% with Tf, which is indicative of selective endosomal sorting. Likewise, preferential sd-rxRNA co-localization was demonstrated with EEA1 but not RBSN-containing endosomes, consistent with preferential EGF-like trafficking through EEA1-containing endosomes. sd-rxRNA cellular uptake is a two-step process, with rapid membrane association followed by internalization through a selective, saturable subset of the endocytic process. However, the mechanistic role of EEA1 is not yet known. This method of visualization can be used to better understand the kinetics and mechanisms of hydrophobic siRNA cellular uptake and will assist in further optimization of these types of compounds for therapeutic intervention. PMID:27899655

Plea to lower English test pass marks for EEA nurses.

PubMed

Longhurst, Chris

2017-08-09

Nurses from a group campaigning for the rights of EU citizens in the UK after Brexit are meeting the Nursing and Midwifery Council (NMC) to press for the pass mark for English language tests to be lowered.

Oral healthcare systems in the extended European union.

PubMed

Widström, Eeva; Eaton, Kenneth A

2004-01-01

This article reports a survey of the systems for the provision of oral healthcare in the 28 member and accession states of the EU/EEA in 2003. Descriptions of the systems were collected from the principal dental advisers to governments in the individual states. In many states these were the Chief Dental Officers (CDOs). In states without a CDO, descriptions were gathered from CDO equivalents or senior academics. A template (model description) was used to guide all respondents. Additional statistical information on oral healthcare costs and workforce was collected from the Council of European Chief Dental Officers, WHO and World Bank websites. The study showed that in broad terms there were six patterns (Beveridgian, Bismarkian, The Eastern European (in transition), Nordic, Southern European and Hybrid) for the administration and financing of oral healthcare in the expanding EU. The extent and nature of government involvement in planning and coordinating oral healthcare services and the numbers and pay of the oral healthcare workforce varied between the different models. The biggest recent changes in European oral healthcare were found to have occurred in Eastern Europe, where there has been wide scale privatization of the previously public dental services. However, most of the EU accession (Eastern European) states seemed to be slowly developing insurance systems to cover oral health treatment costs. In the existing EU/EEA, the public dental services such as those in the Nordic countries still have strong political support and some expansion has occurred. In Southern Europe public dental services seemed to have gained some acceptance for the treatment of children and special needs groups. In UK, which has a unique public dental service system, there are plans to make big changes in the delivery, commissioning and remuneration of dental services in the near future. Some EU member states which operate the Bismarkian system with health insurances offering wide population coverage, comprehensive treatment and benefits connected with frequent dental visits, were reported to be experiencing financial problems. The study also indicated that at present, with the exception of Portugal and Spain, where there is dynamic growth in the numbers of dentists, the overall size of the EU/EEA oral health workforce is expanding fairly slowly. Only a minority of member states appeared to collect data on uptake of services and care costs and there were great difficulties in assessing outcomes of care. The data on costs appeared to show wide variations from member state to member state in per capita spending on oral healthcare. In the majority of states, however, costs, especially those in the private sector, could only be estimated. Nevertheless, at a 'macro' level, the study indicated that, in 2000, the 28 member and accession states of the EU/EEA had a total population of 456 million and an oral health workforce of 900,000 (some 300,000 of whom were dentists) and that the cost of oral healthcare was about EUR 54,000,000,000. The study showed wide variations in oral healthcare provision systems between EU/EEA member and accession states and no evidence of harmonization in the past.

77 FR 42482 - Submission for OMB Review; Comment Request

Federal Register 2010, 2011, 2012, 2013, 2014

2012-07-19

... Public Works and Economic Development program, as well as its Economic Adjustment Assistance (EEA) program. Public Works and Economic Development Program investments help support the construction or rehabilitation of essential public infrastructure and facilities necessary to generate or retain private sector...

Down-modulation of the G-protein-coupled Estrogen Receptor, GPER, from the Cell Surface Occurs via a trans-Golgi-Proteasome Pathway*

PubMed Central

Cheng, Shi-Bin; Quinn, Jeffrey A.; Graeber, Carl T.; Filardo, Edward J.

2011-01-01

GPER is a Gs-coupled seven-transmembrane receptor that has been linked to specific estrogen binding and signaling activities that are manifested by plasma membrane-associated enzymes. However, in many cell types, GPER is predominately localized to the endoplasmic reticulum (ER), and only minor amounts of receptor are detectable at the cell surface, an observation that has caused controversy regarding its role as a plasma membrane estrogen receptor. Here, we show that GPER constitutively buds intracellularly into EEA-1+ endosomes from clathrin-coated pits. Nonvisual arrestins-2/-3 do not co-localize with GPER, and expression of arrestin-2 dominant-negative mutants lacking clathrin- or β-adaptin interaction sites fails to block GPER internalization suggesting that arrestins are not involved in GPER endocytosis. Like β1AR, which recycles to the plasma membrane, GPER co-traffics with transferrin+, Rab11+ recycling endosomes. However, endocytosed GPER does not recycle to the cell surface, but instead returns to the trans-Golgi network (TGN) and does not re-enter the ER. GPER is ubiquitinated at the cell surface, exhibits a short half-life (t½ <1 h), and is protected from degradation by the proteasome inhibitor, MG132. Disruption of the TGN by brefeldin A induces the accumulation of endocytosed GPER in Rab11+ perinuclear endosomes and prevents GPER degradation. Our results provide an explanation as to why GPER is not readily detected on the cell surface in some cell types and further suggest that TGN serves as the checkpoint for degradation of endocytosed GPER. PMID:21540189

Improving influenza virological surveillance in Europe: strain-based reporting of antigenic and genetic characterisation data, 11 European countries, influenza season 2013/14

PubMed Central

Broberg, Eeva; Hungnes, Olav; Schweiger, Brunhilde; Prosenc, Katarina; Daniels, Rod; Guiomar, Raquel; Ikonen, Niina; Kossyvakis, Athanasios; Pozo, Francisco; Puzelli, Simona; Thomas, Isabelle; Waters, Allison; Wiman, Åsa; Meijer, Adam

2016-01-01

Influenza antigenic and genetic characterisation data are crucial for influenza vaccine composition decision making. Previously, aggregate data were reported to the European Centre for Disease Prevention and Control by European Union/European Economic Area (EU/EEA) countries. A system for collecting case-specific influenza antigenic and genetic characterisation data was established for the 2013/14 influenza season. In a pilot study, 11 EU/EEA countries reported through the new mechanism. We demonstrated feasibility of reporting strain-based antigenic and genetic data and ca 10% of influenza virus-positive specimens were selected for further characterisation. Proportions of characterised virus (sub)types were similar to influenza virus circulation levels. The main genetic clades were represented by A/StPetersburg/27/2011(H1N1)pdm09 and A/Texas/50/2012(H3N2). A(H1N1)pdm09 viruses were more prevalent in age groups (by years) < 1 (65%; p = 0.0111), 20–39 (50%; p = 0.0046) and 40–64 (55%; p = 0.00001) while A(H3N2) viruses were most prevalent in those ≥ 65 years (62%*; p = 0.0012). Hospitalised patients in the age groups 6–19 years (67%; p = 0.0494) and ≥ 65 years (52%; p = 0.0005) were more frequently infected by A/Texas/50/2012 A(H3N2)-like viruses compared with hospitalised cases in other age groups. Strain-based reporting enabled deeper understanding of influenza virus circulation among hospitalised patients and substantially improved the reporting of virus characterisation data. Therefore, strain-based reporting of readily available data is recommended to all reporting countries within the EU/EEA. PMID:27762211

Endoscopic endonasal approach to the anteromedial temporal fossa and mobilization of the lateral wall of the cavernous sinus through the inferior orbital fissure and V1-V2 corridor: An anatomical study and clinical considerations.

PubMed

Hanakita, Shunya; Chang, Wei-Chieh; Watanabe, Kentaro; Ronconi, Daniel; Labidi, Moujahed; Park, Hun-Ho; Oyama, Kenichi; Bernat, Anne-Laure; Froelich, Sebastien

2018-04-27

The aim of this study was to identify key anatomical landmarks useful in gaining access to the anteromedial temporal region via the corridor formed by the inferior orbital fissure (IOF), the ophthalmic branch (V1), and the maxillary branch (V2) of the trigeminal nerve via the EEA. An anatomical dissection of six cadaver heads was performed to confirm the feasibility and applicability of the EEA for accessing the anteromedial temporal region. Following middle turbinectomy, the lateral recess of the sphenoid sinus was opened, the orbital apex exposed, and the posterior wall of the maxillary sinus was removed, in sequence. The IOF and the pterygopalatine fossa (PPF) were then identified. After opening the foramen rotundum (FR) and removing the bony structure between the FR, V2 was transposed downward. The orbital muscle of Müller was removed. The PPF was mobilized downward exposing the greater wing of the sphenoid bone (GWS). The GWS between V1 and V2 was drilled, thus exposing the temporal dura. With blunt dissection, the medial temporal dura was peeled away from the cavernous sinus (CS) in order to increase access to the anteromedial temporal region. In this study, the anteromedial temporal fossa was exposed by drilling the V1-V2 triangle corridor via the EEA. Endoscopic endonasal exposure of the anteromedial temporal fossa is feasible and requires limited endonasal work. This approach may be considered as an alternate surgical corridor to the temporomesial lobe that offers the advantages of a direct route with less temporal lobe retraction. Copyright © 2018. Published by Elsevier Inc.

HIV testing in Europe: Evaluating the impact, added value, relevance and usability of the European Centre for Disease Prevention and Control (ECDC)'s 2010 HIV testing guidance.

PubMed

Sullivan, Ann K; Sperle, Ida; Raben, Dorthe; Amato-Gauci, Andrew J; Lundgren, Jens Dilling; Yazdanpanah, Yazdan; Jakobsen, Stine Finne; Tavoschi, Lara

2017-11-01

An evaluation of the 2010 ECDC guidance on HIV testing, conducted in October 2015-January 2016, assessed its impact, added value, relevance and usability and the need for updated guidance. Data sources were two surveys: one for the primary target audience (health policymakers and decision makers, national programme managers and ECDC official contact points in the European Union/European Economic Area (EU/EEA) countries and one for a broader target audience (clinicians, civil society organisations and international public health agencies); two moderated focus group discussions  (17 participants each); webpage access data; a literature citation review; and an expert consultation (18 participants) to discuss the evaluation findings. Twenty-three of 28 primary target audience and 31 of 51 broader target audience respondents indicated the guidance was the most relevant when compared with other international guidance. Primary target audience respondents in 11 of 23 countries reported that they had used the guidance in development, monitoring and/or evaluation of their national HIV testing policy, guidelines, programme and/or strategy, and 29 of 51 of the broader target audience respondents reported having used the guidance in their work. Both the primary and broader target audience considered it important or very important to have an EU/EEA-level HIV testing guidance (23/28 and 46/51, respectively). The guidance has been widely used to develop policies, guidelines, programmes and strategies in the EU/EEA and should be regularly updated due to continuous developments in the field in order to continue to serve as an important reference guidance in the region.

HIV testing in Europe: Evaluating the impact, added value, relevance and usability of the European Centre for Disease Prevention and Control (ECDC)’s 2010 HIV testing guidance

PubMed Central

Sullivan, Ann K; Sperle, Ida; Raben, Dorthe; Amato-Gauci, Andrew J; Lundgren, Jens Dilling; Yazdanpanah, Yazdan; Jakobsen, Stine Finne; Tavoschi, Lara

2017-01-01

Background: An evaluation of the 2010 ECDC guidance on HIV testing, conducted in October 2015–January 2016, assessed its impact, added value, relevance and usability and the need for updated guidance. Methods: Data sources were two surveys: one for the primary target audience (health policymakers and decision makers, national programme managers and ECDC official contact points in the European Union/European Economic Area (EU/EEA) countries and one for a broader target audience (clinicians, civil society organisations and international public health agencies); two moderated focus group discussions  (17 participants each); webpage access data; a literature citation review; and an expert consultation (18 participants) to discuss the evaluation findings. Results: Twenty-three of 28 primary target audience and 31 of 51 broader target audience respondents indicated the guidance was the most relevant when compared with other international guidance. Primary target audience respondents in 11 of 23 countries reported that they had used the guidance in development, monitoring and/or evaluation of their national HIV testing policy, guidelines, programme and/or strategy, and 29 of 51 of the broader target audience respondents reported having used the guidance in their work. Both the primary and broader target audience considered it important or very important to have an EU/EEA-level HIV testing guidance (23/28 and 46/51, respectively). Conclusion: The guidance has been widely used to develop policies, guidelines, programmes and strategies in the EU/EEA and should be regularly updated due to continuous developments in the field in order to continue to serve as an important reference guidance in the region. PMID:29208158

Antibiotic susceptibility and intracellular localization of Diplorickettsia massiliensis.

PubMed

Subramanian, Geetha; Barry, Abdoulaye O; Ghigo, Eric; Raoult, Didier; Mediannikov, Oleg

2012-02-01

Diplorickettsia massiliensis is an obligate intracellular bacterium from the Coxiellaceae family recently isolated from Ixodes ricinus ticks. The inhibitory effects of antimicrobial agents were assessed by two different methods, immunofluorescence and Gimenez staining assay. Different markers (EEA1, Lamp-1, Cathepsin D, and LysoTracker Red DND99) were used to reveal the nature of the vacuole containing the bacterium. Ciprofloxacin, levofloxacin, and rifampin had MIC values of 2 lg mL(-1). We found that 4 lg mL(-1) of Doxycycline inhibited the growth of D. massiliensis strain. Surprisingly, D. massiliensis was resistant to chloramphenicol up to the concentration of 64 lg mL(-1). We found that penicillin G, ammonium chloride, gentamycin, omeprazole, bafilomycin A1, and chloroquine were not active against D. massiliensis. Studies performed with markers EEA1, Lamp-1, Cathepsin D, and LysoTracker Red DND99 showed that D. massiliensis is localized within an acidic compartment that is not an early phagosome, but a late phagosome or a phagolysosome. Gimenez staining stays a good method that will work with a very low number of bacteria and can be used to determine the MICs of new therapeutic antibiotics precisely. The resistance profile of D. massiliensis was found to be quite unusual for intracellular Gram-negative bacterium with marked resistance to chloramphenicol. Despite of localization in acidic compartment, pH-neutralizing agents do not significantly inhibit intracellular growth of bacterium. The results of these studies prove that antibiotic resistance does not depend on pH of vacuole. This pH-related mechanism seems not to play a contributing role in the overall resistance of D. massiliensis.

Single Crystal Substrates for Surface Acoustic Wave Devices.

DTIC Science & Technology

1981-01-01

Research Laboratory 2W 5 University Park PA 16802____________ 1 1. CONTROLLING OFFICE NAME ANO ADDRESS Deputy for Electronic Technology (RADC/ EEA )// Janu...avoid contamination during the preparation of the reacted starting powders. Mixina was performed in plastic hall mills with teflon rollers, and sinterinq

Annual Report to Congress on Foreign Economic Collection and Industrial Espionage - 2004

DTIC Science & Technology

2005-04-01

Administration ( DTSA ) • Department of Energy (DOE) • Department of Justice (DOJ) • Department of State, including the Bureau of Intelligence and...Threat Reduction Agency DTSA Defense Technology Security Administration EAR Export Administration Regulations 18 EEA Economic Espionage Act of 1996

Has the "Equal Environments" assumption been tested in twin studies?

PubMed

Eaves, Lindon; Foley, Debra; Silberg, Judy

2003-12-01

A recurring criticism of the twin method for quantifying genetic and environmental components of human differences is the necessity of the so-called "equal environments assumption" (EEA) (i.e., that monozygotic and dizygotic twins experience equally correlated environments). It has been proposed to test the EEA by stratifying twin correlations by indices of the amount of shared environment. However, relevant environments may also be influenced by genetic differences. We present a model for the role of genetic factors in niche selection by twins that may account for variation in indices of the shared twin environment (e.g., contact between members of twin pairs). Simulations reveal that stratification of twin correlations by amount of contact can yield spurious evidence of large shared environmental effects in some strata and even give false indications of genotype x environment interaction. The stratification approach to testing the equal environments assumption may be misleading and the results of such tests may actually be consistent with a simpler theory of the role of genetic factors in niche selection.

The limits of transsellar/transtuberculum surgery for craniopharyngioma.

PubMed

Koutourousiou, Maria; Fernandez-Miranda, Juan C; Wang, Eric W; Snyderman, Carl H; Gardner, Paul A

2018-06-01

The proximity of craniopharyngiomas to vital neurovascular structures and their high recurrence rates make them one of the most challenging brain tumors to treat. Although surgery remains the first line of therapy and offers the best chance of radical resection and oncological cure, the high recurrence tendency of craniopharyngiomas, even after apparent total removal, often makes adjuvant treatment essential. The endoscopic endonasal approach (EEA) has been recently introduced as a treatment option for both pediatric and adult craniopharyngiomas, rapidly gaining wide acceptance over the traditional transcranial approaches. Although the primary role of EEA over traditional transcranial approaches has been slowly accepted in the literature, little has been written about the limitations and potential contraindications of this approach in the treatment of craniopharyngiomas. This article presents the advantages and highlights the limitations of endoscopic transsellar/transtuberculum surgery for craniopharyngiomas. In every case, surgery should be tailored to individuals based on their age and comorbidities, presenting symptoms, tumor characteristics, prior treatment and treatment tolerance, as well as the surgeon's preference based on personal experience and comfort.

Visualization of early influenza A virus trafficking in human dendritic cells using STED microscopy.

PubMed

Baharom, Faezzah; Thomas, Oliver S; Lepzien, Rico; Mellman, Ira; Chalouni, Cécile; Smed-Sörensen, Anna

2017-01-01

Influenza A viruses (IAV) primarily target respiratory epithelial cells, but can also replicate in immune cells, including human dendritic cells (DCs). Super-resolution microscopy provides a novel method of visualizing viral trafficking by overcoming the resolution limit imposed by conventional light microscopy, without the laborious sample preparation of electron microscopy. Using three-color Stimulated Emission Depletion (STED) microscopy, we visualized input IAV nucleoprotein (NP), early and late endosomal compartments (EEA1 and LAMP1 respectively), and HLA-DR (DC membrane/cytosol) by immunofluorescence in human DCs. Surface bound IAV were internalized within 5 min of infection. The association of virus particles with early endosomes peaked at 5 min when 50% of NP+ signals were also EEA1+. Peak association with late endosomes occurred at 15 min when 60% of NP+ signals were LAMP1+. At 30 min of infection, the majority of NP signals were in the nucleus. Our findings illustrate that early IAV trafficking in human DCs proceeds via the classical endocytic pathway.

Spatiotemporal Dynamics of Adenovirus Membrane Rupture and Endosomal Escape

PubMed Central

Maier, Oana; Marvin, Shauna A.; Wodrich, Harald; Campbell, Edward M.

2012-01-01

A key step in adenovirus cell entry is viral penetration of cellular membranes to gain access to the cytoplasm and deliver the genome to the nucleus. Yet little is known about this important event in the adenoviral life cycle. Using the cytosolic protein galectin-3 (gal3) as a marker of membrane rupture with both live- and fixed-cell imaging, we demonstrate that in the majority of instances, exposure of pVI and recruitment of gal3 to ruptured membranes occur early at or near the cell surface and occur minimally in EEA-1-positive (EEA-1+) early endosomes or LAMP-1+ late endosomes/lysosomes. Live-cell imaging of Ad5 egress from gal3+ endosomes occurs most frequently from perinuclear locations. While the Ad5 capsid is observed escaping from gal3+ endosomes, pVI appears to remain associated with the gal3+ ruptured endosomes. Thus, Ad5 membrane rupture and endosomal escape appear to be both spatially and temporally distinct events. PMID:22855481

European downstream oil industry safety performance : statistical summary of reported incidents, 1998

DOT National Transportation Integrated Search

1999-07-01

This report is the fifth by CONCAWE reviewing the safety performance of the downstream oil industry in Europe. The area of coverage is primarily the EU, EEA and Hungary, but for some companies the data for other European countries such as Poland, Cze...

The potential impact of Brexit and immigration policies on the GP workforce in England: a cross-sectional observational study of GP qualification region and the characteristics of the areas and population they served in September 2016.

PubMed

Esmail, Aneez; Panagioti, Maria; Kontopantelis, Evangelos

2017-11-16

The UK is dependent on international doctors, with a greater proportion of non-UK qualified doctors working in its universal health care system than in any other European country, except Ireland and Norway. The terms of the UK exit from the European Union can reduce the ability of European Economic Area (EEA) qualified doctors to work in the UK, while new visa requirements will significantly restrict the influx of non-EEA doctors. We aimed to explore the implications of policy restrictions on immigration, by regionally and spatially describing the characteristics of general practitioners (GPs) by region of medical qualification and the characteristics of the populations they serve. This is a cross-sectional study on 37,792 of 41,865 GPs in England, as of 30 September 2016. The study involved age, sex, full-time equivalent (FTE), country and region of qualification and geography (organisational regions) of individual GPs. Additionally at the practice and geography levels, we studied patient list size by age groups, average patient location deprivation, the overall morbidity as measured by the Quality and Outcomes Framework (QOF) and the average payment made to primary care per patient. Non-UK qualified GPs comprised 21.1% of the total numbers of GPs, with the largest percentage observed in East England (29.8%). Compared to UK qualified GPs, EEA and elsewhere qualified GPs had higher FTE (medians were 0.80, 0.89 and 0.93, respectively) and worked in practices with higher median patient location deprivation (18.3, 22.5 and 25.2, respectively). Practices with high percentages of EEA and elsewhere qualified GPs served patients who resided in more deprived areas, had lower GP-to-patient ratios and lower GP-to-cumulative QOF register ratios. A decrease in pay as the percentage of elsewhere qualified GPs increased was observed; a 10% increase in elsewhere qualified GPs was linked to a £1 decrease (95% confidence interval 0.5-1.4) in average pay per patient. A large percentage of the UK general practice workforce consists of non-UK qualified GPs who work longer hours, are older and serve a larger number of patients in more deprived areas. Following Brexit, difficulties in replacing this valuable workforce will primarily threaten the care delivery in deprived areas.

High Strength Carbide-Based Fibrous Monolith Materials for Solid Rocket Nozzles

DTIC Science & Technology

2008-02-19

individual powders are blended with thermoplastic melt-extrudable polymer binders and plasticizers , using a high shear mixer (Brabender Technologie Inc...Ethylene Ethyl Acrylate, EEA DuPont MFI 1.5 and 20 DPDA 9169 NT Stearic Acid Crompton Hystrene 97 18 NF FG 65 References 1. J. B

Tuition Fees for International Students in Finland: Where to Go from Here?

ERIC Educational Resources Information Center

Cai, Yuzhuo; Kivisto, Jussi

2013-01-01

Recent Finnish policies have encouraged Finnish higher education institutions to develop a market-oriented approach to international higher education by implementing fee-based educational programmes for students from outside the European Union (EU) and the European Economic Area (EEA). From the Finnish government's perspective, higher education is…

Functional properties of hepatocytes in vitro are correlated with cell polarity maintenance.

PubMed

Zeigerer, Anja; Wuttke, Anne; Marsico, Giovanni; Seifert, Sarah; Kalaidzidis, Yannis; Zerial, Marino

2017-01-01

Exploring the cell biology of hepatocytes in vitro could be a powerful strategy to dissect the molecular mechanisms underlying the structure and function of the liver in vivo. However, this approach relies on appropriate in vitro cell culture systems that can recapitulate the cell biological and metabolic features of the hepatocytes in the liver whilst being accessible to experimental manipulations. Here, we adapted protocols for high-resolution fluorescence microscopy and quantitative image analysis to compare two primary hepatocyte culture systems, monolayer and collagen sandwich, with respect to the distribution of two distinct populations of early endosomes (APPL1 and EEA1-positive), endocytic capacity, metabolic and signaling activities. In addition to the re-acquisition of hepatocellular polarity, primary hepatocytes grown in collagen sandwich but not in monolayer culture recapitulated the apico-basal distribution of EEA1 endosomes observed in liver tissue. We found that such distribution correlated with the organization of the actin cytoskeleton in vitro and, surprisingly, was dependent on the nutritional state in vivo. Hepatocytes in collagen sandwich also exhibited faster kinetics of low-density lipoprotein (LDL) and epidermal growth factor (EGF) internalization, showed improved insulin sensitivity and preserved their ability for glucose production, compared to hepatocytes in monolayer cultures. Although no in vitro culture system can reproduce the exquisite structural features of liver tissue, our data nevertheless highlight the ability of the collagen sandwich system to recapitulate key structural and functional properties of the hepatocytes in the liver and, therefore, support the usage of this system to study aspects of hepatocellular biology in vitro. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Defense Systems Modernization and Sustainment Initiative

DTIC Science & Technology

2008-07-21

surface coatings, including metals and plastics , and coating application processes were developed for repairing bearing surfaces. The Modernization through...technologies and applications utilized by the NC3R team. " : eea -cy c le en- ne Oglsti t trei eci d ’leet ~h SSt re*,0 an tnding ecsio Uiprrts ren.c" O e

Installation Restoration Program. Phase 2. Confirmation/Quantification. Stage 1

DTIC Science & Technology

1986-03-28

Comftin,,,on Fvvem in.eea),a n dftl by bloe * numri ~A Problem Confirmation Study was conducted at Burlington Air National Guard BasE in Vermont. The...cont’d) 3. Characteristics(cont’d)- Biological Agent ( ) Volatile (4f Toxic (). Reactive ( ) 4. Toxicity: TLVs IDLHs 5. Special Hazards 6. Acute

Livestock-associated meticillin-resistant Staphylococcus aureus (MRSA) among human MRSA isolates, European Union/European Economic Area countries, 2013

PubMed Central

Kinross, Pete; Petersen, Andreas; Skov, Robert; Van Hauwermeiren, Evelyn; Pantosti, Annalisa; Laurent, Frédéric; Voss, Andreas; Kluytmans, Jan; Struelens, Marc J; Heuer, Ole; Monnet, Dominique L

2017-01-01

Currently, surveillance of livestock-associated meticillin-resistant Staphylococcus aureus (LA-MRSA) in humans in Europe is not systematic but mainly event-based. In September 2014, the European Centre for Disease Prevention and Control (ECDC) initiated a questionnaire to collect data on the number of LA-MRSA from human samples (one isolate per patient) from national/regional reference laboratories in European Union/European Economic Area (EU/EEA) countries in 2013. Identification of LA-MRSA as clonal complex (CC) 398 by multilocus sequence typing (MLST) was preferred, although surrogate methods such as spa-typing were also accepted. The questionnaire was returned by 28 laboratories in 27 EU/EEA countries. Overall, LA-MRSA represented 3.9% of 13,756 typed MRSA human isolates, but it represented ≥ 10% in five countries (Belgium, Denmark, Spain, the Netherlands and Slovenia). Seven of the reference laboratories did not type MRSA isolates in 2013. To monitor the dispersion of LA-MRSA and facilitate targeted control measures, we advocate periodic systematic surveys or integrated multi-sectorial surveillance. PMID:29113628

Livestock-associated meticillin-resistant Staphylococcus aureus (MRSA) among human MRSA isolates, European Union/European Economic Area countries, 2013.

PubMed

Kinross, Pete; Petersen, Andreas; Skov, Robert; Van Hauwermeiren, Evelyn; Pantosti, Annalisa; Laurent, Frédéric; Voss, Andreas; Kluytmans, Jan; Struelens, Marc J; Heuer, Ole; Monnet, Dominique L

2017-11-01

Currently, surveillance of livestock-associated meticillin-resistant Staphylococcus aureus (LA-MRSA) in humans in Europe is not systematic but mainly event-based. In September 2014, the European Centre for Disease Prevention and Control (ECDC) initiated a questionnaire to collect data on the number of LA-MRSA from human samples (one isolate per patient) from national/regional reference laboratories in European Union/European Economic Area (EU/EEA) countries in 2013. Identification of LA-MRSA as clonal complex (CC) 398 by multilocus sequence typing (MLST) was preferred, although surrogate methods such as spa -typing were also accepted. The questionnaire was returned by 28 laboratories in 27 EU/EEA countries. Overall, LA-MRSA represented 3.9% of 13,756 typed MRSA human isolates, but it represented ≥ 10% in five countries (Belgium, Denmark, Spain, the Netherlands and Slovenia). Seven of the reference laboratories did not type MRSA isolates in 2013. To monitor the dispersion of LA-MRSA and facilitate targeted control measures, we advocate periodic systematic surveys or integrated multi-sectorial surveillance.

Energy Engineering Analysis (EEA) program for Lone Star Army Ammunition Plant, Texas. Executive summary. Final report

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

NONE

The objective of this Energy Engineering Analysis (EEA) for LSAAP is threefold: Develop a systematic plan of projects which will result in reducing energy consumption. Consider renewable energy sources with the objective of establishing an orderly procedure for reducing use of non-renewable energy sources. Determine the feasibility of Total Energy (TE), Selective Energy (SE), and Central Heating Plant (CHP) concepts using alternative fuels. In essence, an assessment of the entire energy picture at LSAAP was undertaken. This report is a summary of that effort. LSAAP was originally built during 1941 and 1942 as a shell loading plant for the Army.more » After World War II, the facility was deactivated until 1951 when it was reactivated as a Government Owned, Contractor Operated (GOCO) facility. Day and Zimmerman was selected as the operator in 1951 and has been the operating contractor ever since. Located just west of Texarkana, Texas, LSAAP encompasses an area of approximately 15,546 acres. The primary mission of LSAAP is to load, assemble and pack ammunition and ammunition components for the Army.« less

Childhood multidrug-resistant tuberculosis in the European Union and European Economic Area: an analysis of tuberculosis surveillance data from 2007 to 2015.

PubMed

Ködmön, Csaba; van den Boom, Martin; Zucs, Phillip; van der Werf, Marieke Johanna

2017-11-01

BackgroundConfirming tuberculosis (TB) in children and obtaining information on drug susceptibility is essential to ensure adequate treatment. We assessed whether there are gaps in diagnosis and treatment of multidrug-resistant (MDR) TB in children in the European Union and European Economic Area (EU/EEA), quantified the burden of MDR TB in children and characterised cases. Methods : We analysed surveillance data from 2007 to 2015 for paediatric cases younger than 15 years. Results : In that period, 26 EU/EEA countries reported 18,826 paediatric TB cases of whom 4,129 (21.9%) were laboratory-confirmed. Drug susceptibility testing results were available for 3,378 (17.9%), representing 81.8% of the confirmed cases. The majority (n = 2,967; 87.8%) had drug-sensitive TB, 249 (7.4%) mono-resistant TB, 64 (1.9%) poly-resistant TB, 90 (2.7%) MDR TB and eight (0.2%) had extensively drug-resistant (XDR) TB. MDR TB was more frequently reported among paediatric cases with foreign background (adjusted odds ratio (aOR) = 1.73; 95% confidence interval (95% CI): 1.12-2.67) or previous TB treatment (aOR: 6.42; 95% CI: 3.24-12.75). Successful treatment outcome was reported for 58 of 74 paediatric MDR TB cases with outcome reported from 2007 to 2013; only the group of 5-9 years-olds was significantly associated with unsuccessful treatment outcome (crude odds ratio (cOR) = 11.45; 95% CI: 1.24-106.04). Conclusions : The burden of MDR TB in children in the EU/EEA appears low, but may be underestimated owing to challenges in laboratory confirmation. Diagnostic improvements are needed for early detection and adequate treatment of MDR TB. Children previously treated for TB or of foreign origin may warrant higher attention.

SVHC in imported articles: REACH authorisation requirement justified under WTO rules.

PubMed

Schenten, Julian; Führ, Martin

2016-01-01

The purpose of the REACH Regulation is to ensure a high level of protection of human health and the environment as well as the free circulation of substances on the internal market while enhancing competitiveness and innovation. To this end, REACH introduces, among other instruments, the authorisation regime for substances of very high concern (SVHC) that are listed on Annex XIV of the regulation. After expiration of the transitional period for each Annex XIV-SVHC, articles, such as most products of daily use, produced in the European Economic Area (EEA) may not contain such substances unless an authorisation was granted for the specific use or this use falls within the scope of an exemption from the authorisation requirement. The authorisation scheme does, however, only apply to SVHC used in the EEA. As a consequence, REACH does not regulate SVHC entering the European market as part of imported articles which burden human health and the environment. Moreover, from an economic perspective, domestic articles are subject to stricter requirements than those which are produced abroad, putting actors from within the EEA at competitive disadvantage and thus impeding the intention of REACH to enhance competitiveness and innovation. One option to close this regulatory gap could be to extend the authorisation requirement to SVHC present in imported articles. A legal appraisal on behalf of the German Environment Agency (UBA) assesses whether such option would be in accordance with the specifications of WTO world trade law. It concludes that, measured by the standards of the WTO dispute settlement practice, such an extended authorisation scheme would neither violate the principles of national treatment and most-favoured nation treatment. Also, such regulation would not constitute an unnecessary obstacle to trade, since the extended authorisation requirement would pursue a legitimate objective covered by the regulatory autonomy of the EU and, furthermore, the regulation would not be more trade-restrictive than necessary. The contribution at hand summarises the main findings while taking into account first reactions to the legal appraisal.

Engagement of the small GTPase Rab31 protein and its effector, early endosome antigen 1, is important for trafficking of the ligand-bound epidermal growth factor receptor from the early to the late endosome.

PubMed

Chua, Christelle En Lin; Tang, Bor Luen

2014-05-02

Rab31 is a member of the Rab5 subfamily of Rab GTPases. Although localized largely to the trans-Golgi network, it shares common guanine nucleotide exchange factors and effectors with other Rab5 subfamily members that have been implicated in endocytic membrane traffic. We investigated whether Rab31 also has a role in the trafficking of the ligand-bound EGF receptor (EGFR) internalized through receptor-mediated endocytosis. We found that loss of Rab31 inhibits, but overexpression enhances, EGFR trafficking to the late endosomes and that the effect of Rab31 silencing could be specifically rescued by overexpression of a silencing-resistant form of Rab31. Rab31 was found to interact with the EGFR by coimmunoprecipitation and affinity pulldown analyses, and the primarily trans-Golgi network-localized Rab31 has increased colocalization with the EGFR in A431 cells 30 min after pulsing with EGF. A glycerol gradient sedimentation assay suggested that Rab31 is sequestered into a high molecular weight complex after stimulation with EGF, as was early endosome antigen 1 (EEA1), a factor responsible for endosomal tethering and fusion events. We found that loss of EEA1 reduced the interaction between Rab31 and the EGFR and abrogated the effect of Rab31 overexpression on the trafficking of the EGFR. Likewise, loss of GAPex5, a Rab31 guanine nucleotide exchange factor that has a role in ubiquitination and degradation of the EGFR, reduced the interaction of Rab31 with the EGFR and its effect on EGFR trafficking. Taken together, our results suggest that Rab31 is an important regulator of endocytic trafficking of the EGFR and functions in an EGFR trafficking complex that includes EEA1 and GAPex5.

Engagement of the Small GTPase Rab31 Protein and Its Effector, Early Endosome Antigen 1, Is Important for Trafficking of the Ligand-bound Epidermal Growth Factor Receptor from the Early to the Late Endosome*

PubMed Central

Chua, Christelle En Lin; Tang, Bor Luen

2014-01-01

Rab31 is a member of the Rab5 subfamily of Rab GTPases. Although localized largely to the trans-Golgi network, it shares common guanine nucleotide exchange factors and effectors with other Rab5 subfamily members that have been implicated in endocytic membrane traffic. We investigated whether Rab31 also has a role in the trafficking of the ligand-bound EGF receptor (EGFR) internalized through receptor-mediated endocytosis. We found that loss of Rab31 inhibits, but overexpression enhances, EGFR trafficking to the late endosomes and that the effect of Rab31 silencing could be specifically rescued by overexpression of a silencing-resistant form of Rab31. Rab31 was found to interact with the EGFR by coimmunoprecipitation and affinity pulldown analyses, and the primarily trans-Golgi network-localized Rab31 has increased colocalization with the EGFR in A431 cells 30 min after pulsing with EGF. A glycerol gradient sedimentation assay suggested that Rab31 is sequestered into a high molecular weight complex after stimulation with EGF, as was early endosome antigen 1 (EEA1), a factor responsible for endosomal tethering and fusion events. We found that loss of EEA1 reduced the interaction between Rab31 and the EGFR and abrogated the effect of Rab31 overexpression on the trafficking of the EGFR. Likewise, loss of GAPex5, a Rab31 guanine nucleotide exchange factor that has a role in ubiquitination and degradation of the EGFR, reduced the interaction of Rab31 with the EGFR and its effect on EGFR trafficking. Taken together, our results suggest that Rab31 is an important regulator of endocytic trafficking of the EGFR and functions in an EGFR trafficking complex that includes EEA1 and GAPex5. PMID:24644286

National Income Inequality and Declining GDP Growth Rates Are Associated with Increases in HIV Diagnoses among People Who Inject Drugs in Europe: A Panel Data Analysis

PubMed Central

Nikolopoulos, Georgios K.; Fotiou, Anastasios; Kanavou, Eleftheria; Richardson, Clive; Detsis, Marios; Pharris, Anastasia; Suk, Jonathan E.; Semenza, Jan C.; Costa-Storti, Claudia; Paraskevis, Dimitrios; Sypsa, Vana; Malliori, Melpomeni-Minerva; Friedman, Samuel R.; Hatzakis, Angelos

2015-01-01

Background There is sparse evidence that demonstrates the association between macro-environmental processes and drug-related HIV epidemics. The present study explores the relationship between economic, socio-economic, policy and structural indicators, and increases in reported HIV infections among people who inject drugs (PWID) in the European Economic Area (EEA). Methods We used panel data (2003–2012) for 30 EEA countries. Statistical analyses included logistic regression models. The dependent variable was taking value 1 if there was an outbreak (significant increase in the national rate of HIV diagnoses in PWID) and 0 otherwise. Explanatory variables included the growth rate of Gross Domestic Product (GDP), the share of the population that is at risk for poverty, the unemployment rate, the Eurostat S80/S20 ratio, the Gini coefficient, the per capita government expenditure on health and social protection, and variables on drug control policy and drug-using population sizes. Lags of one to three years were investigated. Findings In multivariable analyses, using two-year lagged values, we found that a 1% increase of GDP was associated with approximately 30% reduction in the odds of an HIV outbreak. In GDP-adjusted analyses with three-year lagged values, the effect of the national income inequality on the likelihood of an HIV outbreak was significant [S80/S20 Odds Ratio (OR) = 3.89; 95% Confidence Interval (CI): 1.15 to 13.13]. Generally, the multivariable analyses produced similar results across three time lags tested. Interpretation Given the limitations of ecological research, we found that declining economic growth and increasing national income inequality were associated with an elevated probability of a large increase in the number of HIV diagnoses among PWID in EEA countries during the last decade. HIV prevention may be more effective if developed within national and European-level policy contexts that promote income equality, especially among vulnerable groups. PMID:25875598

National income inequality and declining GDP growth rates are associated with increases in HIV diagnoses among people who inject drugs in Europe: a panel data analysis.

PubMed

Nikolopoulos, Georgios K; Fotiou, Anastasios; Kanavou, Eleftheria; Richardson, Clive; Detsis, Marios; Pharris, Anastasia; Suk, Jonathan E; Semenza, Jan C; Costa-Storti, Claudia; Paraskevis, Dimitrios; Sypsa, Vana; Malliori, Melpomeni-Minerva; Friedman, Samuel R; Hatzakis, Angelos

2015-01-01

There is sparse evidence that demonstrates the association between macro-environmental processes and drug-related HIV epidemics. The present study explores the relationship between economic, socio-economic, policy and structural indicators, and increases in reported HIV infections among people who inject drugs (PWID) in the European Economic Area (EEA). We used panel data (2003-2012) for 30 EEA countries. Statistical analyses included logistic regression models. The dependent variable was taking value 1 if there was an outbreak (significant increase in the national rate of HIV diagnoses in PWID) and 0 otherwise. Explanatory variables included the growth rate of Gross Domestic Product (GDP), the share of the population that is at risk for poverty, the unemployment rate, the Eurostat S80/S20 ratio, the Gini coefficient, the per capita government expenditure on health and social protection, and variables on drug control policy and drug-using population sizes. Lags of one to three years were investigated. In multivariable analyses, using two-year lagged values, we found that a 1% increase of GDP was associated with approximately 30% reduction in the odds of an HIV outbreak. In GDP-adjusted analyses with three-year lagged values, the effect of the national income inequality on the likelihood of an HIV outbreak was significant [S80/S20 Odds Ratio (OR) = 3.89; 95% Confidence Interval (CI): 1.15 to 13.13]. Generally, the multivariable analyses produced similar results across three time lags tested. Given the limitations of ecological research, we found that declining economic growth and increasing national income inequality were associated with an elevated probability of a large increase in the number of HIV diagnoses among PWID in EEA countries during the last decade. HIV prevention may be more effective if developed within national and European-level policy contexts that promote income equality, especially among vulnerable groups.

Mouse Polyomavirus Enters Early Endosomes, Requires Their Acidic pH for Productive Infection, and Meets Transferrin Cargo in Rab11-Positive Endosomes

PubMed Central

Liebl, David; Difato, Francesco; Horníková, Lenka; Mannová, Petra; Štokrová, Jitka; Forstová, Jitka

2006-01-01

Mouse polyomavirus (PyV) virions enter cells by internalization into smooth monopinocytic vesicles, which fuse under the cell membrane with larger endosomes. Caveolin-1 was detected on monopinocytic vesicles carrying PyV particles in mouse fibroblasts and epithelial cells (33). Here, we show that PyV can be efficiently internalized by Jurkat cells, which do not express caveolin-1 and lack caveolae, and that overexpression of a caveolin-1 dominant-negative mutant in mouse epithelial cells does not prevent their productive infection. Strong colocalization of VP1 with early endosome antigen 1 (EEA1) and of EEA1 with caveolin-1 in mouse fibroblasts and epithelial cells suggests that the monopinocytic vesicles carrying the virus (and vesicles containing caveolin-1) fuse with EEA1-positive early endosomes. In contrast to SV40, PyV infection is dependent on the acidic pH of endosomes. Bafilomycin A1 abolished PyV infection, and an increase in endosomal pH by NH4Cl markedly reduced its efficiency when drugs were applied during virion transport towards the cell nucleus. The block of acidification resulted in the retention of a fraction of virions in early endosomes. To monitor further trafficking of PyV, we used fluorescent resonance energy transfer (FRET) to determine mutual localization of PyV VP1 with transferrin and Rab11 GTPase at a 2- to 10-nm resolution. Positive FRET between PyV VP1 and transferrin cargo and between PyV VP1 and Rab11 suggests that during later times postinfection (1.5 to 3 h), the virus meets up with transferrin in the Rab11-positive recycling endosome. These results point to a convergence of the virus and the cargo internalized by different pathways in common transitional compartments. PMID:16611921

Determinants of Mobility of Students in Europe: Empirical Evidence for the Period 1998-2009

ERIC Educational Resources Information Center

Caruso, Raul; de Wit, Hans

2015-01-01

This article studies the economic determinants of intra-European student mobility. We constructed a panel of 33 European countries for the period 1998-2009. The dependent variable is the inflow of foreign students (International Standard Classification of Education [ISCED] 5-6) from European Union (EU)-27, European Economic Area (EEA), and…

A Software Framework for Blast Event Simulation

DTIC Science & Technology

2006-11-01

The sequence of images shown in Figure 1 show a steel container filled with PBX9501 (a plastic bonded explosive) that is preheated to the ignition...β ρ ρ α eeA I I bIaIP (2) a, b, A, B, α,and β are constants. The values used in this model are taken as 4 http://www.reaction-eng.com

Consultation Process on the European Commission's "Memorandum on Lifelong Learning": Analysis of National Reports. CEDEFOP Panorama Series.

ERIC Educational Resources Information Center

Ni Cheallaigh, Martina, Ed.

This document presents outcomes of a review of the Memorandum on Lifelong Learning (LL) consultation reports from member states of the European Commission and EEA (European Economic Area) countries that collected all views on how best to make progress in implementing LL at local, regional, and national levels. Section 1 provides an overview of…

Potential adjustment methodology for missing data and reporting delay in the HIV Surveillance System, European Union/European Economic Area, 2015.

PubMed

Rosinska, Magdalena; Pantazis, Nikos; Janiec, Janusz; Pharris, Anastasia; Amato-Gauci, Andrew J; Quinten, Chantal; Ecdc Hiv/Aids Surveillance Network

2018-06-01

Accurate case-based surveillance data remain the key data source for estimating HIV burden and monitoring prevention efforts in Europe. We carried out a literature review and exploratory analysis of surveillance data regarding two crucial issues affecting European surveillance for HIV: missing data and reporting delay. Initial screening showed substantial variability of these data issues, both in time and across countries. In terms of missing data, the CD4+ cell count is the most problematic variable because of the high proportion of missing values. In 20 of 31 countries of the European Union/European Economic Area (EU/EEA), CD4+ counts are systematically missing for all or some years. One of the key challenges related to reporting delays is that countries undertake specific one-off actions in effort to capture previously unreported cases, and that these cases are subsequently reported with excessive delays. Slightly different underlying assumptions and effectively different models may be required for individual countries to adjust for missing data and reporting delays. However, using a similar methodology is recommended to foster harmonisation and to improve the accuracy and usability of HIV surveillance data at national and EU/EEA levels.

Endoscopic endonasal approaches for the management of skull base meningiomas. Selection criteria and clinical outcomes.

PubMed

Todeschini, Alexandre B; Otto, Bradley A; Carrau, Ricardo L; Prevedello, Daniel M

2018-05-28

Meningiomas are the most common primary intracranial tumor, arising from different locations, including the skull base. Despite advances in adjuvant treatments, surgical resection remains the main and best treatment for meningiomas. New surgical strategies, such as the endoscopic endonasal approach, have greatly contributed in achieving maximum and total safe resection, preserving the patient's neurological function. Based on the senior authors large experience and a review of the current literature, we have compiled this chapter. We review the surgical technique used at our institution and the most relevant aspects of patient selection when considering resecting a skull base meningioma using the the EEA. Further consideration is given to some skull base meningiomas arising from specific locations with some case examples. The EEA is not an ideal approach for every skull base meningioma. Careful evaluation of the surrounding neurovascular structures surrounding the tumor is imperative to select the appropriate surgical corridor for a safe resection. Nevertheless, for appropriately selected cases, the endoscopic technique is a very valuable tool with some evidences of being superior to the microscopic transcranial approach. A dual-trained surgeon, in both endoscopic and transcranial approaches, is the best alternative to achieve the best patient outcome.

Petrous apex cholesterol granuloma: importance of pedicled nasoseptal flap in addition to silicone T-tube for prevention of occlusion of drainage route in transsphenoidal approach--a technical note.

PubMed

Shibao, Shunsuke; Toda, Masahiro; Tomita, Toshiki; Saito, Katsuya; Ogawa, Kaoru; Kawase, Takeshi; Yoshida, Kazunari

2015-01-01

Recently, petrous apex cholesterol granulomas (CGs) have been treated via the endoscopic endonasal transsphenoidal approach (EEA) using a silicone tube, to prevent drainage route occlusion. Occlusion of the drainage route has led to problems with recurrence. The aim of this report is to describe the use of a surgical technique to prevent drainage route occlusion. In surgical technique, the posterolateral wall of the sphenoid sinus was opened by EEA. After cyst debridement, a vascularized nasoseptal flap with a width of approximately 4 cm was inserted into the lumen with a silicone T-tube with a diameter of 7 mm. This technique was used in two patients: the first patient during the second operation after recurrence following occlusion of the drainage route, and the second patient during the first operation. Opening of the cyst wall was confirmed endoscopically in both patients 12-24 months after surgery, even after removal of the T-tube. In conclusion, the use of a pedicled nasoseptal flap with a silicone tube is useful to prevent CG recurrence, by paranasal cavitization of the cystic cavity.

Endoscopic endonasal approach for the treatment of anterior skull base tumours.

PubMed

López, Fernando; Suárez, Vanessa; Costales, María; Rodrigo, Juan P; Suárez, Carlos; Llorente, José Luis

2012-01-01

The increasing expertise of transnasal endoscopic surgery has recently expanded its indications to include the management of tumours affecting the skull base. We report our experience with endoscopic management of these tumours, emphasising the indications and surgical technique used. A retrospective analysis was performed of patients treated by an endoscopic endonasal approach (EEA) in our department from 2004 until 2011. Sixty-three patients were analysed. We performed an endoscopic craniofacial resection in 32 patients (51%), an expanded EEA in 22 (35%), a transclival approach in 6 (9%) and a transpterygoid approach in 3 (5%). The most frequent benign tumour was nasopharyngeal angiofibroma (24%), while adenocarcinoma (30%) was the most common among malignancies. Mean follow-up was 26 months (range: 6 to 84 months). The complication rate was 5% and resection was complete in 56 cases (89%). The 5-year overall-survival was 71% in patients with malignant tumours and the effectiveness was 100% in benign tumours. Our results support that endoscopic surgery, when properly planned, represents a valid alternative to standard surgical approaches for the management of skull base tumours. Copyright © 2012 Elsevier España, S.L. All rights reserved.

Using non-local databases for the environmental assessment of industrial activities: The case of Latin America

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

Osses de Eicker, Margarita, E-mail: Margarita.Osses@empa.c; Hischier, Roland, E-mail: Roland.Hischier@empa.c; Hurni, Hans, E-mail: Hans.Hurni@cde.unibe.c

2010-04-15

Nine non-local databases were evaluated with respect to their suitability for the environmental assessment of industrial activities in Latin America. Three assessment methods were considered, namely Life Cycle Assessment (LCA), Environmental Impact Assessment (EIA) and air emission inventories. The analysis focused on data availability in the databases and the applicability of their international data to Latin American industry. The study showed that the European EMEP/EEA Guidebook and the U.S. EPA AP-42 database are the most suitable ones for air emission inventories, whereas the LCI database Ecoinvent is the most suitable one for LCA and EIA. Due to the data coveragemore » in the databases, air emission inventories are easier to develop than LCA or EIA, which require more comprehensive information. One strategy to overcome the limitations of non-local databases for Latin American industry is the combination of validated data from international databases with newly developed local datasets.« less

Chromophoric dissolved organic matter and microbial enzymatic activity. A biophysical approach to understand the marine carbon cycle.

PubMed

Gonnelli, Margherita; Vestri, Stefano; Santinelli, Chiara

2013-12-01

This study reports the first information on extracellular enzymatic activity (EEA) combined with a study of DOM dynamics at the Arno River mouth. DOM dynamics was investigated from both a quantitative (dissolved organic carbon, DOC) and a qualitative (absorption and fluorescence of chromophoric DOM, CDOM) perspective. The data here reported highlight that the Arno River was an important source of both DOC and CDOM for this coastal area. CDOM optical properties suggested that terrestrial DOM did not undergo simple dilution at the river mouth but, other physical-chemical and biological processes were probably at work to change its molecular characteristics. This observation was further supported by the "potential" enzymatic activity of β-glucosidase (BG) and leucine aminopeptidase (LAP). Their Vmax values were markedly higher in the river water than in the seawater and their ratio suggested that most of the DOM used by microbes in the Arno River was polysaccharide-like, while in the seawater it was mainly protein-like. © 2013. Published by Elsevier B.V. All rights reserved.

GlobCorine- A Joint EEA-ESA Project for Operational Land Cover and Land Use Mapping at Pan-European Scale

NASA Astrophysics Data System (ADS)

Bontemps, S.; Defourny, P.; Van Bogaert, E.; Weber, J. L.; Arino, O.

2010-12-01

Regular and global land cover mapping contributes to evaluating the impact of human activities on the environment. Jointly supported by the European Space Agency and the European Environmental Agency, the GlobCorine project builds on the GlobCover findings and aims at making the full use of the MERIS time series for frequent land cover monitoring. The GlobCover automated classification approach has been tuned to the pan-European continent and adjusted towards a classification compatible with the Corine typology. The GlobCorine 2005 land cover map has been achieved, validated and made available to a broad- level stakeholder community from the ESA website. A first version of the GlobCorine 2009 map has also been produced, demonstrating the possibility for an operational production of frequent and updated global land cover maps.

Alimentary Tract as Entry Route for Hantavirus Infection

DTIC Science & Technology

Hantaviruses are zoonotic agents that cause hemorrhagic fever with renal and/or cardiopulmonary manifestations, reaching fatality rates of up to 50...predominant endemic hantavirus in Europe, is associated with mild forms of hemorrhagic fever with renal syndrome. PUUV is transmitted to humans by exposure to...viral association with endosomal antigen EEA-1, followed by virus replication and loss of epithelial barrier function with concomitant basolateral

New estimate of valvuloarterial impedance in aortic valve stenosis: A cardiac magnetic resonance study.

PubMed

Soulat, Gilles; Kachenoura, Nadjia; Bollache, Emilie; Perdrix, Ludivine; Diebold, Benoit; Zhygalina, Valentina; Latremouille, Christian; Laurent, Stephane; Fabiani, Jean-Noel; Mousseaux, Elie

2017-03-01

Valvuloarterial impedance (Z VA ), estimating left ventricle (LV) afterload, has been proposed in transthoracic echocardiography (TTE) as a predictor of mortality in aortic valve stenosis (AVS). However, its calculation differs from arterial characteristic impedance (Z C ). Our aim was to apply the concept of Z C calculation to estimate Z VA from MR with carotid tonometry and to evaluate these indices through their associations with symptoms, LV diastolic function and aortic stiffness. In 40 patients with AVS (76 ± 13 years), Z VA-TI derived from velocity time integral and E/Ea were estimated by TTE. Z VA-INS , based on Z C formula, calculated as the instantaneous pressure gradient to peak flow ratio and aortic compliance were estimated by using MRI at 1.5 Tesla. Both Z VA estimates were higher in symptomatic than asymptomatic patients (707 ± 22 versus 579 ± 53 dyne.s/cm 5 , P = 0.031 for Z VA-INS and 4.35 ± 0.16 versus 3.33 ± 0.38 mmHg.m 2 /mL, P = 0.018 for Z VA-TI ). Although they were both associated with aortic compliance (r = -0.45; P = 0.006 for Z VA-INS and r = -0.43; P = 0.008 for Z VA-TI ) only Z VA-INS was associated with E/Ea (r = 0.50; P < 0.001). In multivariate analysis to identify determinants of E/Ea, a model including age, mean blood pressure, LV ejection fraction, LV mass, and aortic valve area was performed (R 2  = 0.41; P < 0.01). When Z VA-INS was added to the model, its overall significance was higher R 2  = 0.56 (P < 0.01) and Z VA-INS and LV mass were the only significant determinants. Z VA-INS was more strongly associated with diastolic dysfunction than usual parameters quantifying AVS severity. This new Z VA estimate could improve LV afterload evaluation. 1 J. Magn. Reson. Imaging 2017;45:795-803. © 2016 International Society for Magnetic Resonance in Medicine.

Validation of the geographic position of EPER-Spain industries

PubMed Central

García-Pérez, Javier; Boldo, Elena; Ramis, Rebeca; Vidal, Enrique; Aragonés, Nuria; Pérez-Gómez, Beatriz; Pollán, Marina; López-Abente, Gonzalo

2008-01-01

Background The European Pollutant Emission Register in Spain (EPER-Spain) is a public inventory of pollutant industries created by decision of the European Union. The location of these industries is geocoded and the first published data correspond to 2001. Publication of these data will allow for quantification of the effect of proximity to one or more such plant on cancer and all-cause mortality observed in nearby towns. However, as errors have been detected in the geocoding of many of the pollutant foci shown in the EPER, it was decided that a validation study should be conducted into the accuracy of these co-ordinates. EPER-Spain geographic co-ordinates were drawn from the European Environment Agency (EEA) server and the Spanish Ministry of the Environment (MOE). The Farm Plot Geographic Information System (Sistema de Información Geográfica de Parcelas Agrícolas) (SIGPAC) enables orthophotos (digitalized aerial images) of any territorial point across Spain to be obtained. Through a search of co-ordinates in the SIGPAC, all the industrial foci (except farms) were located. The quality criteria used to ascertain possible errors in industrial location were high, medium and low quality, where industries were situated at a distance of less than 500 metres, more than 500 metres but less than 1 kilometre, and more than 1 kilometre from their real locations, respectively. Results Insofar as initial registry quality was concerned, 84% of industrial complexes were inaccurately positioned (low quality) according to EEA data versus 60% for Spanish MOE data. The distribution of the distances between the original and corrected co-ordinates for each of the industries on the registry revealed that the median error was 2.55 kilometres for Spain overall (according to EEA data). The Autonomous Regions that displayed most errors in industrial geocoding were Murcia, Canary Islands, Andalusia and Madrid. Correct co-ordinates were successfully allocated to 100% of EPER-Spain industries. Conclusion Knowing the exact location of pollutant foci is vital to obtain reliable and valid conclusions in any study where distance to the focus is a decisive factor, as in the case of the consequences of industrial pollution on the health of neighbouring populations. PMID:18190678

Circumferential suture technique for esophageal transection to treat esophageal variceal bleeding.

PubMed

Jeng, L B; Chen, M F

1993-01-01

The EEA stapler has been used routinely for esophageal transection to treat esophageal variceal bleeding for some time. It carries the risk of postoperative leakage and is not suitable in those cases receiving recent sclerotherapy. The circumferential suture technique presented in this paper can be used in any situation requiring esophageal transection. It has been utilized by us in twenty-two emergent cases with good results.

Calculation of the Moments of Polygons.

DTIC Science & Technology

1987-06-01

2.1) VowUK-1N0+IDIO TUUNTKPlNO.YKNO C Calculate AREA YKXK-YKPIND*IKNO-YKNO*XKP1NO AIKA-hEEA4YKXX C Calculate ACEIT ACENT (1)- ACEIT ( 1) VSUNI4TKIK... ACEIT (2) -ACENT(2) .VSUNYKXK C Calculate SECHON 3ECNON (1) -SCNON( 1) TKXK*(XX~PIdO*VSUNXKKO**2) SECNO(2) -SEn N(2) .yrf* (XKP114*YKP1MO.XKO*YXO+VB1hi

Large and prolonged food-borne multistate hepatitis A outbreak in Europe associated with consumption of frozen berries, 2013 to 2014.

PubMed

Severi, E; Verhoef, L; Thornton, L; Guzman-Herrador, B R; Faber, M; Sundqvist, L; Rimhanen-Finne, R; Roque-Afonso, A M; Ngui, S L; Allerberger, F; Baumann-Popczyk, A; Muller, L; Parmakova, K; Alfonsi, V; Tavoschi, L; Vennema, H; Fitzgerald, M; Myrmel, M; Gertler, M; Ederth, J; Kontio, M; Vanbockstael, C; Mandal, S; Sadkowska-Todys, M; Tosti, M E; Schimmer, B; O Gorman, J; Stene-Johansen, K; Wenzel, J J; Jones, G; Balogun, K; Ciccaglione, A R; O' Connor, L; Vold, L; Takkinen, J; Rizzo, C

2015-07-23

In May 2013, Italy declared a national outbreak of hepatitis A, which also affected several foreign tourists who had recently visited the country. Molecular investigations identified some cases as infected with an identical strain of hepatitis A virus subgenotype IA. After additional European Union/European Economic Area (EU/EEA) countries reported locally acquired and travel-related cases associated with the same outbreak, an international outbreak investigation team was convened, a European outbreak case definition was issued and harmonisation of the national epidemiological and microbiological investigations was encouraged. From January 2013 to August 2014, 1,589 hepatitis A cases were reported associated with the multistate outbreak; 1,102 (70%) of the cases were hospitalised for a median time of six days; two related deaths were reported. Epidemiological and microbiological investigations implicated mixed frozen berries as the vehicle of infection of the outbreak. In order to control the spread of the outbreak, suspected or contaminated food batches were recalled, the public was recommended to heat-treat berries, and post-exposure prophylaxis of contacts was performed. The outbreak highlighted how large food-borne hepatitis A outbreaks may affect the increasingly susceptible EU/EEA general population and how, with the growing international food trade, frozen berries are a potential high-risk food.

Indocyanine Green Fluorescence to Evaluate Nasoseptal Flap Viability in Endoscopic Endonasal Cranial Base Surgery.

PubMed

Kerr, Edward E; Jamshidi, Ali; Carrau, Ricardo L; Campbell, Raewyn G; Filho, Leo F Ditzel; Otto, Bradley A; Prevedello, Daniel M

2017-10-01

Objectives  The pedicled nasoseptal flap (NSF) has dramatically reduced postoperative cerebrospinal fluid leakage following endoscopic endonasal approach (EEA) surgery. Although rare, its arterial supply may be damaged during harvest or may be preoperatively damaged for numerous reasons. Early recognition permits harvesting a contralateral flap before sacrificing its pedicle as part of the surgical exposure or use of an alternative flap. Design  Technical feasibility study and case series. Setting  Tertiary care university-associated medical center. Participants  Five patients requiring an EEA with NSF reconstruction. Main Outcome Measures  During NSF harvest, intravenous indocyanine green (IVICG) was administered, and a customized endoscopic system was used to visualize the emerging fluorescence. At the end of each case, just before final positioning of the NSF, additional IVICG was administered, and the custom endoscope was again introduced to evaluate fluorescence. Results  In four patients, the entire NSF fluoresced brightly with IVICG on initial harvest and before final positioning. One patient showed heterogeneous fluorescence of the pedicle and distal parts of the NSF at both stages. All NSFs healed well without complication. Conclusion  IVICG facilitates real-time evaluation NSF's arterial supply. This may provide early recognition of arterial compromise, allowing the harvest of alternate flaps or modification of surgery.

Endoscopic endonasal approach for craniopharyngioma: the importance of the relationship between pituitary stalk and tumor.

PubMed

Dho, Yun-Sik; Kim, Yong Hwy; Se, Young-Bem; Han, Doo Hee; Kim, Jung Hee; Park, Chul-Kee; Wang, Kyu-Chang; Kim, Dong Gyu

2017-09-29

OBJECTIVE The endoscopic endonasal approach (EEA) is commonly used for the treatment of craniopharyngioma; therefore, it is essential to analyze outcomes in order to understand the benefits and drawbacks. The goal of this paper was to evaluate the clinical features and outcomes associated with this treatment approach. METHODS From July 2010 to March 2016, 82 adult craniopharyngioma patients underwent an EEA at the authors' institution. Of these cases, intraoperative records and immediate postoperative MR images were available for 68 patients. The patients underwent systemized endocrinological evaluation. Eighteen of 68 patients who underwent EEA for recurrence or regrowth of residual lesions after previous surgical management were excluded in the analysis of the anatomical tumor classification. The authors retrospectively analyzed preoperative clinical features and previous anatomical classifications, focusing on the relationship of the pituitary stalk and tumor, to determine predictive factors for the clinical outcome, such as the extent of resection, visual function, endocrinological function, recurrence rate, and complications. RESULTS The mean tumor size was 2.5 cm (3.1 cm for primary tumors and 1.9 cm for recurrent lesions). Gross-total resection (GTR) was achieved in 62 (91.1%) patients (48 [96.0%] patients with primary tumors and 14 [77.8%] patients with recurrent tumors). The rate of GTR was higher in the primary group than in the group with recurrence (p = 0.038). The overall pre- and postoperative visual impairment scale (VIS) scores were 40.8 and 22.1, respectively (50.9 and 14.3 in the primary group and 30.7 and 29.9 in patients with recurrence, respectively). The improvement rate in VIS score was higher in the primary group than in the recurrent group (p = 0.001). Endocrinological function was improved in 4 patients (5.9%) and deteriorated in 32 of 68 patients (47.1%). Tumor invasion into the center of the pituitary stalk affected the postoperative outcomes most significantly. Cognitive dysfunction was observed in 22 patients before surgery and improved in 20 patients (90.9%) after surgery. Hydrocephalus was found in 7 patients and resolved after surgery in all cases. CSF leakage occurred in 2 (2.9%) of 68 patients and was repaired by revision surgery in both patients. Ten patients without CSF leakage also received antibiotics for the treatment of meningitis. The infection rate was higher in the recurrent group. Postoperative endocrinological evaluation showed no deficits in 12 patients and panhypopituitarism in 55 patients. The remaining patient had growth hormone deficiency. Forty-three patients had new-onset diabetes insipidus, and 1 patient had persistent diabetes insipidus after surgery. There were 2 (2.9%) cases of recurrence during the mean 30.7-month follow-up period; one patient underwent radiosurgery and the other underwent reoperation. CONCLUSIONS The EEA resulted in excellent surgical outcomes and acceptable morbidity rates, regardless of the anatomical location of the tumor. Invasion of the craniopharyngioma into the center of the pituitary stalk has strong predictive power for postoperative endocrinological outcome.

Burden of Six Healthcare-Associated Infections on European Population Health: Estimating Incidence-Based Disability-Adjusted Life Years through a Population Prevalence-Based Modelling Study

PubMed Central

Eckmanns, Tim; Abu Sin, Muna; Ducomble, Tanja; Harder, Thomas; Sixtensson, Madlen; Velasco, Edward; Weiß, Bettina; Kramarz, Piotr; Monnet, Dominique L.; Kretzschmar, Mirjam E.; Suetens, Carl

2016-01-01

Background Estimating the burden of healthcare-associated infections (HAIs) compared to other communicable diseases is an ongoing challenge given the need for good quality data on the incidence of these infections and the involved comorbidities. Based on the methodology of the Burden of Communicable Diseases in Europe (BCoDE) project and 2011–2012 data from the European Centre for Disease Prevention and Control (ECDC) point prevalence survey (PPS) of HAIs and antimicrobial use in European acute care hospitals, we estimated the burden of six common HAIs. Methods and Findings The included HAIs were healthcare-associated pneumonia (HAP), healthcare-associated urinary tract infection (HA UTI), surgical site infection (SSI), healthcare-associated Clostridium difficile infection (HA CDI), healthcare-associated neonatal sepsis, and healthcare-associated primary bloodstream infection (HA primary BSI). The burden of these HAIs was measured in disability-adjusted life years (DALYs). Evidence relating to the disease progression pathway of each type of HAI was collected through systematic literature reviews, in order to estimate the risks attributable to HAIs. For each of the six HAIs, gender and age group prevalence from the ECDC PPS was converted into incidence rates by applying the Rhame and Sudderth formula. We adjusted for reduced life expectancy within the hospital population using three severity groups based on McCabe score data from the ECDC PPS. We estimated that 2,609,911 new cases of HAI occur every year in the European Union and European Economic Area (EU/EEA). The cumulative burden of the six HAIs was estimated at 501 DALYs per 100,000 general population each year in EU/EEA. HAP and HA primary BSI were associated with the highest burden and represented more than 60% of the total burden, with 169 and 145 DALYs per 100,000 total population, respectively. HA UTI, SSI, HA CDI, and HA primary BSI ranked as the third to sixth syndromes in terms of burden of disease. HAP and HA primary BSI were associated with the highest burden because of their high severity. The cumulative burden of the six HAIs was higher than the total burden of all other 32 communicable diseases included in the BCoDE 2009–2013 study. The main limitations of the study are the variability in the parameter estimates, in particular the disease models’ case fatalities, and the use of the Rhame and Sudderth formula for estimating incident number of cases from prevalence data. Conclusions We estimated the EU/EEA burden of HAIs in DALYs in 2011–2012 using a transparent and evidence-based approach that allows for combining estimates of morbidity and of mortality in order to compare with other diseases and to inform a comprehensive ranking suitable for prioritization. Our results highlight the high burden of HAIs and the need for increased efforts for their prevention and control. Furthermore, our model should allow for estimations of the potential benefit of preventive measures on the burden of HAIs in the EU/EEA. PMID:27755545

The Effects of Permafrost Thaw on Organic Matter Quality and Availability Along a Hill Slope in Northeastern Siberia

NASA Astrophysics Data System (ADS)

Connolly, C. T.; Spawn, S.; Ludwig, S.; Schade, J. D.; Natali, S.

2014-12-01

Climate warming and permafrost thaw in northeastern Siberia are expected to change the quantity and quality of organic matter (OM) transported through watersheds, releasing previously frozen carbon (C) to biologically available pool. Hill slopes have shown to influence the distribution of OM, resulting in a downhill accumulation of available C and nutrients relative to uphill. Here we examine how future permafrost thaw will change OM quality and availability along a hill slope in a larch-dominated watershed. We collected soils from the thawed organic and mineral layers, and 1m deep permafrost cores for dissolved organic C (DOC) and total dissolved N (TDN), C composition from measures of colored dissolved organic matter (CDOM), DOC lability from biodegradable DOC (BDOC) incubations, C and nutrient availability from extracellular-enzyme assays (EEA's), and microbial respiration from aerobic soil incubations. Here we show that organic soils (O), in comparison to mineral soils (M) and permafrost (P) are the most abundant source of C (avg O DOC: 51.6mg/L), exhibiting low molecular complexity (avg O SUVA254: 4.05) and high quality. Evidence suggests permafrost OM may be an equally abundant, and more labile source of C than mineral soils (highest P DOC: 16.1 mg/L, lowest P SUVA254: 6.32; median M DOC: 18.5 mg/L, median M SUVA254: 24.0). Furthermore, we demonstrate that there may be a positive relationship in the rate of C mineralization and distance downhill, showing 15-30% greater CO2 production/gC downhill relative to uphill. Evidence also supports a similar relationship in permafrost DOC content and molecular complexity, showing more DOC of a lower complexity further downhill. This indicates DOC transport may have been occurring through the active layer and downhill during ice-rich permafrost formation, and may supply a labile source of carbon to lowland areas and adjacent stream networks upon thaw.

Experimental Crack Sealing in Asphaltic Concrete Pavements. Thule Air Base. Greenland, 8-24 August 1960

DTIC Science & Technology

1961-07-01

material prgpsr-ed by A"lied ihnterialls Corp., Stroud., VkI.1 th at is supposed to be soft and more pla-stic in cold cliwmtes. ___b. Para- Plastic ...uimenUsed in Tests .11 heeuiprzent, an-I accesscoia *_ e in the w .rk x; eea iso --router, 4-’- ; eJ. jnt =nd crack ’- ’, ir-power-ed wire brush, spIalt E

Embankment Criteria and Performance Report. Cheyenne River Basin, South Dakota, Red Dale Gulch Area, Cedar Canyon Dam, Rapid City, South Dakota.

DTIC Science & Technology

1983-01-01

to 14.0 percent in the reservoir area. Liquid limits and plastic indexes ranged from 27 to 30 and 7 to 20, respectively, in the spillway area and 29...1 0 t~ EEA e~ c Ea tt , bedoc All elvthoe scton.. refe Itofeet aofe 9𔃾 -v 12 2*, De toeld Ivsit atod aus rae shon oia o adthe field investmation shwn

Epitaxial Garnets and Hexagonal Ferrites.

DTIC Science & Technology

1980-02-28

shaped LPE garnet samples with 31.5um film thickness. We were informed that initial evalu- ation showed acceptably low insertion loss and that the material...frequencies above 25 GHz. c. Furnish up to eight (8) liquid phase epitaxy yttrium iron garnet films to RADC/EEA for testing and evaluation. These tasks...a "Method for Controlling Resonance Frequency of Yttrium Iron Garnet Films ." A patent, "Epitaxial Growth of M-type Hexagonal Ferrite Films on Spinel

EC-92: Catalyst for Change

DTIC Science & Technology

1991-04-02

rules of membership and meet the prevailing democratic and judicial standards of the other member states. 16 While only two years ago this possibility...EES/EEA rules affect everyone, not just the EC, then all participants require a role in formulating and accepting the rules . The EC counters that...18 stabilized; members must accept binding rules for the transfer of monetary pol icy to the new bank .34 The most probable shape of the Eurofed, as

Short-term and long-term effects of protein kinase C on the trafficking and stability of human organic anion transporter 3

PubMed Central

Zhang, Qiang; Suh, Wonmo; Pan, Zui; You, Guofeng

2012-01-01

Human organic anion transporter 3 (hOAT3) belongs to a family of organic anion transporters that play critical roles in the body disposition of numerous clinically important drugs. Therefore, understanding the regulation of this transporter has profound clinical significance. In the current study, we investigated the short-term and long-term regulation of hOAT3 by protein kinase C (PKC). We showed that short-term activation of PKC by phobol 12-Myristate 13-Acetate (PMA) inhibited hOAT3 activity through accelerating its internalization from cell surface to intracellular recycling endosomes. The colocalization of hOAT3 with EEA1-positive recycling endosomes was demonstrated by immunolocalization with confocal microscopy. Furthermore, we showed that long-term activation of PKC resulted in the enhanced degradation of cell surface hOAT3. The pathways for hOAT3 degradation were further examined using proteasomal and lysosomal inhibitors. Our results showed that both proteasomal inhibitors and the lysosomal inhibitors significantly blocked hOAT3 degradation. These results demonstrate that PKC plays critical roles in the trafficking and the stability of hOAT3. PMID:22773962

Tissue Doppler Imaging Combined with Advanced 12-Lead ECG Analysis Might Improve Early Diagnosis of Hypertrophic Cardiomyopathy in Childhood

NASA Technical Reports Server (NTRS)

Femlund, E.; Schlegel, T.; Liuba, P.

2011-01-01

Optimization of early diagnosis of childhood hypertrophic cardiomyopathy (HCM) is essential in lowering the risk of HCM complications. Standard echocardiography (ECHO) has shown to be less sensitive in this regard. In this study, we sought to assess whether spatial QRS-T angle deviation, which has shown to predict HCM in adults with high sensitivity, and myocardial Tissue Doppler Imaging (TDI) could be additional tools in early diagnosis of HCM in childhood. Methods: Children and adolescents with familial HCM (n=10, median age 16, range 5-27 years), and without obvious hypertrophy but with heredity for HCM (n=12, median age 16, range 4-25 years, HCM or sudden death with autopsy-verified HCM in greater than or equal to 1 first-degree relative, HCM-risk) were additionally investigated with TDI and advanced 12-lead ECG analysis using Cardiax(Registered trademark) (IMED Co Ltd, Budapest, Hungary and Houston). Spatial QRS-T angle (SA) was derived from Kors regression-related transformation. Healthy age-matched controls (n=21) were also studied. All participants underwent thorough clinical examination. Results: Spatial QRS-T angle (Figure/ Panel A) and septal E/Ea ratio (Figure/Panel B) were most increased in HCM group as compared to the HCM-risk and control groups (p less than 0.05). Of note, these 2 variables showed a trend toward higher levels in HCM-risk group than in control group (p=0.05 for E/Ea and 0.06 for QRS/T by ANOVA). In a logistic regression model, increased SA and septal E/Ea ratio appeared to significantly predict both the disease (Chi-square in HCM group: 9 and 5, respectively, p less than 0.05 for both) and the risk for HCM (Chi-square in HCM-risk group: 5 and 4 respectively, p less than 0.05 for both), with further increased predictability level when these 2 variables were combined (Chi-square 10 in HCM group, and 7 in HCM-risk group, p less than 0.01 for both). Conclusions: In this small material, Tissue Doppler Imaging and spatial mean QRS-T angle deviation, particularly when combined, appear to be sensitive in predicting the risk for developing childhood HCM. Large-scale, prospective studies are needed to confirm this hypothesis.

Exergy and extended exergy accounting of very large complex systems with an application to the province of Siena, Italy.

PubMed

Sciubba, Enrico; Bastianoni, Simone; Tiezzi, Enzo

2008-01-01

This paper describes the application of exergy and extended exergy analyses to large complex systems. The system to be analysed is assumed to be at steady state, and the input and output fluxes of matter and energy are expressed in units of exergy. Human societies of any reasonable extent are indeed Very Large Complex Systems and can be represented as interconnected networks of N elementary "components", their Subsystems; the detail of the disaggregation depends on the type and quality of the available data. The structural connectivity of the "model" of the System must correctly describe the interactions of each mass or energy flow with each sector of the society: since it is seldom the case that all of these fluxes are available in detail, some preliminary mass- and energy balances must be completed and constitute in fact a part of the initial assumptions. Exergy accounting converts the total amount of resources inflow into their equivalent exergetic form with the help of a table of "raw exergy data" available in the literature. The quantification of each flow on a homogeneous exergetic basis paves the way to the evaluation of the efficiency of each energy and mass transfer between the N sectors and makes it possible to quantify the irreversible losses and identify their sources. The advantage of the EEA, compared to a classical exergy accounting, is the inclusion in the system balance of the exergetic equivalents of three additional "Production Factors": human Labour, Capital and Environmental Remediation costs. EEA has an additional advantage: it allows for the calculation of the efficiency of the domestic sector (impossible to evaluate with any other energy- or exergy-based method) by considering the working hours as its product. As implied in the title, an application of the method was made to a model of the province of Siena (on a year 2000 database): the results show that the sectors of this Province have values of efficiency close to the Italian average, with the exception of the commercial and energy conversion sectors that are more efficient, in agreement with the rather peculiar socio-economic situation of the Province. The largest inefficiency is found to be in the transportation sector, which has an efficiency lower then 30% in EEA and lower than 10% in classical exergy accounting.

The impact of Circulation Weather Types in Urban Air Quality in Portugal

NASA Astrophysics Data System (ADS)

Russo, Ana; Trigo, Ricardo

2013-04-01

It is now clear that emissions of the main air pollutants in Europe have declined significantly in recent decades (EEA, 2011). Nevertheless, many European countries (including Portugal) do not expect to comply with one (or more) pollutant emission ceilings and to air quality limit values, especially for particulate matter (PM), ground level ozone (O3) and nitrogen dioxide (NO2) (EEA, 2011). Consequently, and considering that air pollution (AP) plays a role as a major cause of human mortality and morbidity, exposure to pollutants remains a key environment-related health concern (EEA 2010). Thus, and to comply with the new limits, new strategies must be applied for air quality management. The main objective of this work is to present an objective classification of pre-defined and widely used CWTs affecting Portugal and, based on the most relevant patterns, provide a framework that is useful to characterise the occurrence of pollution episodes, namely its inter-annual and intra-annual variability, as well as the occurrence of extreme events. CWTs were determined using the simple Geostrophic approximation according to the methodology proposed by Trigo and DaCamara (2000). The interannual variability of the resulting CWTs was determined for the period with AP data (2002-2010) and the number of days for each CWT and season for the same period was accounted for. During this period, the most frequent CWTs were found to be the anticyclonic (A), the north (N) and the northeast (NE) types, accounting respectively for 34.7%, 10.9% and 14% of the days. However, higher-than average episodes tend to occur associated predominantly with situations characterized by a few less frequent CWTs, namely easterly (E), northeasterly (NE) and southeasterly (SE) types (that together contributed to less than one fourth of all observed days), are the ones which are associated to higher median and maximum concentrations of the three pollutants. Results obtained highlight the existence of strong links between the interannual variability of daily air quality and interannual variability of CWTs. Additionally, three specific extreme episodes were assessed in more detail including a comparison with results obtained with the HYSPLIT system model. In general, all the pollutants' extreme events occur associated predominantly with situations characterized by an Eastern component and advection of dry air masses.

Training model for control of an internal carotid artery injury during transsphenoidal surgery.

PubMed

Muto, Jun; Carrau, Ricardo L; Oyama, Kenichi; Otto, Brad A; Prevedello, Daniel M

2017-01-01

As the adoption of endoscopic endonasal approaches (EEA) continues to proliferate, increasing numbers of internal carotid artery (ICA) injuries are reported. The objective of this study was to develop a synthetic ICA injury-training model that could mimic this clinical scenario and be portable, repeatable, reproducible, and without risk of biological contamination. Based on computed tomography of a human head, we constructed a synthetic model using selective laser sintering with polyamide nylon and glass beads. Subsequently, the model was connected to a pulsatile pump using 6-mm silicon tubing. The pump maintains a pulsatile flow of an artificial blood-like fluid at a variable pressure to simulate heart beats. Volunteer surgeons with different levels of training and experience were provided simulation training sessions with the models. Pre- and posttraining questionnaires were completed by each of the participants. Pre- and posttraining questionnaires suggest that repeated simulation sessions improve the surgical skills and self-confidence of trainees. This ICA injury model is portable; reproducible; and avoids ethical, biohazard, religious, and legal problems associated with cadaveric models. A synthetic ICA injury model for EEA allows recurring training that may improve the surgeon's ability to maintain endoscopic visualization, control catastrophic bleeding, decrease psychomotor stress, and develop effective team strategies to achieve hemostasis. NA Laryngoscope, 127:38-43, 2017. © 2016 The American Laryngological, Rhinological and Otological Society, Inc.

Green Infrastructure Design Based on Spatial Conservation Prioritization and Modeling of Biodiversity Features and Ecosystem Services.

PubMed

Snäll, Tord; Lehtomäki, Joona; Arponen, Anni; Elith, Jane; Moilanen, Atte

2016-02-01

There is high-level political support for the use of green infrastructure (GI) across Europe, to maintain viable populations and to provide ecosystem services (ES). Even though GI is inherently a spatial concept, the modern tools for spatial planning have not been recognized, such as in the recent European Environment Agency (EEA) report. We outline a toolbox of methods useful for GI design that explicitly accounts for biodiversity and ES. Data on species occurrence, habitats, and environmental variables are increasingly available via open-access internet platforms. Such data can be synthesized by statistical species distribution modeling, producing maps of biodiversity features. These, together with maps of ES, can form the basis for GI design. We argue that spatial conservation prioritization (SCP) methods are effective tools for GI design, as the overall SCP goal is cost-effective allocation of conservation efforts. Corridors are currently promoted by the EEA as the means for implementing GI design, but they typically target the needs of only a subset of the regional species pool. SCP methods would help to ensure that GI provides a balanced solution for the requirements of many biodiversity features (e.g., species, habitat types) and ES simultaneously in a cost-effective manner. Such tools are necessary to make GI into an operational concept for combating biodiversity loss and promoting ES.

Spatial Relationships between Markers for Secretory and Endosomal Machinery in Human Cytomegalovirus-Infected Cells versus Those in Uninfected Cells▿†

PubMed Central

Das, Subhendu; Pellett, Philip E.

2011-01-01

Human cytomegalovirus (HCMV) induces extensive remodeling of the secretory apparatus to form the cytoplasmic virion assembly compartment (cVAC), where virion tegumentation and envelopment take place. We studied the structure of the cVAC by confocal microscopy to assess the three-dimensional distribution of proteins specifically associated with individual secretory organelles. In infected cells, early endosome antigen 1 (EEA1)-positive vesicles are concentrated at the center of the cVAC and, as previously seen, are distinct from structures visualized by markers for the endoplasmic reticulum, Golgi apparatus, and trans-Golgi network (TGN). EEA1-positive vesicles can be strongly associated with markers for recycling endosomes, to a lesser extent with markers associated with components of the endosomal sorting complex required for transport III (ESCRT III) machinery, and then with markers of late endosomes. In comparisons of uninfected and infected cells, we found significant changes in the structural associations and colocalization of organelle markers, as well as in net organelle volumes. These results provide new evidence that the HCMV-induced remodeling of the membrane transport apparatus involves much more than simple relocation and expansion of preexisting structures and are consistent with the hypothesis that the shift in identity of secretory organelles in HCMV-infected cells results in new functional profiles. PMID:21471245

Long-term trends in oil and gas discovery rates in lower 48 United States

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

Woods, T.J.

1985-09-01

The Gas Research Institute (GRI), in association with Energy and Environmental Analysis, Inc. (EEA), has developed a data base characterizing the discovered oil and gas fields in the lower 48 United States. The number of fields in this data base reported to have been discovered since 1947 substantially exceeds the count presented in the AAPG survey of new-field discoveries since 1947. The greatest relative difference between the field counts is for fields larger than 10 million bbl of oil equivalent (BOE) (AAPG Class C fields or larger). Two factors contribute to the difference in reported discoveries by field size. First,more » the AAPG survey does not capture all new-field discoveries, particularly in the offshore. Second, the AAPG survey does not update field sizes past 6 years after the field discovery date. Because of reserve appreciation to discovered fields, discovery-trend data based on field-size data should be used with caution, particularly when field-size estimates have not been updated for a substantial period of time. Based on the GRI/EEA data base, the major decline in the discovery rates of large, new oil and gas fields in the lower 48 United States appears to have ended by the early 1960s. Since then, discovery rates seem to have improved. Thus, the outlook for future discoveries of large fields may be much better than previously believed.« less

Green Infrastructure Design Based on Spatial Conservation Prioritization and Modeling of Biodiversity Features and Ecosystem Services

NASA Astrophysics Data System (ADS)

Snäll, Tord; Lehtomäki, Joona; Arponen, Anni; Elith, Jane; Moilanen, Atte

2016-02-01

There is high-level political support for the use of green infrastructure (GI) across Europe, to maintain viable populations and to provide ecosystem services (ES). Even though GI is inherently a spatial concept, the modern tools for spatial planning have not been recognized, such as in the recent European Environment Agency (EEA) report. We outline a toolbox of methods useful for GI design that explicitly accounts for biodiversity and ES. Data on species occurrence, habitats, and environmental variables are increasingly available via open-access internet platforms. Such data can be synthesized by statistical species distribution modeling, producing maps of biodiversity features. These, together with maps of ES, can form the basis for GI design. We argue that spatial conservation prioritization (SCP) methods are effective tools for GI design, as the overall SCP goal is cost-effective allocation of conservation efforts. Corridors are currently promoted by the EEA as the means for implementing GI design, but they typically target the needs of only a subset of the regional species pool. SCP methods would help to ensure that GI provides a balanced solution for the requirements of many biodiversity features (e.g., species, habitat types) and ES simultaneously in a cost-effective manner. Such tools are necessary to make GI into an operational concept for combating biodiversity loss and promoting ES.

Installation Restoration Program Phase 2. Confirmation/Quantification. Stage 1. Volume 1.

DTIC Science & Technology

1986-10-10

FIGUR 1- EEA LNO EFIG I AiNLanfl GUARD BASESHOWING PHSE II INVTIGTOIE I 1- ........ Site No. - I MAA Site No.Northwest Landfill TS Site.No.4 West...Protective Steel PVC Cap Flush Mount in Casing Plastic Nelson Valve Box Land Surface PVC Cap . Cement Grout 0 L n aBentonite Pellet 4" Diameter Seal...12.0 feet BLS. Clays of high plasticity predominate in all three wells. No sand lenses were encountered and the topsoil at all three well locations is

Fracture Damage and Failure of Cannon Components by Service Loading

DTIC Science & Technology

1983-02-01

the result of normal service :onALtiolt. )etaits of the failure and the redesign of the cannon h3’e eea iLOs’rtbed elsewhere.| , The brief review...here is intenlded to ie.,Acrlbe the extreme situation of very severe damage and failure of a cannon. In fact, this failure led to many fracture- safe ...criterion; elastic-perfectly plastic material properties. The experiments summarized in Figure 6 used cannon tubes in which a 6.4 mm deep semi

[Creation of a colostomy using a circular mechanical stapler].

PubMed

Ruscalla, L; Delemont, M; Ligresti, C; Farinella, M; Rossi, R

1991-09-15

The paper describes the method used to create a preternatural anus in terminal stomas using a mechanical circular stapler (Model EEA-31). Two methods are put forward: Chung's and Burke's methods (the latter of which was used by our department). A mechanical circular stapler has been used several times (13) to perform this type of stoma, with excellent esthetic and functional results, both immediate and long-term. It was only necessary to reoperate in one case in order to suspend the affected colic loop.

Enzyme activity assay of glycoprotein enzymes based on a boronate affinity molecularly imprinted 96-well microplate.

PubMed

Bi, Xiaodong; Liu, Zhen

2014-12-16

Enzyme activity assay is an important method in clinical diagnostics. However, conventional enzyme activity assay suffers from apparent interference from the sample matrix. Herein, we present a new format of enzyme activity assay that can effectively eliminate the effects of the sample matrix. The key is a 96-well microplate modified with molecularly imprinted polymer (MIP) prepared according to a newly proposed method called boronate affinity-based oriented surface imprinting. Alkaline phosphatase (ALP), a glycoprotein enzyme that has been routinely used as an indicator for several diseases in clinical tests, was taken as a representative target enzyme. The prepared MIP exhibited strong affinity toward the template enzyme (with a dissociation constant of 10(-10) M) as well as superb tolerance for interference. Thus, the enzyme molecules in a complicated sample matrix could be specifically captured and cleaned up for enzyme activity assay, which eliminated the interference from the sample matrix. On the other hand, because the boronate affinity MIP could well retain the enzymatic activity of glycoprotein enzymes, the enzyme captured by the MIP was directly used for activity assay. Thus, additional assay time and possible enzyme or activity loss due to an enzyme release step required by other methods were avoided. Assay of ALP in human serum was successfully demonstrated, suggesting a promising prospect of the proposed method in real-world applications.

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.

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.

Finding Biomass Degrading Enzymes Through an Activity-Correlated Quantitative Proteomics Platform (ACPP).

PubMed

Ma, Hongyan; Delafield, Daniel G; Wang, Zhe; You, Jianlan; Wu, Si

2017-04-01

The microbial secretome, known as a pool of biomass (i.e., plant-based materials) degrading enzymes, can be utilized to discover industrial enzyme candidates for biofuel production. Proteomics approaches have been applied to discover novel enzyme candidates through comparing protein expression profiles with enzyme activity of the whole secretome under different growth conditions. However, the activity measurement of each enzyme candidate is needed for confident "active" enzyme assignments, which remains to be elucidated. To address this challenge, we have developed an Activity-Correlated Quantitative Proteomics Platform (ACPP) that systematically correlates protein-level enzymatic activity patterns and protein elution profiles using a label-free quantitative proteomics approach. The ACPP optimized a high performance anion exchange separation for efficiently fractionating complex protein samples while preserving enzymatic activities. The detected enzymatic activity patterns in sequential fractions using microplate-based assays were cross-correlated with protein elution profiles using a customized pattern-matching algorithm with a correlation R-score. The ACPP has been successfully applied to the identification of two types of "active" biomass-degrading enzymes (i.e., starch hydrolysis enzymes and cellulose hydrolysis enzymes) from Aspergillus niger secretome in a multiplexed fashion. By determining protein elution profiles of 156 proteins in A. niger secretome, we confidently identified the 1,4-α-glucosidase as the major "active" starch hydrolysis enzyme (R = 0.96) and the endoglucanase as the major "active" cellulose hydrolysis enzyme (R = 0.97). The results demonstrated that the ACPP facilitated the discovery of bioactive enzymes from complex protein samples in a high-throughput, multiplexing, and untargeted fashion. Graphical Abstract ᅟ.

Finding Biomass Degrading Enzymes Through an Activity-Correlated Quantitative Proteomics Platform (ACPP)

NASA Astrophysics Data System (ADS)

Ma, Hongyan; Delafield, Daniel G.; Wang, Zhe; You, Jianlan; Wu, Si

2017-04-01

The microbial secretome, known as a pool of biomass (i.e., plant-based materials) degrading enzymes, can be utilized to discover industrial enzyme candidates for biofuel production. Proteomics approaches have been applied to discover novel enzyme candidates through comparing protein expression profiles with enzyme activity of the whole secretome under different growth conditions. However, the activity measurement of each enzyme candidate is needed for confident "active" enzyme assignments, which remains to be elucidated. To address this challenge, we have developed an Activity-Correlated Quantitative Proteomics Platform (ACPP) that systematically correlates protein-level enzymatic activity patterns and protein elution profiles using a label-free quantitative proteomics approach. The ACPP optimized a high performance anion exchange separation for efficiently fractionating complex protein samples while preserving enzymatic activities. The detected enzymatic activity patterns in sequential fractions using microplate-based assays were cross-correlated with protein elution profiles using a customized pattern-matching algorithm with a correlation R-score. The ACPP has been successfully applied to the identification of two types of "active" biomass-degrading enzymes (i.e., starch hydrolysis enzymes and cellulose hydrolysis enzymes) from Aspergillus niger secretome in a multiplexed fashion. By determining protein elution profiles of 156 proteins in A. niger secretome, we confidently identified the 1,4-α-glucosidase as the major "active" starch hydrolysis enzyme (R = 0.96) and the endoglucanase as the major "active" cellulose hydrolysis enzyme (R = 0.97). The results demonstrated that the ACPP facilitated the discovery of bioactive enzymes from complex protein samples in a high-throughput, multiplexing, and untargeted fashion.

Co-immobilization of multiple enzymes by metal coordinated nucleotide hydrogel nanofibers: improved stability and an enzyme cascade for glucose detection

NASA Astrophysics Data System (ADS)

Liang, Hao; Jiang, Shuhui; Yuan, Qipeng; Li, Guofeng; Wang, Feng; Zhang, Zijie; Liu, Juewen

2016-03-01

Preserving enzyme activity and promoting synergistic activity via co-localization of multiple enzymes are key topics in bionanotechnology, materials science, and analytical chemistry. This study reports a facile method for co-immobilizing multiple enzymes in metal coordinated hydrogel nanofibers. Specifically, four types of protein enzymes, including glucose oxidase, Candida rugosa lipase, α-amylase, and horseradish peroxidase, were respectively encapsulated in a gel nanofiber made of Zn2+ and adenosine monophosphate (AMP) with a simple mixing step. Most enzymes achieved quantitative loading and retained full activity. At the same time, the entrapped enzymes were more stable against temperature variation (by 7.5 °C), protease attack, extreme pH (by 2-fold), and organic solvents. After storing for 15 days, the entrapped enzyme still retained 70% activity while the free enzyme nearly completely lost its activity. Compared to nanoparticles formed with AMP and lanthanide ions, the nanofiber gels allowed much higher enzyme activity. Finally, a highly sensitive and selective biosensor for glucose was prepared using the gel nanofiber to co-immobilize glucose oxidase and horseradish peroxidase for an enzyme cascade system. A detection limit of 0.3 μM glucose with excellent selectivity was achieved. This work indicates that metal coordinated materials using nucleotides are highly useful for interfacing with biomolecules.Preserving enzyme activity and promoting synergistic activity via co-localization of multiple enzymes are key topics in bionanotechnology, materials science, and analytical chemistry. This study reports a facile method for co-immobilizing multiple enzymes in metal coordinated hydrogel nanofibers. Specifically, four types of protein enzymes, including glucose oxidase, Candida rugosa lipase, α-amylase, and horseradish peroxidase, were respectively encapsulated in a gel nanofiber made of Zn2+ and adenosine monophosphate (AMP) with a simple mixing step. Most enzymes achieved quantitative loading and retained full activity. At the same time, the entrapped enzymes were more stable against temperature variation (by 7.5 °C), protease attack, extreme pH (by 2-fold), and organic solvents. After storing for 15 days, the entrapped enzyme still retained 70% activity while the free enzyme nearly completely lost its activity. Compared to nanoparticles formed with AMP and lanthanide ions, the nanofiber gels allowed much higher enzyme activity. Finally, a highly sensitive and selective biosensor for glucose was prepared using the gel nanofiber to co-immobilize glucose oxidase and horseradish peroxidase for an enzyme cascade system. A detection limit of 0.3 μM glucose with excellent selectivity was achieved. This work indicates that metal coordinated materials using nucleotides are highly useful for interfacing with biomolecules. Electronic supplementary information (ESI) available: Additional methods, IR and XRD spectroscopy, enzyme loading capacity, enzyme kinetic parameters, and enzyme stability data. See DOI: 10.1039/c5nr08734a

County-scale spatial distribution of soil enzyme activities and enzyme activity indices in agricultural land: implications for soil quality assessment.

PubMed

Tan, Xiangping; Xie, Baoni; Wang, Junxing; He, Wenxiang; Wang, Xudong; Wei, Gehong

2014-01-01

Here the spatial distribution of soil enzymatic properties in agricultural land was evaluated on a county-wide (567 km(2)) scale in Changwu, Shaanxi Province, China. The spatial variations in activities of five hydrolytic enzymes were examined using geostatistical methods. The relationships between soil enzyme activities and other soil properties were evaluated using both an integrated total enzyme activity index (TEI) and the geometric mean of enzyme activities (GME). At the county scale, soil invertase, phosphatase, and catalase activities were moderately spatially correlated, whereas urease and dehydrogenase activities were weakly spatially correlated. Correlation analysis showed that both TEI and GME were better correlated with selected soil physicochemical properties than single enzyme activities. Multivariate regression analysis showed that soil OM content had the strongest positive effect while soil pH had a negative effect on the two enzyme activity indices. In addition, total phosphorous content had a positive effect on TEI and GME in orchard soils, whereas alkali-hydrolyzable nitrogen and available potassium contents, respectively, had negative and positive effects on these two enzyme indices in cropland soils. The results indicate that land use changes strongly affect soil enzyme activities in agricultural land, where TEI provides a sensitive biological indicator for soil quality.

Glycyl radical activating enzymes: Structure, mechanism, and substrate interactions☆

PubMed Central

Shisler, Krista A.; Broderick, Joan B.

2014-01-01

The glycyl radical enzyme activating enzymes (GRE–AEs) are a group of enzymes that belong to the radical S-adenosylmethionine (SAM) superfamily and utilize a [4Fe–4S] cluster and SAM to catalyze H-atom abstraction from their substrate proteins. GRE–AEs activate homodimeric proteins known as glycyl radical enzymes (GREs) through the production of a glycyl radical. After activation, these GREs catalyze diverse reactions through the production of their own substrate radicals. The GRE–AE pyruvate formate lyase activating enzyme (PFL-AE) is extensively characterized and has provided insights into the active site structure of radical SAM enzymes including GRE–AEs, illustrating the nature of the interactions with their corresponding substrate GREs and external electron donors. This review will highlight research on PFL-AE and will also discuss a few GREs and their respective activating enzymes. PMID:24486374

[Interaction between CYP450 enzymes and metabolism of traditional Chinese medicine as well as enzyme activity assay].

PubMed

Lu, Tu-lin; Su, Lian-lin; Ji, De; Gu, Wei; Mao, Chun-qin

2015-09-01

Drugs are exogenous compounds for human bodies, and will be metabolized by many enzymes after administration. CYP450 enzyme, as a major metabolic enzyme, is an important phase I drug metabolizing enzyme. In human bodies, about 75% of drug metabolism is conducted by CYP450 enzymes, and CYP450 enzymes is the key factor for drug interactions between traditional Chinese medicine( TCM) -TCM, TCM-medicine and other drug combination. In order to make clear the interaction between metabolic enzymes and TCM metabolism, we generally chose the enzymatic activity as an evaluation index. That is to say, the enhancement or reduction of CYP450 enzyme activity was used to infer the inducing or inhibitory effect of active ingredients and extracts of traditional Chinese medicine on enzymes. At present, the common method for measuring metabolic enzyme activity is Cocktail probe drugs, and it is the key to select the suitable probe substrates. This is of great significance for study drug's absorption, distribution, metabolism and excretion (ADME) process in organisms. The study focuses on the interaction between TCMs, active ingredients, herbal extracts, cocktail probe substrates as well as CYP450 enzymes, in order to guide future studies.

A sorting nexin 17-binding domain within the LRP1 cytoplasmic tail mediates receptor recycling through the basolateral sorting endosome

PubMed Central

Farfán, Pamela; Lee, Jiyeon; Larios, Jorge; Sotelo, Pablo; Bu, Guojun; Marzolo, María-Paz

2013-01-01

Sorting nexin 17 (SNX17) is an adaptor protein present in EEA1-positive sorting endosomes that promotes the efficient recycling of low-density lipoprotein receptor-related protein 1 (LRP1) to the plasma membrane through recognition of the first NPxY motif in the cytoplasmic tail of this receptor. The interaction of LRP1 with SNX17 also regulates the basolateral recycling of the receptor from the basolateral sorting endosome (BSE). In contrast, megalin, which is apically distributed in polarized epithelial cells and localizes poorly to EEA1-positive sorting endosomes, does not interact with SNX17, despite containing three NPxY motifs, indicating that this motif is not sufficient for receptor recognition by SNX17. Here, we identified a cluster of 32 amino acids within the cytoplasmic domain of LRP1 that is both necessary and sufficient for SNX17 binding. To delineate the function of this SNX17-binding domain, we generated chimeric proteins in which the SNX17-binding domain was inserted into the cytoplasmic tail of megalin. This insertion mediated the binding of megalin to SNX17 and modified the cell surface expression and recycling of megalin in non-polarized cells. However, the polarized localization of chimeric megalin was not modified in polarized MDCK cells. These results provide evidence regarding the molecular and cellular mechanisms underlying the specificity of SNX17-binding receptors and the restricted function of SNX17 in the BSE. PMID:23593972

[Experimental model for the examination of inner pressure tolerance of telescopic anastomosis and other frequently performed anastomosis types of the esophagus].

PubMed

Szúcs, G; Tóth, I; Bráth, E; Gyáni, K; Miko, I

2001-08-01

We have good results with telescopic anastomosis technique in partial oesophagectomies and gastrectomies. As we could not find data about the healing process of telescopic anastomoses so we started experimenting. Inside pressure tolerance was examined immediately after performing anastomoses by measuring the bursting pressure using the organs of pigs slaughtered in the meat industry. Both oesophago-gastrostomies and oesophago-jejunostomies were performed with telescopic, single layer interrupted, single layer continuous, double layer interrupted and double layer continuous-interrupted technique, 9 of each anastomosis. A series of oesophago-jejunostomies were performed with EEA stapler. 99 anastomoses of 11 types were investigated. We found, that the inner pressure tolerance of telescopic oesophago-gastrostomy is better than any other single layer type variant. On the other hand the double layer type variants have much better pressure tolerance than the telescopic and other two type single layer anastomoses. The difference is statistically significant. In oesophago-jejunostomies the pressure tolerance of telescopic anastomosis is better than of the single layer interrupted type but the difference between the telescopic and single layer continuous type anastomoses is not significant. The pressure tolerance of double layer anastomosis is higher than the telescopic one but the difference is significant only in the continuous-interrupted type. The inner pressure tolerance of telescopic and EEA stapler anastomoses are equal. The investigation of additional features in anastomosis healing is in progress.

Distribution of enzyme activity hotspots induced by earthworms in top- and subsoil

NASA Astrophysics Data System (ADS)

Hoang, D. T. T.

2016-12-01

Earthworms (Lumbricus terrestris L.) not only affect soil physics, but they also boost microbial activities and consequently create important hotspots of microbial mediated carbon and nutrient turnover through their burrowing activity. However, it is still unknown to which extend earthworms change the enzyme distribution and activity inside their burrows in top- and subsoil horizons. We hypothesized that earthworm burrows, which are enriched in available substrates, have higher percentage of enzyme activity hotspots than soil without earthworms, and that enzyme activities decreased with increasing depth because of the increasing recalcitrance of organic matter in subsoil. We visualized enzyme distribution inside and outside of worm burrows (biopores) by in situ soil zymography and measured enzyme kinetics of 6 enzymes - β-glucosidase (GLU), cellobiohydrolase (CBH), xylanase (XYL), chitinase (NAG), leucine aminopeptidase (LAP) and acid phosphatase (APT) - in pore and bulk soil material up to 105 cm. Zymography showed a heterogeneous distribution of hotspots in worm burrows. The hotspot areas was 2.4 to 14 times larger in the burrows than in soil without earthworms. However, the dispersion index of hotspot distribution showed more aggregated hotspots in soil without earthworms than in soil with earthworms and burrow wall. Enzyme activities decreased with depth, by a factor of 2 to 8 due to fresh C input from the soil surface. Compared to bulk soil, enzyme activities in topsoil biopores were up to 11 times higher for all enzymes, but in the subsoil activities of XYL, NAG and APT were lower in earthworm biopores than bulk soil. In conclusion, hotspots were twice as concentrated close to earthworm burrows as in surrounding soil. Earthworms exerted stronger effects on enzyme activities in biopores in the topsoil than in subsoil. Keywords: Earthworms, hotspots, enzyme activities, enzyme distribution, subsoil

Glycyl radical activating enzymes: structure, mechanism, and substrate interactions.

PubMed

Shisler, Krista A; Broderick, Joan B

2014-03-15

The glycyl radical enzyme activating enzymes (GRE-AEs) are a group of enzymes that belong to the radical S-adenosylmethionine (SAM) superfamily and utilize a [4Fe-4S] cluster and SAM to catalyze H-atom abstraction from their substrate proteins. GRE-AEs activate homodimeric proteins known as glycyl radical enzymes (GREs) through the production of a glycyl radical. After activation, these GREs catalyze diverse reactions through the production of their own substrate radicals. The GRE-AE pyruvate formate lyase activating enzyme (PFL-AE) is extensively characterized and has provided insights into the active site structure of radical SAM enzymes including GRE-AEs, illustrating the nature of the interactions with their corresponding substrate GREs and external electron donors. This review will highlight research on PFL-AE and will also discuss a few GREs and their respective activating enzymes. Copyright © 2014. Published by Elsevier Inc.

Purification and characterization of the enzyme cholesterol oxidase from a new isolate of Streptomyces sp.

PubMed

Praveen, Vandana; Srivastava, Akanksha; Tripathi, C K M

2011-11-01

An extracellular cholesterol oxidase (cho) enzyme was isolated from the Streptomyces parvus, a new source and purified 18-fold by ion exchange and gel filtration chromatography. Specific activity of the purified enzyme was found to be 20 U/mg with a 55 kDa molecular mass. The enzyme was stable at pH 7.2 and 50 °C. The enzyme activity was inhibited in the presence of Pb(2+), Ag(2+), Hg(2+), and Zn(2+) and enhanced in the presence of Mn(2+). The enzyme activity was inhibited by the thiol-reducing reagents (DTT, β-mercaptoethanol), suggesting that disulfide linkage is essential for the enzyme activity. The enzyme activity was found to be maximum in the presence of Triton X-100 and X-114 detergents whereas sodium dodecyl sulfate fully inactivated the enzyme. The enzyme showed moderate stability towards all organic solvents except acetone, benzene, chloroform and the activity increased in the presence of isopropanol and ethanol. The K(m) value for the oxidation of cholesterol by this enzyme was 0.02 mM.

Spatial distribution of enzyme activities along the root and in the rhizosphere of different plants

NASA Astrophysics Data System (ADS)

Razavi, Bahar S.; Zarebanadkouki, Mohsen; Blagodatskaya, Evgenia; Kuzyakov, Yakov

2015-04-01

Extracellular enzymes are important for decomposition of many biological macromolecules abundant in soil such as cellulose, hemicelluloses and proteins. Activities of enzymes produced by both plant roots and microbes are the primary biological drivers of organic matter decomposition and nutrient cycling. So far acquisition of in situ data about local activity of different enzymes in soil has been challenged. That is why there is an urgent need in spatially explicit methods such as 2-D zymography to determine the variation of enzymes along the roots in different plants. Here, we developed further the zymography technique in order to quantitatively visualize the enzyme activities (Spohn and Kuzyakov, 2013), with a better spatial resolution We grew Maize (Zea mays L.) and Lentil (Lens culinaris) in rhizoboxes under optimum conditions for 21 days to study spatial distribution of enzyme activity in soil and along roots. We visualized the 2D distribution of the activity of three enzymes:β-glucosidase, leucine amino peptidase and phosphatase, using fluorogenically labelled substrates. Spatial resolution of fluorescent images was improved by direct application of a substrate saturated membrane to the soil-root system. The newly-developed direct zymography shows different pattern of spatial distribution of enzyme activity along roots and soil of different plants. We observed a uniform distribution of enzyme activities along the root system of Lentil. However, root system of Maize demonstrated inhomogeneity of enzyme activities. The apical part of an individual root (root tip) in maize showed the highest activity. The activity of all enzymes was the highest at vicinity of the roots and it decreased towards the bulk soil. Spatial patterns of enzyme activities as a function of distance from the root surface were enzyme specific, with highest extension for phosphatase. We conclude that improved zymography is promising in situ technique to analyze, visualize and quantify spatial distribution of enzyme activities in the rhizosphere hotspots. References Spohn, M., Kuzyakov, Y., 2013. Phosphorus mineralization can be driven by microbial need for carbon. Soil Biology & Biochemistry 61: 69-75

Erythrocyte enzymes in sheep: comparison of activity in fetal, newborn, maternal and nonpregnant ewe erythrocytes.

PubMed

Noble, N A; Cabalum, T C; Nathanielsz, P W; Tanaka, K R

1982-01-01

Hematological data and the activities of 21 red cell enzymes were measured in 8 nonpregnant ewes, 13 chronically catheterized fetuses at 125-135 days of gestation, and 8 of their mothers. In addition, 7 lambs were followed from birth to 17 days of age. Fetal sheep red cells have dramatically increased activities for 17 of 21 enzymes measured compared with adult nonpregnant ewes. The pattern of decline of enzyme activities with development varies considerably among enzymes. The activity of seven enzymes showed an orderly decline from fetal to adult life. For seven enzymes very little or no decline in activity was observed between 125 and 135 days of gestation and birth. Pyruvate kinase activity declined to adult levels by birth. Phosphoglucose isomerase and nucleoside phosphorylase activity increased, and glutathione peroxidase activity decreased in newborn lamb red cells compared to fetal cells. Differences in blood cell variables were also found among these groups.

Process for preparing multilayer enzyme coating on a fiber

DOEpatents

Kim, Jungbae [Richland, WA; Kwak, Ja Hun [Richland, WA; Grate, Jay W [West Richland, WA

2009-11-03

A process for preparing high stability, high activity biocatalytic materials is disclosed and processes for using the same. The process involves coating of a material or fiber with enzymes and enzyme aggregate providing a material or fiber with high biocatalytic activity and stability useful in heterogeneous environments. In one illustrative approach, enzyme "seeds" are covalently attached to polymer nanofibers followed by treatment with a reagent that crosslinks additional enzyme molecules to the seed enzymes forming enzyme aggregates thereby improving biocatalytic activity due to increased enzyme loading and enzyme stability. This approach creates a useful new biocatalytic immobilized enzyme system with potential applications in bioconversion, bioremediation, biosensors, and biofuel cells.

In Situ Enzyme Activity in the Dissolved and Particulate Fraction of the Fluid from Four Pitcher Plant Species of the Genus Nepenthes

PubMed Central

Takeuchi, Yayoi; Salcher, Michaela M.; Ushio, Masayuki; Shimizu-Inatsugi, Rie; Kobayashi, Masaki J.; Diway, Bibian; von Mering, Christian; Pernthaler, Jakob; Shimizu, Kentaro K.

2011-01-01

The genus Nepenthes, a carnivorous plant, has a pitcher to trap insects and digest them in the contained fluid to gain nutrient. A distinctive character of the pitcher fluid is the digestive enzyme activity that may be derived from plants and dwelling microbes. However, little is known about in situ digestive enzymes in the fluid. Here we examined the pitcher fluid from four species of Nepenthes. High bacterial density was observed within the fluids, ranging from 7×106 to 2.2×108 cells ml−1. We measured the activity of three common enzymes in the fluid: acid phosphatases, β-d-glucosidases, and β-d-glucosaminidases. All the tested enzymes detected in the liquid of all the pitcher species showed activity that considerably exceeded that observed in aquatic environments such as freshwater, seawater, and sediment. Our results indicate that high enzyme activity within a pitcher could assist in the rapid decomposition of prey to maximize efficient nutrient use. In addition, we filtered the fluid to distinguish between dissolved enzyme activity and particle-bound activity. As a result, filtration treatment significantly decreased the activity in all enzymes, while pH value and Nepenthes species did not affect the enzyme activity. It suggested that enzymes bound to bacteria and other organic particles also would significantly contribute to the total enzyme activity of the fluid. Since organic particles are themselves usually colonized by attached and highly active bacteria, it is possible that microbe-derived enzymes also play an important role in nutrient recycling within the fluid and affect the metabolism of the Nepenthes pitcher plant. PMID:21949872

In situ enzyme activity in the dissolved and particulate fraction of the fluid from four pitcher plant species of the genus Nepenthes.

PubMed

Takeuchi, Yayoi; Salcher, Michaela M; Ushio, Masayuki; Shimizu-Inatsugi, Rie; Kobayashi, Masaki J; Diway, Bibian; von Mering, Christian; Pernthaler, Jakob; Shimizu, Kentaro K

2011-01-01

The genus Nepenthes, a carnivorous plant, has a pitcher to trap insects and digest them in the contained fluid to gain nutrient. A distinctive character of the pitcher fluid is the digestive enzyme activity that may be derived from plants and dwelling microbes. However, little is known about in situ digestive enzymes in the fluid. Here we examined the pitcher fluid from four species of Nepenthes. High bacterial density was observed within the fluids, ranging from 7×10(6) to 2.2×10(8) cells ml(-1). We measured the activity of three common enzymes in the fluid: acid phosphatases, β-D-glucosidases, and β-D-glucosaminidases. All the tested enzymes detected in the liquid of all the pitcher species showed activity that considerably exceeded that observed in aquatic environments such as freshwater, seawater, and sediment. Our results indicate that high enzyme activity within a pitcher could assist in the rapid decomposition of prey to maximize efficient nutrient use. In addition, we filtered the fluid to distinguish between dissolved enzyme activity and particle-bound activity. As a result, filtration treatment significantly decreased the activity in all enzymes, while pH value and Nepenthes species did not affect the enzyme activity. It suggested that enzymes bound to bacteria and other organic particles also would significantly contribute to the total enzyme activity of the fluid. Since organic particles are themselves usually colonized by attached and highly active bacteria, it is possible that microbe-derived enzymes also play an important role in nutrient recycling within the fluid and affect the metabolism of the Nepenthes pitcher plant.

Enhanced enzyme stability through site-directed covalent immobilization.

PubMed

Wu, Jeffrey Chun Yu; Hutchings, Christopher Hayden; Lindsay, Mark Jeffrey; Werner, Christopher James; Bundy, Bradley Charles

2015-01-10

Breakthroughs in enzyme immobilization have enabled increased enzyme recovery and reusability, leading to significant decreases in the cost of enzyme use and fueling biocatalysis growth. However, current enzyme immobilization techniques suffer from leaching, enzyme stability, and recoverability and reusability issues. Moreover, these techniques lack the ability to control the orientation of the immobilized enzymes. To determine the impact of orientation on covalently immobilized enzyme activity and stability, we apply our PRECISE (Protein Residue-Explicit Covalent Immobilization for Stability Enhancement) system to a model enzyme, T4 lysozyme. The PRECISE system uses non-canonical amino acid incorporation and the Huisgen 1,3-dipolar cycloaddition "click" reaction to enable directed enzyme immobilization at rationally chosen residues throughout an enzyme. Unlike previous site-specific systems, the PRECISE system is a truly covalent immobilization method. Utilizing this system, enzymes immobilized at proximate and distant locations from the active site were tested for activity and stability under denaturing conditions. Our results demonstrate that orientation control of covalently immobilized enzymes can provide activity and stability benefits exceeding that of traditional random covalent immobilization techniques. PRECISE immobilized enzymes were 50 and 73% more active than randomly immobilized enzymes after harsh freeze-thaw and chemical denaturant treatments. Copyright © 2014 Elsevier B.V. All rights reserved.

Spatial characterization of proteolytic enzyme activity in the foregut region of the adult necrophagous fly, Protophormia terraenovae.

PubMed

Rivers, David B; Acca, Gillian; Fink, Marc; Brogan, Rebecca; Schoeffield, Andrew

2014-08-01

The spatial distribution of proteolytic enzymes in the adult foregut of Protophormia terraenovae was studied in the context of protein digestion and regurgitation. Based on substrate specificity, pH optima, and use of specific protease inhibitors, all adults tested displayed enzyme activity in the foregut consistent with pepsin, trypsin and chymotrypsin. Chymotrypsin-like and trypsin-like enzyme activity were detected in all gut fluids and tissues tested, with chymotrypsin displaying the highest activity in saliva and salivary gland tissue, whereas maximal trypsin activity was evident in the crop. Pepsin-like activity was only evident in crop fluids and tissues. The activity of all three enzymes was low or undetectable (pepsin) in the fluids and tissue homogenates derived from the esophagus and cardia of any of the adults assayed. Fed adult females displayed higher enzyme activities than fed males, and the activity of all three enzymes were much more prevalent in fed adults than starved. The pH optimum of the trypsin-like enzyme was between pH 7.0 and 8.0; chymotrypsin was near pH 8.0; and maximal pepsin-like activity occurred between pH 1.0 and 2.0. Regurgitate from fed adult females displayed enzyme activity consistent with the proteolytic enzymes detected in crop gut fluids. Enzymes in regurgitate were not derived from food sources based on assays of bovine liver samples. These latter observations suggest that adult flies release fluids from foregut when encountering dry foods, potentially as a means to initiate extra-oral digestion. Copyright © 2014 Elsevier Ltd. All rights reserved.

Application of activity-based protein profiling to study enzyme function in adipocytes.

PubMed

Galmozzi, Andrea; Dominguez, Eduardo; Cravatt, Benjamin F; Saez, Enrique

2014-01-01

Activity-based protein profiling (ABPP) is a chemical proteomics approach that utilizes small-molecule probes to determine the functional state of enzymes directly in native systems. ABPP probes selectively label active enzymes, but not their inactive forms, facilitating the characterization of changes in enzyme activity that occur without alterations in protein levels. ABPP can be a tool superior to conventional gene expression and proteomic profiling methods to discover new enzymes active in adipocytes and to detect differences in the activity of characterized enzymes that may be associated with disorders of adipose tissue function. ABPP probes have been developed that react selectively with most members of specific enzyme classes. Here, using as an example the serine hydrolase family that includes many enzymes with critical roles in adipocyte physiology, we describe methods to apply ABPP analysis to the study of adipocyte enzymatic pathways. © 2014 Elsevier Inc. All rights reserved.

Cave Buttes Dam Foundation Report. Gila River Basin: Phoenix, Arizona and Vicinity (Including New River).

DTIC Science & Technology

1983-08-01

bedrock. Reservoir deposits are rich in silt and clay as shown by the plastic nature of material behind Cave Creek Dam. Recent alluvium is directly...formation and the zone I, impervious material. Then rolling with the rubber tired equipment would fill voids and small cracks with the clayey, plastic ...Assoclates US " 2 t 64urvey air J.1y 1969. ___OSOWRS g.Aorizatnfa( conirol ji 45 on G1A - EEA fSI Alaon~ral ieodtc Survey Dafum. Yerftci 4 1 -5 AtE

Transport Properties of Selected Elements and Compounds in the Gaseous State

DTIC Science & Technology

1957-01-01

coefficient (D) can be defined from the equations dp 0 -jA 6 dt M I dq = -kA dt dm a -DA a dt which relate the fluxes of momentum, heat and mass to the...000 ? 0080080 4 8aq ~ 0 Uot - 0- to 4 ; 𔃾 14𔃾 L ý * ; 4 40Ir w 4- 511 1 r14 0 Lv EEA 8 NI191 ;A 441 La 00 La~ 9-4 P f-4Ř 36 ~~~~~- 00 0 t00o L000

Effects of non-starch polysaccharides enzymes on pancreatic and small intestinal digestive enzyme activities in piglet fed diets containing high amounts of barley.

PubMed

Li, Wei-Fen; Feng, Jie; Xu, Zi-Rong; Yang, Cai-Mei

2004-03-15

To investigate effects of non-starch polysaccharides(NSP) enzymes on pancreatic and small intestinal digestive enzyme activities in piglet fed diets containing high amounts of barley. Sixty crossbred piglets averaging 13.5 kg were randomly assigned to two treatment groups with three replications (pens) based on sex and mass. Each group was fed on the diet based on barley with or without added NSP enzymes (0.15%) for a 40-d period. At the end of the experiment the pigs were weighed. Three piglets of each group were chosen and slaughtered. Pancreas, digesta from the distal end of the duodenum and jejunal mucosa were collected for determination. Activities of the digestive enzymes trypsin, chymotrypsin, amylase and lipase were determined in the small intestinal sections as well as in homogenates of pancreatic tissue. Maltase, sucrase, lactase and gamma-glutamyl transpeptidase (gamma-GT) activities were analyzed in jejunal mucosa. Supplementation with NSP enzymes improved growth performance of piglets. It showed that NSP enzymes had no effect on digestive enzyme activities in pancreas, but decreased the activities of proteolytic enzyme, trypsin, amylase and lipase in duodenal contents by 57.56%, 76.08%, 69.03% and 40.22%(P<0.05) compared with control, and increased gamma-GT activities in jejunal mucosa by 118.75%(P<0.05). Supplementation with NSP enzymes in barley based diets could improve piglets' growth performance, decrease activities of proteolytic enzyme, trypsin, amylase and lipase in duodenal contents and increase gamma-GT activities in jejunal mucosa.

A New Versatile Microarray-based Method for High Throughput Screening of Carbohydrate-active Enzymes*

PubMed Central

Vidal-Melgosa, Silvia; Pedersen, Henriette L.; Schückel, Julia; Arnal, Grégory; Dumon, Claire; Amby, Daniel B.; Monrad, Rune Nygaard; Westereng, Bjørge; Willats, William G. T.

2015-01-01

Carbohydrate-active enzymes have multiple biological roles and industrial applications. Advances in genome and transcriptome sequencing together with associated bioinformatics tools have identified vast numbers of putative carbohydrate-degrading and -modifying enzymes including glycoside hydrolases and lytic polysaccharide monooxygenases. However, there is a paucity of methods for rapidly screening the activities of these enzymes. By combining the multiplexing capacity of carbohydrate microarrays with the specificity of molecular probes, we have developed a sensitive, high throughput, and versatile semiquantitative enzyme screening technique that requires low amounts of enzyme and substrate. The method can be used to assess the activities of single enzymes, enzyme mixtures, and crude culture broths against single substrates, substrate mixtures, and biomass samples. Moreover, we show that the technique can be used to analyze both endo-acting and exo-acting glycoside hydrolases, polysaccharide lyases, carbohydrate esterases, and lytic polysaccharide monooxygenases. We demonstrate the potential of the technique by identifying the substrate specificities of purified uncharacterized enzymes and by screening enzyme activities from fungal culture broths. PMID:25657012

Effects of epithalon on activities gastrointestinal enzymes in young and old rats.

PubMed

Khavinson, V Kh; Malinin, V V; Timofeeva, N M; Egorova, V V; Nikitina, A A

2002-03-01

Peroral administration of Epithalon (Ala-Glu-Asp-Gly) to male and female Wistar rats aging 3 and 11 months changed activity of enzymes hydrolyzing carbohydrates, proteins, and phosphoric acid esters in various portions of the gastrointestinal tract. The most pronounced activation of enzymes was observed in 11-month-old animals. This effect diminished the differences in enzyme activities between young and old rats (compared to untreated animals). Our results indicate that Epithalon modulates activity of gastrointestinal enzymes during aging.

Co-immobilization of multiple enzymes by metal coordinated nucleotide hydrogel nanofibers: improved stability and an enzyme cascade for glucose detection.

PubMed

Liang, Hao; Jiang, Shuhui; Yuan, Qipeng; Li, Guofeng; Wang, Feng; Zhang, Zijie; Liu, Juewen

2016-03-21

Preserving enzyme activity and promoting synergistic activity via co-localization of multiple enzymes are key topics in bionanotechnology, materials science, and analytical chemistry. This study reports a facile method for co-immobilizing multiple enzymes in metal coordinated hydrogel nanofibers. Specifically, four types of protein enzymes, including glucose oxidase, Candida rugosa lipase, α-amylase, and horseradish peroxidase, were respectively encapsulated in a gel nanofiber made of Zn(2+) and adenosine monophosphate (AMP) with a simple mixing step. Most enzymes achieved quantitative loading and retained full activity. At the same time, the entrapped enzymes were more stable against temperature variation (by 7.5 °C), protease attack, extreme pH (by 2-fold), and organic solvents. After storing for 15 days, the entrapped enzyme still retained 70% activity while the free enzyme nearly completely lost its activity. Compared to nanoparticles formed with AMP and lanthanide ions, the nanofiber gels allowed much higher enzyme activity. Finally, a highly sensitive and selective biosensor for glucose was prepared using the gel nanofiber to co-immobilize glucose oxidase and horseradish peroxidase for an enzyme cascade system. A detection limit of 0.3 μM glucose with excellent selectivity was achieved. This work indicates that metal coordinated materials using nucleotides are highly useful for interfacing with biomolecules.

[Enzymatic degradation of organophosphorus insecticide chlorpyrifos by fungus WZ-I].

PubMed

Xie, Hui; Zhu, Lu-sheng; Wang, Jun; Wang, Xiu-guo; Liu, Wei; Qian, Bo; Wang, Qian

2005-11-01

Degradation characteristics of chlorpyrifos insecticides was determined by the crude enzyme extracted from the isolated strain WZ-I ( Fusarium LK. ex Fx). The best separating condition and the degrading characteristic of chlorpyrifos were studied. Rate of degradation for chlorpyrifos by its intracellular enzyme, extracellular enzyme and cell fragment was 60.8%, 11.3% and 48%, respectively. The degrading enzyme was extracted after this fungus was incubated for 8 generations in the condition of noninducement, and its enzymic activity lost less, the results show that this enzyme is an intracellular and connatural enzyme. The solubility protein of the crude enzyme was determined with Albumin (bovine serum) as standard protein and the solubility protein of the crude enzyme was 3.36 mg x mL(-1). The pH optimum for crude enzyme was 6.8 for enzymatic degradation of chlorpyrifos, and it had comparatively high activity in the range of pH 6.0 - 9.0. The optimum temperature for enzymatic activity was at 40 degrees C, it still had comparatively high activity in the range of temperature 20-50 degrees C, the activity of enzyme rapidly reduced at 55 degrees C, its activity was 41% of the maximal activity. The crude enzyme showed Km value for chlorpyrifos of 1.049 26 mmol x L(-1), and the maximal enzymatic degradation rate was 0.253 5 micromol x (mg x min)(-1). Additional experimental evidence suggests that the enzyme had the stability of endure for temperature and pH, the crude enzyme of fungus WZ-I could effectively degrade chlorpyrifos.

Molecular architectures and functions of radical enzymes and their (re)activating proteins.

PubMed

Shibata, Naoki; Toraya, Tetsuo

2015-10-01

Certain proteins utilize the high reactivity of radicals for catalysing chemically challenging reactions. These proteins contain or form a radical and therefore named 'radical enzymes'. Radicals are introduced by enzymes themselves or by (re)activating proteins called (re)activases. The X-ray structures of radical enzymes and their (re)activases revealed some structural features of these molecular apparatuses which solved common enigmas of radical enzymes—i.e. how the enzymes form or introduce radicals at the active sites, how they use the high reactivity of radicals for catalysis, how they suppress undesired side reactions of highly reactive radicals and how they are (re)activated when inactivated by extinction of radicals. This review highlights molecular architectures of radical B12 enzymes, radical SAM enzymes, tyrosyl radical enzymes, glycyl radical enzymes and their (re)activating proteins that support their functions. For generalization, comparisons of the recently reported structures of radical enzymes with those of canonical radical enzymes are summarized here. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Defying the activity-stability trade-off in enzymes: taking advantage of entropy to enhance activity and thermostability.

PubMed

Siddiqui, Khawar Sohail

2017-05-01

The biotechnological applications of enzymes are limited due to the activity-stability trade-off, which implies that an increase in activity is accompanied by a concomitant decrease in protein stability. This premise is based on thermally adapted homologous enzymes where cold-adapted enzymes show high intrinsic activity linked to enhanced thermolability. In contrast, thermophilic enzymes show low activity around ambient temperatures. Nevertheless, genetically and chemically modified enzymes are beginning to show that the activity-stability trade-off can be overcome. In this review, the origin of the activity-stability trade-off, the thermodynamic basis for enhanced activity and stability, and various approaches for escaping the activity-stability trade-off are discussed. The role of entropy in enhancing both the activity and the stability of enzymes is highlighted with a special emphasis placed on the involvement of solvent water molecules. This review is concluded with suggestions for further research, which underscores the implications of these findings in the context of productivity curves, the Daniel-Danson equilibrium model, catalytic antibodies, and life on cold planets.

Effects of Nanoparticle Size on Multilayer Formation and Kinetics of Tethered Enzymes.

PubMed

Lata, James P; Gao, Lizeng; Mukai, Chinatsu; Cohen, Roy; Nelson, Jacquelyn L; Anguish, Lynne; Coonrod, Scott; Travis, Alexander J

2015-09-16

Despite numerous applications, we lack fundamental understanding of how variables such as nanoparticle (NP) size influence the activity of tethered enzymes. Previously, we showed that biomimetic oriented immobilization yielded higher specific activities versus nonoriented adsorption or carboxyl-amine binding. Here, we standardize NP attachment strategy (oriented immobilization via hexahistidine tags) and composition (Ni-NTA coated gold NPs), to test the impact of NP size (⌀5, 10, 20, and 50 nm) on multilayer formation, activity, and kinetic parameters (kcat, KM, kcat/KM) of enzymes representing three different classes: glucose-6-phosphate isomerase (GPI), an isomerase; Glyceraldehyde-3-phosphate dehydrogenase S (GAPDHS), an oxidoreductase; and pyruvate kinase (PK), a transferase. Contrary to other reports, we observed no trend in kinetic parameters for individual enzymes when found in monolayers (<100% enzyme coverage), suggesting an advantage for oriented immobilization versus other attachment strategies. Saturating the NPs to maximize activity per NP resulted in enzyme multilayer formation. Under these conditions, total activity per NP increased with increasing NP size. Conversely, specific activity for all three enzymes was highest when tethered to the smallest NPs, retaining a remarkable 73-94% of the activity of free/untethered enzymes. Multilayer formations caused a clear trend of kcat decreasing with increasing NP size, yet negligible change in KM. Understanding the fundamental relationships between NP size and tethered enzyme activity enables optimized design of various applications, maximizing activity per NP or activity per enzyme molecule.

[Seasonal variations of soil enzyme activities in typical plant communities in the Ebinur Lake wetland, China].

PubMed

Zhu, Hai Qiang; Li, Yan Hong; Li, Fa Dong

2017-04-18

In this study, the soil catalase, phosphatase and urease activities of typical plant communities of reed (Phragmites australis) and tamarisk (Tamarix ramosissima) and their influencing factors were investigated in Ebinur Lake wetland. The results showed that three soil enzyme activities of reed and tamarisk had seasonal dynamic characteristics during different growth periods. For the reed community, the peak concentrations of soil catalase, phosphatase and urease appeared at vigorous stage with 3.26, 0.60 and 0.33 mg·g -1 , respectively, and the minimum value occurred at budding stage and leaf-expansion stage. For the tamarisk community, the peak values of three soil enzyme activities appeared at withered stage with values of 6.33, 0.58 and 0.21 mg·g -1 , respectively, and the valley values were observed at flowering and vigorous stages. Urease was stable during different growth periods, and it could be used as an indicator to identify the differences of soil enzyme activities in the wetlands. The enzyme activities of reed and tamarisk had significant positive correlation with soil organic matter and total P in all growth periods, while there was no significant relationship between enzyme activities and soil water content. The enzyme activities of reed had significant positive correlation with ammonium nitrogen in the rapid growth period. There were no significant relationships between enzyme activities and soil salinity in both communities. The soil enzyme activities of reed and tamarisk were controlled by many factors. Soil organic matter, soil water and soil temperature were the main factors influencing the enzyme activities in the Ebinur Lake wetland.

Assessment of digestive enzymes activity during the fry development of the endangered Caspian brown trout Salmo caspius.

PubMed

Zamani, A; Hajimoradloo, A; Madani, R; Farhangi, M

2009-09-01

The study of digestive enzymes activity at Salmo caspius fry showed that enzymes were available at the moment of mouth opening on the first day post hatching (dph) and the activity of enzymes showed no significant difference from the hatching day 28 dph. An increased activity was seen between 32 and 43 dph and this activity was significantly higher than the activity during the first 28 days. In the primary stages after yolk sac resorption (43-58 dph), enzymes activity showed an increased profile, however none of them showed a significant difference between 43 and 58 dph.

Nanocaged enzymes with enhanced catalytic activity and increased stability against protease digestion

NASA Astrophysics Data System (ADS)

Zhao, Zhao; Fu, Jinglin; Dhakal, Soma; Johnson-Buck, Alexander; Liu, Minghui; Zhang, Ting; Woodbury, Neal W.; Liu, Yan; Walter, Nils G.; Yan, Hao

2016-02-01

Cells routinely compartmentalize enzymes for enhanced efficiency of their metabolic pathways. Here we report a general approach to construct DNA nanocaged enzymes for enhancing catalytic activity and stability. Nanocaged enzymes are realized by self-assembly into DNA nanocages with well-controlled stoichiometry and architecture that enabled a systematic study of the impact of both encapsulation and proximal polyanionic surfaces on a set of common metabolic enzymes. Activity assays at both bulk and single-molecule levels demonstrate increased substrate turnover numbers for DNA nanocage-encapsulated enzymes. Unexpectedly, we observe a significant inverse correlation between the size of a protein and its activity enhancement. This effect is consistent with a model wherein distal polyanionic surfaces of the nanocage enhance the stability of active enzyme conformations through the action of a strongly bound hydration layer. We further show that DNA nanocages protect encapsulated enzymes against proteases, demonstrating their practical utility in functional biomaterials and biotechnology.

Nanocaged enzymes with enhanced catalytic activity and increased stability against protease digestion

PubMed Central

Zhao, Zhao; Fu, Jinglin; Dhakal, Soma; Johnson-Buck, Alexander; Liu, Minghui; Zhang, Ting; Woodbury, Neal W.; Liu, Yan; Walter, Nils G.; Yan, Hao

2016-01-01

Cells routinely compartmentalize enzymes for enhanced efficiency of their metabolic pathways. Here we report a general approach to construct DNA nanocaged enzymes for enhancing catalytic activity and stability. Nanocaged enzymes are realized by self-assembly into DNA nanocages with well-controlled stoichiometry and architecture that enabled a systematic study of the impact of both encapsulation and proximal polyanionic surfaces on a set of common metabolic enzymes. Activity assays at both bulk and single-molecule levels demonstrate increased substrate turnover numbers for DNA nanocage-encapsulated enzymes. Unexpectedly, we observe a significant inverse correlation between the size of a protein and its activity enhancement. This effect is consistent with a model wherein distal polyanionic surfaces of the nanocage enhance the stability of active enzyme conformations through the action of a strongly bound hydration layer. We further show that DNA nanocages protect encapsulated enzymes against proteases, demonstrating their practical utility in functional biomaterials and biotechnology. PMID:26861509

Nanocaged enzymes with enhanced catalytic activity and increased stability against protease digestion.

PubMed

Zhao, Zhao; Fu, Jinglin; Dhakal, Soma; Johnson-Buck, Alexander; Liu, Minghui; Zhang, Ting; Woodbury, Neal W; Liu, Yan; Walter, Nils G; Yan, Hao

2016-02-10

Cells routinely compartmentalize enzymes for enhanced efficiency of their metabolic pathways. Here we report a general approach to construct DNA nanocaged enzymes for enhancing catalytic activity and stability. Nanocaged enzymes are realized by self-assembly into DNA nanocages with well-controlled stoichiometry and architecture that enabled a systematic study of the impact of both encapsulation and proximal polyanionic surfaces on a set of common metabolic enzymes. Activity assays at both bulk and single-molecule levels demonstrate increased substrate turnover numbers for DNA nanocage-encapsulated enzymes. Unexpectedly, we observe a significant inverse correlation between the size of a protein and its activity enhancement. This effect is consistent with a model wherein distal polyanionic surfaces of the nanocage enhance the stability of active enzyme conformations through the action of a strongly bound hydration layer. We further show that DNA nanocages protect encapsulated enzymes against proteases, demonstrating their practical utility in functional biomaterials and biotechnology.

A new versatile microarray-based method for high throughput screening of carbohydrate-active enzymes.

PubMed

Vidal-Melgosa, Silvia; Pedersen, Henriette L; Schückel, Julia; Arnal, Grégory; Dumon, Claire; Amby, Daniel B; Monrad, Rune Nygaard; Westereng, Bjørge; Willats, William G T

2015-04-03

Carbohydrate-active enzymes have multiple biological roles and industrial applications. Advances in genome and transcriptome sequencing together with associated bioinformatics tools have identified vast numbers of putative carbohydrate-degrading and -modifying enzymes including glycoside hydrolases and lytic polysaccharide monooxygenases. However, there is a paucity of methods for rapidly screening the activities of these enzymes. By combining the multiplexing capacity of carbohydrate microarrays with the specificity of molecular probes, we have developed a sensitive, high throughput, and versatile semiquantitative enzyme screening technique that requires low amounts of enzyme and substrate. The method can be used to assess the activities of single enzymes, enzyme mixtures, and crude culture broths against single substrates, substrate mixtures, and biomass samples. Moreover, we show that the technique can be used to analyze both endo-acting and exo-acting glycoside hydrolases, polysaccharide lyases, carbohydrate esterases, and lytic polysaccharide monooxygenases. We demonstrate the potential of the technique by identifying the substrate specificities of purified uncharacterized enzymes and by screening enzyme activities from fungal culture broths. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Measuring the Enzyme Activity of Arabidopsis Deubiquitylating Enzymes.

PubMed

Kalinowska, Kamila; Nagel, Marie-Kristin; Isono, Erika

2016-01-01

Deubiquitylating enzymes, or DUBs, are important regulators of ubiquitin homeostasis and substrate stability, though the molecular mechanisms of most of the DUBs in plants are not yet understood. As different ubiquitin chain types are implicated in different biological pathways, it is important to analyze the enzyme characteristic for studying a DUB. Quantitative analysis of DUB activity is also important to determine enzyme kinetics and the influence of DUB binding proteins on the enzyme activity. Here, we show methods to analyze DUB activity using immunodetection, Coomassie Brilliant Blue staining, and fluorescence measurement that can be useful for understanding the basic characteristic of DUBs.

Enzyme activities in plasma, kidney, liver, and muscle of five avian species

USGS Publications Warehouse

Franson, J.C.; Murray, H.C.; Bunck, C.

1985-01-01

Activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), creatine phosphokinase (CPK), and lactate dehydrogenase (LDH) were determined in plasma, kidney, liver, and muscle from five species of captive birds. Few differences occurred in plasma activities between sexes but considerable differences occurred between species. All five enzymes were detected in each of the tissues sampled. Relative enzyme activities in liver, kidney, and muscle were similar for each species. CPK activity was much higher in muscle than in liver or kidney and, of the five enzymes studied, may be the best indicator of muscle damage. Most of the other enzymes were more evenly distributed among the three tissues, and no organ-specific enzyme could be identified for liver or kidney. Because of interspecific variations in plasma enzyme activities, it is important to establish baseline values for each species to ensure accurate interpretation of results.

The molecular basis of the effect of temperature on enzyme activity.

PubMed

Daniel, Roy M; Peterson, Michelle E; Danson, Michael J; Price, Nicholas C; Kelly, Sharon M; Monk, Colin R; Weinberg, Cristina S; Oudshoorn, Matthew L; Lee, Charles K

2009-12-23

Experimental data show that the effect of temperature on enzymes cannot be adequately explained in terms of a two-state model based on increases in activity and denaturation. The Equilibrium Model provides a quantitative explanation of enzyme thermal behaviour under reaction conditions by introducing an inactive (but not denatured) intermediate in rapid equilibrium with the active form. The temperature midpoint (Teq) of the rapid equilibration between the two forms is related to the growth temperature of the organism, and the enthalpy of the equilibrium (DeltaHeq) to its ability to function over various temperature ranges. In the present study, we show that the difference between the active and inactive forms is at the enzyme active site. The results reveal an apparently universal mechanism, independent of enzyme reaction or structure, based at or near the active site, by which enzymes lose activity as temperature rises, as opposed to denaturation which is global. Results show that activity losses below Teq may lead to significant errors in the determination of DeltaG*cat made on the basis of the two-state ('Classical') model, and the measured kcat will then not be a true indication of an enzyme's catalytic power. Overall, the results provide a molecular rationale for observations that the active site tends to be more flexible than the enzyme as a whole, and that activity losses precede denaturation, and provide a general explanation in molecular terms for the effect of temperature on enzyme activity.

A review on the effects of supercritical carbon dioxide on enzyme activity.

PubMed

Wimmer, Zdenek; Zarevúcka, Marie

2010-01-19

Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO(2). The stability and the activity of enzymes exposed to carbon dioxide under high pressure depend on enzyme species, water content in the solution and on the pressure and temperature of the reaction system. The three-dimensional structure of enzymes may be significantly altered under extreme conditions, causing their denaturation and consequent loss of activity. If the conditions are less adverse, the protein structure may be largely retained. Minor structural changes may induce an alternative active protein state with altered enzyme activity, specificity and stability.

A Review on the Effects of Supercritical Carbon Dioxide on Enzyme Activity

PubMed Central

Wimmer, Zdeněk; Zarevúcka, Marie

2010-01-01

Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO2. The stability and the activity of enzymes exposed to carbon dioxide under high pressure depend on enzyme species, water content in the solution and on the pressure and temperature of the reaction system. The three-dimensional structure of enzymes may be significantly altered under extreme conditions, causing their denaturation and consequent loss of activity. If the conditions are less adverse, the protein structure may be largely retained. Minor structural changes may induce an alternative active protein state with altered enzyme activity, specificity and stability. PMID:20162013

Mechanobiocatalysis: Modulating Enzymatic Activity with Mechanical Force

DTIC Science & Technology

2015-09-28

displayed by enzymes and other materials. It was demonstrated that the application of forces to enzymes properly outfitted with polymers resulted in...distortions at the active sites of the corresponding enzymes . For example, polymer-protein composites were found to display photophysical properties that...intrinsic activities displayed by enzymes and other materials. It was demonstrated that the application of forces to enzymes properly outfitted with polymers

Seasonal influenza immunisation in Europe. Overview of recommendations and vaccination coverage for three seasons: pre-pandemic (2008/09), pandemic (2009/10) and post-pandemic (2010/11).

PubMed

Mereckiene, J; Cotter, S; Nicoll, A; Lopalco, P; Noori, T; Weber, Jt; D'Ancona, F; Levy-Bruhl, D; Dematte, L; Giambi, C; Valentiner-Branth, P; Stankiewicz, I; Appelgren, E; O Flanagan, D

2014-04-24

Since 2008, annual surveys of influenza vaccination policies, practices and coverage have been undertaken in 29 European Union (EU)/ European Economic Area (EEA) countries. After 2009, this monitored the impact of European Council recommendation to increase vaccination coverage to 75% among risk groups. This paper summarises the results of three seasonal influenza seasons: 2008/09, 2009/10 and 2010/11. In 2008/09, 27/29 countries completed the survey; in 2009/10 and 2010/11, 28/29 completed it. All or almost all countries recommended vaccination of older people (defined as those aged ≥50, ≥55, ≥59, ≥60 or ≥65 years), and people aged ≥6 months with clinical risk and healthcare workers. A total of 23 countries provided vaccination coverage data for older people, but only 7 and 10 had data for the clinical risk groups and healthcare workers, respectively. The number of countries recommending vaccination for some or all pregnant women increased from 10 in 2008/09 to 22 in 2010/11. Only three countries could report coverage among pregnant women. Seasonal influenza vaccination coverage during and after the pandemic season in older people and clinical groups remained unchanged in countries with higher coverage. However, small decreases were seen in most countries during this period. The results of the surveys indicate that most EU/EEA countries recommend influenza vaccination for the main target groups; however, only a few countries have achieved the target of 75% coverage among risk groups. Coverage among healthcare workers remained low.

A sorting nexin 17-binding domain within the LRP1 cytoplasmic tail mediates receptor recycling through the basolateral sorting endosome.

PubMed

Farfán, Pamela; Lee, Jiyeon; Larios, Jorge; Sotelo, Pablo; Bu, Guojun; Marzolo, María-Paz

2013-07-01

Sorting nexin 17 (SNX17) is an adaptor protein present in early endosomal antigen 1 (EEA1)-positive sorting endosomes that promotes the efficient recycling of low-density lipoprotein receptor-related protein 1 (LRP1) to the plasma membrane through recognition of the first NPxY motif in the cytoplasmic tail of this receptor. The interaction of LRP1 with SNX17 also regulates the basolateral recycling of the receptor from the basolateral sorting endosome (BSE). In contrast, megalin, which is apically distributed in polarized epithelial cells and localizes poorly to EEA1-positive sorting endosomes, does not interact with SNX17, despite containing three NPxY motifs, indicating that this motif is not sufficient for receptor recognition by SNX17. Here, we identified a cluster of 32 amino acids within the cytoplasmic domain of LRP1 that is both necessary and sufficient for SNX17 binding. To delineate the function of this SNX17-binding domain, we generated chimeric proteins in which the SNX17-binding domain was inserted into the cytoplasmic tail of megalin. This insertion mediated the binding of megalin to SNX17 and modified the cell surface expression and recycling of megalin in non-polarized cells. However, the polarized localization of chimeric megalin was not modified in polarized Madin-Darby canine kidney cells. These results provide evidence regarding the molecular and cellular mechanisms underlying the specificity of SNX17-binding receptors and the restricted function of SNX17 in the BSE. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Analysis of Recent European Surface Ozone Trends Considering Site Representativeness and Mete-orology

NASA Astrophysics Data System (ADS)

Henne, S.; Fleming, Z.; Brunner, D.; Klausen, J.; Buchmann, B.

2009-04-01

Recent trends of surface ozone (O3) within Europe vary substantially depending on the location and surroundings of a measurement site. The influence of long-range transport from North America and Asia, changes in stratosphere-troposphere exchange, increase in lower stratospheric O3 and changes in advection patterns are possible drivers for the observed O3 trends. O3 concentrations greatly depend on meteorology (temperature and radiation) and local to regional emissions of precursor gases and therefore on the representativeness of a site (e.g. background vs. urban site) and regional emission trends. We investigated the representativeness of 1264 "rural" and "suburban" background sites (as available through the European Environment Agency (EEA )Airbase database) by analysing population density, land cover and topography in the surrounding of the sites. A hierarchical clustering method was applied to derive an independent site categorization. The two area types as specified by EEA are split into 7 categories: elevated, lowered, remote, rural, rural/coastal, rural/polluted, suburban. Furthermore, we analysed the trend of surface O3 and Ox (O3+NO2) for the mentioned sites based on the above site categorization, local meteorology and precursor emission trends. Of the 1264 sites 161 possess sufficiently long and complete O3 data series suitable for robust trend estimation, while for 100 sites both O3 and NO2 data are available. We present a strategy for further data exclusion based on available data quality information and a break detection algorithm. First results of the trend analysis applying different statistical approaches are discussed.

Internationally trained pharmacists' perception of their communication proficiency and their views on the impact on patient safety.

PubMed

Ziaei, Zainab; Hassell, Karen; Schafheutle, Ellen I

2015-01-01

According to Great Britain (GB)'s pharmacy regulator's standards of conduct, ethics and performance, pharmacists have a responsibility to ensure that they have sufficient linguistic skills to communicate and perform their job safely. Yet, very little is known about internationally trained pharmacists' (ITPs) linguistic proficiency. The purpose of this study was to investigate ITPs' perceptions of their communication proficiency and the resultant impact on patient safety. Eight focus groups were conducted between May and July 2010, with 31 European Economic Area (EEA) and 11 non-EEA pharmacists who, at the time of the study, practiced in community pharmacy (n = 29) or hospital (n = 13), in London, Manchester, Liverpool and Glasgow. The framework method was used to analyze qualitative data, and the Model of Communicative Proficiency (MCP) served as a framework to handle and explain the data obtained. ITPs experienced communication difficulties through new dialects, use of idioms and colloquial language in their workplace. The differences between the "BBC English" they learned formally and the "Street English" used in GB also led to difficulties. Culture was also recognized as an important aspect of communication. ITPs in this study were adamant that communication difficulty did not compromise patient safety. Communicative deficiency of ITPs arose primarily from two sources: linguistic competence and socio-cultural competence. These deficiencies could have negative implications for patient safety. The findings of this study should be taken into account when designing adaptation programs for ITPs. Copyright © 2015 Elsevier Inc. All rights reserved.

[Study on relationship between effective components and soil enzyme activity in different growth patterns of Panax ginseng].

PubMed

Yang, Yan-Wen; Jiang, Yuan-Tong

2016-08-01

Study on 5 effective components and 6 soil enzyme activities of 2 different growth patterns, analyse the dates with the canonical correlation analysis, In order to reveal the relations between the effective components and soil enzyme activities. The result showed that they had a great relation between the effective components and soil enzyme activities, the activity of the same enzyme in humus soil was higher than that in farmland soil. Growth pattern of farmland soil, if the invertase and phosphatase activity were too high, which would inhibit the accumulation of total ginsenoside, water-miscible total proteins and total amino acid; Growth pattern of humus soil, if the invertase, urease and phosphatase activity were too high, which would inhibit the accumulation of total ginsenoside and the total essential oils. Integral soil enzyme activity can be used as a index of soil quality, which, together with other growth factors. The appropriate enzyme activity can accelerate the circulation and transformation of all kinds of material in the soil, improve effectively components accumulation. Copyright© by the Chinese Pharmaceutical Association.

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.

Correlation Among Soil Enzyme Activities, Root Enzyme Activities, and Contaminant Removal in Two-Stage In Situ Constructed Wetlands Purifying Domestic Wastewater.

PubMed

Ni, Lixiao; Xu, Jiajun; Chu, Xianglin; Li, Shiyin; Wang, Peifang; Li, Yiping; Li, Yong; Zhu, Liang; Wang, Chao

2016-07-01

Two-stage in situ wetlands (two vertical flow constructed wetlands in parallel and a horizontal flow constructed wetland) were constructed for studying domestic wastewater purification and the correlations between contaminant removal and plant and soil enzyme activities. Results indicated the removal efficiency of NH4 (+) and NO3 (-) were significantly correlated with both urease and protease activity, and the removal of total phosphorus was significantly correlated with phosphatase activity. Chemical oxygen demand removal was not correlated with enzyme activity in constructed wetlands. Plant root enzyme (urease, phosphatase, protease and cellulose) activity correlation was apparent with all contaminant removal in the two vertical flow constructed wetlands. However, the correlation between the plant root enzyme activity and contaminant removal was poor in horizontal flow constructed wetlands. Results indicated that plant roots clearly played a role in the removal of contaminants.

Heparin/heparan sulfate 6-O-sulfatase from Flavobacterium heparinum: integrated structural and biochemical investigation of enzyme active site and substrate specificity.

PubMed

Myette, James R; Soundararajan, Venkataramanan; Shriver, Zachary; Raman, Rahul; Sasisekharan, Ram

2009-12-11

Heparin and heparan sulfate glycosaminoglycans (HSGAGs) comprise a chemically heterogeneous class of sulfated polysaccharides. The development of structure-activity relationships for this class of polysaccharides requires the identification and characterization of degrading enzymes with defined substrate specificity and enzymatic activity. Toward this end, we report here the molecular cloning and extensive structure-function analysis of a 6-O-sulfatase from the Gram-negative bacterium Flavobacterium heparinum. In addition, we report the recombinant expression of this enzyme in Escherichia coli in a soluble, active form and identify it as a specific HSGAG sulfatase. We further define the mechanism of action of the enzyme through biochemical and structural studies. Through the use of defined substrates, we investigate the kinetic properties of the enzyme. This analysis was complemented by homology-based molecular modeling studies that sought to rationalize the substrate specificity of the enzyme and mode of action through an analysis of the active-site topology of the enzyme including identifying key enzyme-substrate interactions and assigning key amino acids within the active site of the enzyme. Taken together, our structural and biochemical studies indicate that 6-O-sulfatase is a predominantly exolytic enzyme that specifically acts on N-sulfated or N-acetylated 6-O-sulfated glucosamines present at the non-reducing end of HSGAG oligosaccharide substrates. This requirement for the N-acetyl or N-sulfo groups on the glucosamine substrate can be explained through eliciting favorable interactions with key residues within the active site of the enzyme. These findings provide a framework that enables the use of 6-O-sulfatase as a tool for HSGAG structure-activity studies as well as expand our biochemical and structural understanding of this important class of enzymes.

Stabilizing effect of biochar on soil extracellular enzymes after a denaturing stress.

PubMed

Elzobair, Khalid A; Stromberger, Mary E; Ippolito, James A

2016-01-01

Stabilizing extracellular enzymes may maintain enzymatic activity while protecting enzymes from proteolysis and denaturation. A study determined whether a fast pyrolysis hardwood biochar (CQuest™) would reduce evaporative losses, subsequently stabilizing soil extracellular enzymes and prohibiting potential enzymatic activity loss following a denaturing stress (microwaving). Soil was incubated in the presence of biochar (0%, 1%, 2%, 5%, or 10% by wt.) for 36 days and then exposed to microwave energies (0, 400, 800, 1600, or 3200 J g(-1) soil). Soil enzymes (β-glucosidase, β-d-cellobiosidase, N-acetyl-β-glucosaminidase, phosphatase, leucine aminopeptidase, β-xylosidase) were analyzed by fluorescence-based assays. Biochar amendment reduced leucine aminopeptidase and β-xylosidase potential activity after the incubation period and prior to stress exposure. The 10% biochar rate reduced soil water loss at the lowest stress level (400 J microwave energy g(-1) soil). Enzyme stabilization was demonstrated for β-xylosidase; intermediate biochar application rates prevented a complete loss of this enzyme's potential activity after soil was exposed to 400 (1% biochar treatment) or 1600 (5% biochar treatment) J microwave energy g(-1) soil. Remaining enzyme potential activities were not affected by biochar, and activities decreased with increasing stress levels. We concluded that biochar has the potential to reduce evaporative soil water losses and stabilize certain extracellular enzymes where activity is maintained after a denaturing stress; this effect was biochar rate and enzyme dependent. While biochar may reduce the potential activity of certain soil extracellular enzymes, this phenomenon was not universal as the majority of enzymes assayed in this study were unaffected by exposure to biochar. Copyright © 2015 Elsevier Ltd. All rights reserved.

Rapid intranasal delivery of chloramphenicol acetyltransferase in the active form to different brain regions as a model for enzyme therapy in the CNS.

PubMed

Appu, Abhilash P; Arun, Peethambaran; Krishnan, Jishnu K S; Moffett, John R; Namboodiri, Aryan M A

2016-02-01

The blood brain barrier (BBB) is critical for maintaining central nervous system (CNS) homeostasis by restricting entry of potentially toxic substances. However, the BBB is a major obstacle in the treatment of neurotoxicity and neurological disorders due to the restrictive nature of the barrier to many medications. Intranasal delivery of active enzymes to the brain has therapeutic potential for the treatment of numerous CNS enzyme deficiency disorders and CNS toxicity caused by chemical threat agents. The aim of this work is to provide a sensitive model system for analyzing the rapid delivery of active enzymes into various regions of the brain with therapeutic bioavailability. We tested intranasal delivery of chloramphenicol acetyltransferase (CAT), a relatively large (75kD) enzyme, in its active form into different regions of the brain. CAT was delivered intranasally to anaesthetized rats and enzyme activity was measured in different regions using a highly specific High Performance Thin Layer Chromatography (HP-TLC)-radiometry coupled assay. Active enzyme reached all examined areas of the brain within 15min (the earliest time point tested). In addition, the yield of enzyme activity in the brain was almost doubled in the brains of rats pre-treated with matrix metalloproteinase-9 (MMP-9). Intranasal administration of active enzymes in conjunction with MMP-9 to the CNS is both rapid and effective. The present results suggest that intranasal enzyme therapy is a promising method for counteracting CNS chemical threat poisoning, as well as for treating CNS enzyme deficiency disorders. Published by Elsevier B.V.

Microbial responses to multi-factor climate change: effects on soil enzymes.

PubMed

Steinweg, J Megan; Dukes, Jeffrey S; Paul, Eldor A; Wallenstein, Matthew D

2013-01-01

The activities of extracellular enzymes, the proximate agents of decomposition in soils, are known to depend strongly on temperature, but less is known about how they respond to changes in precipitation patterns, and the interaction of these two components of climate change. Both enzyme production and turnover can be affected by changes in temperature and soil moisture, thus it is difficult to predict how enzyme pool size may respond to altered climate. Soils from the Boston-Area Climate Experiment (BACE), which is located in an old field (on abandoned farmland), were used to examine how climate variables affect enzyme activities and microbial biomass carbon (MBC) in different seasons and in soils exposed to a combination of three levels of precipitation treatments (ambient, 150% of ambient during growing season, and 50% of ambient year-round) and four levels of warming treatments (unwarmed to ~4°C above ambient) over the course of a year. Warming, precipitation and season had very little effect on potential enzyme activity. Most models assume that enzyme dynamics follow microbial biomass, because enzyme production should be directly controlled by the size and activity of microbial biomass. We observed differences among seasons and treatments in mass-specific potential enzyme activity, suggesting that this assumption is invalid. In June 2009, mass-specific potential enzyme activity, using chloroform fumigation-extraction MBC, increased with temperature, peaking under medium warming and then declining under the highest warming. This finding suggests that either enzyme production increased with temperature or turnover rates decreased. Increased maintenance costs associated with warming may have resulted in increased mass-specific enzyme activities due to increased nutrient demand. Our research suggests that allocation of resources to enzyme production could be affected by climate-induced changes in microbial efficiency and maintenance costs.

Allosteric regulation of epigenetic modifying enzymes.

PubMed

Zucconi, Beth E; Cole, Philip A

2017-08-01

Epigenetic enzymes including histone modifying enzymes are key regulators of gene expression in normal and disease processes. Many drug development strategies to target histone modifying enzymes have focused on ligands that bind to enzyme active sites, but allosteric pockets offer potentially attractive opportunities for therapeutic development. Recent biochemical studies have revealed roles for small molecule and peptide ligands binding outside of the active sites in modulating the catalytic activities of histone modifying enzymes. Here we highlight several examples of allosteric regulation of epigenetic enzymes and discuss the biological significance of these findings. Copyright © 2017 Elsevier Ltd. All rights reserved.

Activity-based proteomics of enzyme superfamilies: serine hydrolases as a case study.

PubMed

Simon, Gabriel M; Cravatt, Benjamin F

2010-04-09

Genome sequencing projects have uncovered thousands of uncharacterized enzymes in eukaryotic and prokaryotic organisms. Deciphering the physiological functions of enzymes requires tools to profile and perturb their activities in native biological systems. Activity-based protein profiling has emerged as a powerful chemoproteomic strategy to achieve these objectives through the use of chemical probes that target large swaths of enzymes that share active-site features. Here, we review activity-based protein profiling and its implementation to annotate the enzymatic proteome, with particular attention given to probes that target serine hydrolases, a diverse superfamily of enzymes replete with many uncharacterized members.

Profiling the orphan enzymes.

PubMed

Sorokina, Maria; Stam, Mark; Médigue, Claudine; Lespinet, Olivier; Vallenet, David

2014-06-06

The emergence of Next Generation Sequencing generates an incredible amount of sequence and great potential for new enzyme discovery. Despite this huge amount of data and the profusion of bioinformatic methods for function prediction, a large part of known enzyme activities is still lacking an associated protein sequence. These particular activities are called "orphan enzymes". The present review proposes an update of previous surveys on orphan enzymes by mining the current content of public databases. While the percentage of orphan enzyme activities has decreased from 38% to 22% in ten years, there are still more than 1,000 orphans among the 5,000 entries of the Enzyme Commission (EC) classification. Taking into account all the reactions present in metabolic databases, this proportion dramatically increases to reach nearly 50% of orphans and many of them are not associated to a known pathway. We extended our survey to "local orphan enzymes" that are activities which have no representative sequence in a given clade, but have at least one in organisms belonging to other clades. We observe an important bias in Archaea and find that in general more than 30% of the EC activities have incomplete sequence information in at least one superkingdom. To estimate if candidate proteins for local orphans could be retrieved by homology search, we applied a simple strategy based on the PRIAM software and noticed that candidates may be proposed for an important fraction of local orphan enzymes. Finally, by studying relation between protein domains and catalyzed activities, it appears that newly discovered enzymes are mostly associated with already known enzyme domains. Thus, the exploration of the promiscuity and the multifunctional aspect of known enzyme families may solve part of the orphan enzyme issue. We conclude this review with a presentation of recent initiatives in finding proteins for orphan enzymes and in extending the enzyme world by the discovery of new activities.

Study on the Correlation between Gene Expression and Enzyme Activity of Seven Key Enzymes and Ginsenoside Content in Ginseng in Over Time in Ji'an, China.

PubMed

Yin, Juxin; Zhang, Daihui; Zhuang, Jianjian; Huang, Yi; Mu, Ying; Lv, Shaowu

2017-12-11

Panax ginseng is a traditional medicine. Fresh ginseng is one of the most important industries related to ginseng development, and fresh ginseng of varying ages has different medicinal properties. Previous research has not systematically reported the correlation between changes in key enzyme activity with changes in ginsenoside content in fresh ginseng over time. In this study, for the first time, we use ginseng samples of varying ages in Ji'an and systematically reported the changes in the activity of seven key enzymes (HMGR, FPS, SS, SE, DS, CYP450, and GT). We investigated the content of ginsenoside and gene expression of these key enzymes. Ginsenoside content was measured using HPLC. HPLC, GC-MS, and LC-MS were combined to measure the enzyme activity of the key enzymes. Quantitative PCR was used in the investigation of gene expression. By analyzing the correlation between the enzyme activity and the transcription level of the key enzymes with ginsenoside content, we found that DS and GT enzyme activities are significantly correlated with the ginsenoside content in different ages of ginseng. Our findings might provide a new strategy to discriminate between ginseng of different years. Meanwhile, this research provides important information for the in-depth study of ginsenoside biosynthesis.

Effects of protease and non-starch polysaccharide enzyme on performance, digestive function, activity and gene expression of endogenous enzyme of broilers.

PubMed

Yuan, Lin; Wang, Mingfa; Zhang, Xiaotu; Wang, Zhixiang

2017-01-01

Three hundred one-day-old male broiler chickens (Ross-308) were fed corn-soybean basal diets containing non-starch polysaccharide (NSP) enzyme and different levels of acid protease from 1 to 42 days of age to investigate the effects of exogenous enzymes on growth performance, digestive function, activity of endogenous digestive enzymes in the pancreas and mRNA expression of pancreatic digestive enzymes. For days 1-42, compared to the control chickens, average daily feed intake (ADFI) and average daily gain (ADG) were significantly enhanced by the addition of NSP enzyme in combination with protease supplementation at 40 or 80 mg/kg (p<0.05). Feed-to-gain ratio (FGR) was significantly improved by supplementation with NSP enzymes or NSP enzyme combined with 40 or 80 mg/kg protease compared to the control diet (p<0.05). Apparent digestibility of crude protein (ADCP) was significantly enhanced by the addition of NSP enzyme or NSP enzyme combined with 40 or 80 mg/kg protease (p<0.05). Cholecystokinin (CCK) level in serum was reduced by 31.39% with NSP enzyme combined with protease supplementation at 160 mg/kg (p<0.05), but the CCK level in serum was increased by 26.51% with NSP enzyme supplementation alone. After 21 days, supplementation with NSP enzyme and NSP enzyme combined with 40 or 80 mg/kg protease increased the activity of pancreatic trypsin by 74.13%, 70.66% and 42.59% (p<0.05), respectively. After 42 days, supplementation with NSP enzyme and NSP enzyme combined with 40 mg/kg protease increased the activity of pancreatic trypsin by 32.45% and 27.41%, respectively (p<0.05). However, supplementation with NSP enzyme and 80 or 160 mg/kg protease decreased the activity of pancreatic trypsin by 10.75% and 25.88%, respectively (p<0.05). The activities of pancreatic lipase and amylase were significantly higher in treated animals than they were in the control group (p<0.05). Supplementation with NSP enzyme, NSP enzyme combined with 40 or 80 mg/kg protease increased pancreatic trypsin mRNA levels by 40%, 44% and 28%, respectively. Supplementation with NSP enzyme and 160 mg/kg protease decreased pancreatic trypsin mRNA levels by 13%. Pancreatic lipase and amylase mRNA expression were significantly elevated in treated animals compared to the control group (p<0.05). These results suggest that the amount of NSP enzyme and acid protease in the diet significantly affects digestive function, endogenous digestive-enzyme activity and mRNA expression in broilers.

Oxidation of indole-3-acetic acid to oxindole-3-acetic acid by an enzyme preparation from Zea mays

NASA Technical Reports Server (NTRS)

Reinecke, D. M.; Bandurski, R. S.

1988-01-01

Indole-3-acetic acid is oxidized to oxindole-3-acetic acid by Zea mays tissue extracts. Shoot, root, and endosperm tissues have enzyme activities of 1 to 10 picomoles per hour per milligram protein. The enzyme is heat labile, is soluble, and requires oxygen for activity. Cofactors of mixed function oxygenase, peroxidase, and intermolecular dioxygenase are not stimulatory to enzymic activity. A heat-stable, detergent-extractable component from corn enhances enzyme activity 6- to 10-fold. This is the first demonstration of the in vitro enzymic oxidation of indole-3-acetic acid to oxindole-3-acetic acid in higher plants.

Activation of immobilized enzymes by acoustic wave resonance oscillation.

PubMed

Nishiyama, Hiroshi; Watanabe, Tomoya; Inoue, Yasunobu

2014-12-01

Acoustic wave resonance oscillation has been used successfully in the development of methods to activate immobilized enzyme catalysts. In this study, resonance oscillation effects were demonstrated for enzyme reactions on galactose oxidase (GAD), D-amino acid oxidase (DAAO), and L-amino acid oxidase (LAAO), all of which were immobilized covalently on a ferroelectric lead zirconate titanate (PZT) device that could generate thickness-extensional resonance oscillations (TERO) of acoustic waves. For galactose oxidation on immobilized GAD in a microreactor, TERO generation immediately increased enzyme activity 2- to 3-fold. Eliminating TERO caused a slight decrease in the activity, with ∼90% of the enhanced activity retained while the reaction proceeded. Contact of the enhanced enzyme with a galactose-free solution caused almost complete reversion of the activity to the original low level before TERO generation, indicating that, not only TERO-induced GAD activation, but also preservation of the increased activity, required a galactose substrate. Similar activity changes with TERO were observed for enzyme reactions on DAAO and LAAO. Kinetic analysis demonstrated that TERO helped strengthen the interactions of the immobilized enzyme with the reactant substrate and promoted formation of an activation complex. Copyright © 2014 Elsevier Inc. All rights reserved.

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.

Physics-based enzyme design: predicting binding affinity and catalytic activity.

PubMed

Sirin, Sarah; Pearlman, David A; Sherman, Woody

2014-12-01

Computational enzyme design is an emerging field that has yielded promising success stories, but where numerous challenges remain. Accurate methods to rapidly evaluate possible enzyme design variants could provide significant value when combined with experimental efforts by reducing the number of variants needed to be synthesized and speeding the time to reach the desired endpoint of the design. To that end, extending our computational methods to model the fundamental physical-chemical principles that regulate activity in a protocol that is automated and accessible to a broad population of enzyme design researchers is essential. Here, we apply a physics-based implicit solvent MM-GBSA scoring approach to enzyme design and benchmark the computational predictions against experimentally determined activities. Specifically, we evaluate the ability of MM-GBSA to predict changes in affinity for a steroid binder protein, catalytic turnover for a Kemp eliminase, and catalytic activity for α-Gliadin peptidase variants. Using the enzyme design framework developed here, we accurately rank the most experimentally active enzyme variants, suggesting that this approach could provide enrichment of active variants in real-world enzyme design applications. © 2014 Wiley Periodicals, Inc.

Stability improvement of immobilized lactoperoxidase using polyaniline polymer.

PubMed

Jafary, Fariba; Kashanian, Soheila; Sharieat, Ziadin Samsam; Jafary, Farzaneh; Omidfar, Kobra; Paknejad, Maliheh

2012-12-01

Enzyme engineering via immobilization techniques is perfectly compatible against the other chemical or biological approximate to improve enzyme functions and stability. In this study lactoperoxidase was immobilized onto polyaniline polymer activated with glutaraldehyde as a bifunctional agent, to improve enzyme properties. Polyaniline polymer was used due its unique physical and chemical properties to immobilize lactoperoxidase (LPO). The optimum activity of immobilized LPO was observed at pH 6 and 55 °C, which has been increased about 10 °C for the immobilized enzyme. The immobilized enzyme maintained absolutely active for 60 days whereas the native enzyme lost 80 % of its initial activity within this period of time. Moreover, the immobilized enzyme can be reused for several times without loss of activity. The kinetic parameter studies showed slight differences between free and immobilized enzymes. The K(m) and K(m.app) were calculated to be 0.6 and 0.4; also V(max) and V(max.app) were 1.3 and 0.9 respectively.

Engineering a hyper-catalytic enzyme by photo-activated conformation modulation

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

Agarwal, Pratul K

2012-01-01

Enzyme engineering for improved catalysis has wide implications. We describe a novel chemical modification of Candida antarctica lipase B that allows modulation of the enzyme conformation to promote catalysis. Computational modeling was used to identify dynamical enzyme regions that impact the catalytic mechanism. Surface loop regions located distal to active site but showing dynamical coupling to the reaction were connected by a chemical bridge between Lys136 and Pro192, containing a derivative of azobenzene. The conformational modulation of the enzyme was achieved using two sources of light that alternated the azobenzene moiety in cis and trans conformations. Computational model predicted thatmore » mechanical energy from the conformational fluctuations facilitate the reaction in the active-site. The results were consistent with predictions as the activity of the engineered enzyme was found to be enhanced with photoactivation. Preliminary estimations indicate that the engineered enzyme achieved 8-52 fold better catalytic activity than the unmodulated enzyme.« less

Dipeptidyl peptidase IV in angiotensin-converting enzyme inhibitor associated angioedema.

PubMed

Byrd, James Brian; Touzin, Karine; Sile, Saba; Gainer, James V; Yu, Chang; Nadeau, John; Adam, Albert; Brown, Nancy J

2008-01-01

Angioedema is a potentially life-threatening adverse effect of angiotensin-converting enzyme inhibitors. Bradykinin and substance P, substrates of angiotensin-converting enzyme, increase vascular permeability and cause tissue edema in animals. Studies indicate that amino-terminal degradation of these peptides, by aminopeptidase P and dipeptidyl peptidase IV, may be impaired in individuals with angiotensin-converting enzyme inhibitor-associated angioedema. This case-control study tested the hypothesis that dipeptidyl peptidase IV activity and antigen are decreased in sera of patients with a history of angiotensin-converting enzyme inhibitor-associated angioedema. Fifty subjects with a history of angiotensin-converting enzyme inhibitor-associated angioedema and 176 angiotensin-converting enzyme inhibitor-exposed control subjects were ascertained. Sera were assayed for angiotensin-converting enzyme activity, aminopeptidase P activity, aminopeptidase N activity, dipeptidyl peptidase IV activity, and antigen and the ex vivo degradation half-lives of bradykinin, des-Arg(9)-bradykinin, and substance P in a subset. The prevalence of smoking was increased and of diabetes decreased in case versus control subjects. Overall, dipeptidyl peptidase IV activity (26.6+/-7.8 versus 29.6+/-7.3 nmol/mL per minute; P=0.026) and antigen (465.8+/-260.8 versus 563.1+/-208.6 ng/mL; P=0.017) were decreased in sera from individuals with angiotensin-converting enzyme inhibitor-associated angioedema compared with angiotensin-converting enzyme inhibitor-exposed control subjects without angioedema. Dipeptidyl peptidase IV activity (21.5+/-4.9 versus 29.8+/-6.7 nmol/mL per minute; P=0.001) and antigen (354.4+/-124.7 versus 559.8+/-163.2 ng/mL; P=0.003) were decreased in sera from cases collected during angiotensin-converting enzyme inhibition but not in the absence of angiotensin-converting enzyme inhibition. The degradation half-life of substance P correlated inversely with dipeptidyl peptidase IV antigen during angiotensin-converting enzyme inhibition. Environmental or genetic factors that reduce dipeptidyl peptidase IV activity may predispose individuals to angioedema.

Dipeptidyl Peptidase IV in Angiotensin-Converting Enzyme Inhibitor–Associated Angioedema

PubMed Central

Byrd, James Brian; Touzin, Karine; Sile, Saba; Gainer, James V.; Yu, Chang; Nadeau, John; Adam, Albert; Brown, Nancy J.

2009-01-01

Angioedema is a potentially life-threatening adverse effect of angiotensin-converting enzyme inhibitors. Bradykinin and substance P, substrates of angiotensin-converting enzyme, increase vascular permeability and cause tissue edema in animals. Studies indicate that amino-terminal degradation of these peptides, by aminopeptidase P and dipeptidyl peptidase IV, may be impaired in individuals with angiotensin-converting enzyme inhibitor–associated angioedema. This case-control study tested the hypothesis that dipeptidyl peptidase IV activity and antigen are decreased in sera of patients with a history of angiotensin-converting enzyme inhibitor–associated angioedema. Fifty subjects with a history of angiotensin-converting enzyme inhibitor–associated angioedema and 176 angiotensin-converting enzyme inhibitor–exposed control subjects were ascertained. Sera were assayed for angiotensin-converting enzyme activity, aminopeptidase P activity, aminopeptidase N activity, dipeptidyl peptidase IV activity, and antigen and the ex vivo degradation half-lives of bradykinin, des-Arg9-bradykinin, and substance P in a subset. The prevalence of smoking was increased and of diabetes decreased in case versus control subjects. Overall, dipeptidyl peptidase IV activity (26.6±7.8 versus 29.6±7.3 nmol/mL per minute; P=0.026) and antigen (465.8±260.8 versus 563.1±208.6 ng/mL; P=0.017) were decreased in sera from individuals with angiotensin-converting enzyme inhibitor–associated angioedema compared with angiotensin-converting enzyme inhibitor–exposed control subjects without angioedema. Dipeptidyl peptidase IV activity (21.5±4.9 versus 29.8±6.7 nmol/mL per minute; P=0.001) and antigen (354.4±124.7 versus 559.8±163.2 ng/mL; P=0.003) were decreased in sera from cases collected during angiotensin-converting enzyme inhibition but not in the absence of angiotensin-converting enzyme inhibition. The degradation half-life of substance P correlated inversely with dipeptidyl peptidase IV antigen during angiotensin-converting enzyme inhibition. Environmental or genetic factors that reduce dipeptidyl peptidase IV activity may predispose individuals to angioedema. PMID:18025295

ORENZA: a web resource for studying ORphan ENZyme activities

PubMed Central

Lespinet, Olivier; Labedan, Bernard

2006-01-01

Background Despite the current availability of several hundreds of thousands of amino acid sequences, more than 36% of the enzyme activities (EC numbers) defined by the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology (NC-IUBMB) are not associated with any amino acid sequence in major public databases. This wide gap separating knowledge of biochemical function and sequence information is found for nearly all classes of enzymes. Thus, there is an urgent need to explore these sequence-less EC numbers, in order to progressively close this gap. Description We designed ORENZA, a PostgreSQL database of ORphan ENZyme Activities, to collate information about the EC numbers defined by the NC-IUBMB with specific emphasis on orphan enzyme activities. Complete lists of all EC numbers and of orphan EC numbers are available and will be periodically updated. ORENZA allows one to browse the complete list of EC numbers or the subset associated with orphan enzymes or to query a specific EC number, an enzyme name or a species name for those interested in particular organisms. It is possible to search ORENZA for the different biochemical properties of the defined enzymes, the metabolic pathways in which they participate, the taxonomic data of the organisms whose genomes encode them, and many other features. The association of an enzyme activity with an amino acid sequence is clearly underlined, making it easy to identify at once the orphan enzyme activities. Interactive publishing of suggestions by the community would provide expert evidence for re-annotation of orphan EC numbers in public databases. Conclusion ORENZA is a Web resource designed to progressively bridge the unwanted gap between function (enzyme activities) and sequence (dataset present in public databases). ORENZA should increase interactions between communities of biochemists and of genomicists. This is expected to reduce the number of orphan enzyme activities by allocating gene sequences to the relevant enzymes. PMID:17026747

Remote enzyme activation using gold coated magnetite as antennae for radio frequency fields

NASA Astrophysics Data System (ADS)

Collins, Christian B.; Ackerson, Christopher J.

2018-02-01

The emerging field of remote enzyme activation, or the ability to remotely turn thermophilic increase enzyme activity, could be a valuable tool for understanding cellular processes. Through exploitation of the temperature dependence of enzymatic processes and high thermal stability of thermophilic enzymes these experiments utilize nanoparticles as `antennae' that convert radiofrequency (RF) radiation into local heat, increasing activity of the enzymes without increasing the temperature of the surrounding bulk solution. To investigate this possible tool, thermolysin, a metalloprotease was covalently conjugated to 4nm gold coated magnetite particles via peptide bond formation with the protecting ligand shell. RF stimulated protease activity at 17.76 MHz in a solenoid shaped antenna, utilizing both electric and magnetic field interactions was investigated. On average 40 percent higher protease activity was observed in the radio frequency fields then when bulk heating the sample to the same temperature. This is attributed to electrophoretic motion of the nanoparticle enzyme conjugates and local regions of heat generated by the relaxation of the magnetite cores with the oscillating field. Radio frequency local heating of nanoparticles conjugated to enzymes as demonstrated could be useful in the activation of specific enzymes in complex cellular environments.

High activity CAZyme cassette for improving biomass degradation in thermophiles

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

Brunecky, Roman; Chung, Daehwan; Sarai, Nicholas S.

Currently, Thermophilic microorganisms and their enzymes offer several advantages for industrial application over their mesophilic counterparts. For example, a hyperthermophilic anaerobe, Caldicellulosiruptor bescii, was recently isolated from hot springs in Kamchatka, Siberia, and shown to have very high cellulolytic activity. Additionally, it is one of a few microorganisms being considered as viable candidates for consolidated bioprocessing applications. Moreover, C. bescii is capable of deconstructing plant biomass without enzymatic or chemical pretreatment. This ability is accomplished by the production and secretion of free, multi-modular and multi-functional enzymes, one of which, CbCel9A/Cel48A also secretion of free, multi-modular and multi-functional enzymes, one ofmore » which, CbCel9A/Cel48A also known as CelA, is able to outperform enzymes found in commercial enzyme preparations. Furthermore, the complete C. bescii exoproteome is extremely thermostable and highly active at elevated temperatures, unlike commercial fungal cellulases. Understanding the functional diversity of enzymes in the C. bescii exoproteome and how inter-molecular synergy between them confers C. bescii with its high cellulolytic activity is an important endeavor to enable the production more efficient biomass degrading enzyme formulations and in turn, better cellulolytic industrial microorganisms. We found that the combination of three or four of the most highly expressed enzymes in the C. bescii exoproteome exhibits such synergistic activity. For example, some discrete combinations of these enzymes mimic and even improve upon the activity of the exoproteome, even though some of the enzymes lack significant activity on their own. We have demonstrated that it is possible to replicate the cellulolytic activity of the native C. bescii exoproteome utilizing a minimal gene set, and that these minimal gene sets are more active than the whole exoproteome. In the future, this may lead to more simplified and efficient cellulolytic enzyme preparations or yield improvements when these enzymes are expressed in microorganisms engineered for consolidated bioprocessing.« less

High activity CAZyme cassette for improving biomass degradation in thermophiles

DOE PAGES

Brunecky, Roman; Chung, Daehwan; Sarai, Nicholas S.; ...

2018-02-01

Currently, Thermophilic microorganisms and their enzymes offer several advantages for industrial application over their mesophilic counterparts. For example, a hyperthermophilic anaerobe, Caldicellulosiruptor bescii, was recently isolated from hot springs in Kamchatka, Siberia, and shown to have very high cellulolytic activity. Additionally, it is one of a few microorganisms being considered as viable candidates for consolidated bioprocessing applications. Moreover, C. bescii is capable of deconstructing plant biomass without enzymatic or chemical pretreatment. This ability is accomplished by the production and secretion of free, multi-modular and multi-functional enzymes, one of which, CbCel9A/Cel48A also secretion of free, multi-modular and multi-functional enzymes, one ofmore » which, CbCel9A/Cel48A also known as CelA, is able to outperform enzymes found in commercial enzyme preparations. Furthermore, the complete C. bescii exoproteome is extremely thermostable and highly active at elevated temperatures, unlike commercial fungal cellulases. Understanding the functional diversity of enzymes in the C. bescii exoproteome and how inter-molecular synergy between them confers C. bescii with its high cellulolytic activity is an important endeavor to enable the production more efficient biomass degrading enzyme formulations and in turn, better cellulolytic industrial microorganisms. We found that the combination of three or four of the most highly expressed enzymes in the C. bescii exoproteome exhibits such synergistic activity. For example, some discrete combinations of these enzymes mimic and even improve upon the activity of the exoproteome, even though some of the enzymes lack significant activity on their own. We have demonstrated that it is possible to replicate the cellulolytic activity of the native C. bescii exoproteome utilizing a minimal gene set, and that these minimal gene sets are more active than the whole exoproteome. In the future, this may lead to more simplified and efficient cellulolytic enzyme preparations or yield improvements when these enzymes are expressed in microorganisms engineered for consolidated bioprocessing.« less

Role of Rab5 in the formation of macrophage-derived foam cell.

PubMed

Chan, Lokwern; Hong, Jin; Pan, Junjie; Li, Jian; Wen, Zhichao; Shi, Haiming; Ding, Jianping; Luo, Xinping

2017-09-12

Foam cells play a key role in the occurrence and pathogenesis of atherosclerosis. Its formation starts with the ingestion of oxidized low-density lipoprotein (oxLDL). The process is associated with Ras related protein in brain 5 (Rab5) which plays a critical role in regulating endocytosis and early endosomal trafficking. Base on this, we presumed that Rab5 might participate in the maturation of foam cell. The aim of this study is to investigate the effect of Rab5 on macrophage cholesterol during the evolvement of macrophage when induced by oxLDL to the formation of foam cell. Immunohistochemistry was performed to analyze the distribution of macrophages and Rab5 in atherosclerotic plaque. RNA inteference study and transfection of inactive mutant (GFP-Rab5-S34N) and active mutant (GFP-Rab5-Q79L) in U937-derived macrophage were utilized to investigate the impact of Rab5 on the process of macrophage cholesterol, which could be detected by oil red O staining, determination of intracellular lipid content, filipin staining, nile red staining and the costaining of early endosome antigen-1 (EEA-1) and 1,1'-dioctadecyl-3,3,3',3'-tetramethylin dicarbocyanine (Dil)-labelled oxLDL (Dil-oxLDL). Rab5 was found abundantly localized in macrophage rich areas of human atherosclerotic lesions. On the foam cell study, the expression of Rab5 was increased after the incubation of oxLDL. The inteference study indicated the depletion of Rab5 led to the decreases of oil red O staining areas, total cholesterol and cholesterol esters in U937-derived marophages. Moreover, the fluorescence intensity of filipin and nile red staining were lower in GFP-Rab5-S34N as compared with GFP-Rab5-Q79L. The confocal study demonstrated less Dil-oxLDL was internalized in GFP-Rab5-S34N as compared with GFP-Rab5-Q79L; the result showed also the decrease in colocalization of internalized Dil-oxLDL and EEA-1 for GFP-Rab5-S34N as compared with GFP-Rab5-Q79L. Rab5 plays an important role in modulating the intracellular cholesterol of macrophages and consequently mediating the formation of foam cells.

Mapping specific soil functions based on digital soil property maps

NASA Astrophysics Data System (ADS)

Pásztor, László; Fodor, Nándor; Farkas-Iványi, Kinga; Szabó, József; Bakacsi, Zsófia; Koós, Sándor

2016-04-01

Quantification of soil functions and services is a great challenge in itself even if the spatial relevance is supposed to be identified and regionalized. Proxies and indicators are widely used in ecosystem service mapping. Soil services could also be approximated by elementary soil features. One solution is the association of soil types with services as basic principle. Soil property maps however provide quantified spatial information, which could be utilized more versatilely for the spatial inference of soil functions and services. In the frame of the activities referred as "Digital, Optimized, Soil Related Maps and Information in Hungary" (DOSoReMI.hu) numerous soil property maps have been compiled so far with proper DSM techniques partly according to GSM.net specifications, partly by slightly or more strictly changing some of its predefined parameters (depth intervals, pixel size, property etc.). The elaborated maps have been further utilized, since even DOSoReMI.hu was intended to take steps toward the regionalization of higher level soil information (secondary properties, functions, services). In the meantime the recently started AGRAGIS project requested spatial soil related information in order to estimate agri-environmental related impacts of climate change and support the associated vulnerability assessment. One of the most vulnerable services of soils in the context of climate change is their provisioning service. In our work it was approximated by productivity, which was estimated by a sequential scenario based crop modelling. It took into consideration long term (50 years) time series of both measured and predicted climatic parameters as well as accounted for the potential differences in agricultural practice and crop production. The flexible parametrization and multiple results of modelling was then applied for the spatial assessment of sensitivity, vulnerability, exposure and adaptive capacity of soils in the context of the forecasted changes in climatic conditions in the Carpathian Basin. In addition to soil fertility, degradation risk due to N-leaching was also assessed by the model runs by taking into account the movement of nitrate in the profile during the simulated periods. Our paper will present the resulted national maps and some conclusions drawn from the experiences. Acknowledgement: Our work was supported by Iceland, Liechtenstein and Norway through the EEA Grants and the REC (Project No: EEA C12-12) and the Hungarian National Scientific Research Foundation (OTKA, Grant No. K105167).

Optimization of process variables by central composite design for the immobilization of urease enzyme on functionalized gold nanoparticles for various applications.

PubMed

Talat, Mahe; Singh, Ashwani Kumar; Srivastava, O N

2011-08-01

In the present study, enzyme urease has been immobilized on amine-functionalized gold nanoparticles (AuNPs). AuNPs were synthesized using natural precursor, i.e., clove extract and amine functionalized through 0.004 M L: -cysteine. Enzyme (urease) was extracted and purified from the vegetable waste, i.e., seeds of pumpkin to apparent homogeneity (sp. activity 353 U/mg protein). FTIR spectroscopy and transmission electron microscopy was used to characterize the immobilized enzyme. The immobilized enzyme exhibited enhanced activity as compared with the enzyme in the solution, especially, at lower enzyme concentration. Based on the evaluation of activity assay of the immobilized enzyme, it was found that the immobilized enzyme was quite stable for about a month and could successfully be used even after eight cycles having enzyme activity of about 47%. In addition to this central composite design (CCD) with the help of MINITAB version 15 Software was utilized to optimize the process variables viz., pH and temperature affecting the enzyme activity upon immobilization on AuNPs. The results predicted by the design were found in good agreement (R2 = 96.38%) with the experimental results indicating the applicability of proposed model. The multiple regression analysis and ANOVA showed the individual and cumulative effect of pH and temperature on enzyme activity indicating that the activity increased with the increase of pH up to 7.5 and temperature 75 °C. The effects of each variables represented by main effect plot, 3D surface plot, isoresponse contour plot and optimized plot were helpful in predicting results by performing a limited set of experiments.

Metagenomics as a Tool for Enzyme Discovery: Hydrolytic Enzymes from Marine-Related Metagenomes.

PubMed

Popovic, Ana; Tchigvintsev, Anatoly; Tran, Hai; Chernikova, Tatyana N; Golyshina, Olga V; Yakimov, Michail M; Golyshin, Peter N; Yakunin, Alexander F

2015-01-01

This chapter discusses metagenomics and its application for enzyme discovery, with a focus on hydrolytic enzymes from marine metagenomic libraries. With less than one percent of culturable microorganisms in the environment, metagenomics, or the collective study of community genetics, has opened up a rich pool of uncharacterized metabolic pathways, enzymes, and adaptations. This great untapped pool of genes provides the particularly exciting potential to mine for new biochemical activities or novel enzymes with activities tailored to peculiar sets of environmental conditions. Metagenomes also represent a huge reservoir of novel enzymes for applications in biocatalysis, biofuels, and bioremediation. Here we present the results of enzyme discovery for four enzyme activities, of particular industrial or environmental interest, including esterase/lipase, glycosyl hydrolase, protease and dehalogenase.

Ectomycorrhizal Fungal Communities and Enzymatic Activities Vary across an Ecotone between a Forest and Field.

PubMed

Rúa, Megan A; Moore, Becky; Hergott, Nicole; Van, Lily; Jackson, Colin R; Hoeksema, Jason D

2015-08-28

Extracellular enzymes degrade macromolecules into soluble substrates and are important for nutrient cycling in soils, where microorganisms, such as ectomycorrhizal (ECM) fungi, produce these enzymes to obtain nutrients. Ecotones between forests and fields represent intriguing arenas for examining the effect of the environment on ECM community structure and enzyme activity because tree maturity, ECM composition, and environmental variables may all be changing simultaneously. We studied the composition and enzymatic activity of ECM associated with loblolly pine (Pinus taeda) across an ecotone between a forest where P. taeda is established and an old field where P. taeda saplings had been growing for <5 years. ECM community and environmental characteristics influenced enzyme activity in the field, indicating that controls on enzyme activity may be intricately linked to the ECM community, but this was not true in the forest. Members of the Russulaceae were associated with increased phenol oxidase activity and decreased peroxidase activity in the field. Members of the Atheliaceae were particularly susceptible to changes in their abiotic environment, but this did not mediate differences in enzyme activity. These results emphasize the complex nature of factors that dictate the distribution of ECM and activity of their enzymes across a habitat boundary.

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.

Evaluation of 2-Thioxo-2,3,5,6,7,8-hexahydropyrimido[4,5-d]pyrimidin-4(1H)-one analogues as GAA Activators

PubMed Central

Marugan, Juan J.; Zheng, Wei; Motabar, Omid; Southall, Noel; Goldin, Ehud; Sidransky, Ellen; Aungst, Ronald A.; Liu, Ke; Sadhukhan, Subir Kumar; Austin, Christopher P.

2010-01-01

Pompe disease is a lysosomal storage disease (LSD) caused by a deficiency in the lysosomal enzyme acid α-glucosidase. In several LSDs, enzyme inhibitors have been used as small molecule chaperones to facilitate and increase the translocation of mutant protein from the endoplasmic reticulum to the lysosome. Enzyme activators with chaperone activity would be even more desirable as they would not inhibit the enzyme after translocation and might potentiate the activity of the enzyme that is successfully translocated. Herein we report our initial findings of a new series of acid α-glucosidase activators. PMID:20206419

Evaluation of 2-thioxo-2,3,5,6,7,8-hexahydropyrimido[4,5-d]pyrimidin-4(1H)-one analogues as GAA activators.

PubMed

Marugan, Juan J; Zheng, Wei; Motabar, Omid; Southall, Noel; Goldin, Ehud; Sidransky, Ellen; Aungst, Ronald A; Liu, Ke; Sadhukhan, Subir Kumar; Austin, Christopher P

2010-05-01

Pompe disease is a lysosomal storage disease (LSD) caused by a deficiency in the lysosomal enzyme acid alpha-glucosidase. In several LSDs, enzyme inhibitors have been used as small molecule chaperones to facilitate and increase the translocation of mutant protein from the endoplasmic reticulum to the lysosome. Enzyme activators with chaperone activity would be even more desirable as they would not inhibit the enzyme after translocation and might potentiate the activity of the enzyme that is successfully translocated. Herein we report our initial findings of a new series of acid alpha-glucosidase activators.

Nanoarmoring of Enzymes by Interlocking in Cellulose Fibers With Poly(Acrylic Acid).

PubMed

Riccardi, Caterina M; Kasi, Rajeswari M; Kumar, Challa V

2017-01-01

A simple method for interlocking glucose oxidase (GOx) and horseradish peroxidase (HRP) in cellulose fibers using poly(acrylic acid) (PAA) as an armor around the enzyme, without any need for activation of the cellulose support, is reported here. The resulting enzyme paper is an inexpensive, stable, simple, wearable, and washable biosensor. PAA functions as a multifunctional tether to interlock the enzyme molecules around the paper fibers so that the enzymes are protected against thermal/chemical denaturation and not released from the paper when washed with a detergent. The decreased conformational entropy of the interlocked enzyme protected by the nanoarmor is likely responsible for increased enzyme stability to heat and chemical denaturants (retained ≥70 percent enzyme activity after washing with urea or SDS for 30min), and the polymer protects the enzyme against inactivation by proteases, bacteria, inhibitors, etc. The kinetics of the interlocked enzyme were similar to that of the enzyme in solution. The V max was 6(±0.5)mM per minute before washing, then increased slightly to 9(±1.4)mM per minute after washing with water. The K m was 22(±6.4mM), which was slightly higher compared to GOx in solution (25-27mM). Because the surface area of the paper does not limit the enzyme loading, about 20% of enzyme was successfully loaded onto the paper (0.2g enzyme per gram of paper), and ≥95% of the enzyme was retained after washing. Interlocking works with other enzymes such as laccase, where ≥60% of the enzyme activity is retained. This novel methodology provides a low cost, simple, modular approach of achieving high enzyme loadings in ordinary filter paper, not limited by cellulose surface area, and there has been no need for complex methods of enzyme engineering or toxic methods of activation of the solid support to prepare highly active biocatalysts. © 2017 Elsevier Inc. All rights reserved.

A Model for the Management of Technical Risk in New Technology Development Programs

DTIC Science & Technology

1989-09-01

Heppenheimer , T. A . "Requiem for a Heavy-weight," Air and Snace/Smithsonian, 4: 50-55-- June/July 1989). 91. Jane’s Information Group, Inc. Jane’s...8217-- A MODEL FOR THE MANAGEMENT OF TECHNICAL RISK IN NEW TECHNOLOGY DEVELOPMENT PROGRAMS THESIS Peter B. King Captain, USA~F A FIT /G SM /L SR /89...S-22 1 DISI~1UT0N TAEMEN𔃻 A , S’o11 _44110h, eea;EL ECTE EApproVed 0 ulcrl~;DC2 0 1989 Dis-,irbuticf Unlimited D DEPARTMENT OF THE AIR FORCE-- - AIR

Evaluation of Visibility Sensors at the Eglin Air Force Base Climatic Chamber.

DTIC Science & Technology

1983-10-01

s o t LI a of to -NAIC 6/81- -11 * VIow:1710 tons e- I: l - llO 26 5 6 Os *15 eeas IIg " 25 l5 .0 36 15 lOI (C) RYC (d) FIGURE 5-15. HOMOGENEITY OF...4 inch thick white plastic scattering disk. Since the exact attenuation of a filter can vary by as much as a factor of two, there was no way to...translucent plastic disk as scatterer and was found to be unchanged. The difference in calibration between Event 1 (slope - 0.88) and Event 2 (slope

Achieving Generality over Conditions: Combining the Multitrait Multimethod Matrix and the Representative Design of Experiments.

DTIC Science & Technology

1984-08-02

We take that aim to have been achieved in principle (no one has challenged it), if not in practice. We aim therefore to build upon that achievement by...NJ + (J + C~ C + 2 C EU E E 4-)C o.C ~ ~+ (NJ + (N Er E o - ~.. ~____ U) C fl - C C C CE E le E0 I0 U.- E EEa E - Ui Ei . U (alKE E E E 2 2- 2 E Ca...RESEARCH Engineering Psychology Group • TECHNICAL REPORTS DISTRIBUTION LIST OSD Department of the Nay CAPT Paul R. Chatelier Tactical Development

[Marketing of medicinal products in the European Community. The Mutual Recognition and Decentralised Procedures].

PubMed

Bachmann, Peter

2008-07-01

The Mutual Recognition Procedure (MRP) and the Decentralised Procedure (DCP), which were first established in late 2005, can be regarded as the backbone for marketing authorisation of medicinal products in the European Community (EC) and the European Economic Area (EEA). Both procedures are compared and advantages and disadvantages are discussed. However, the focus is more related to current developments than the detailed comparison of both procedures. The role of the Coordination Group for Mutual Recognition and Decentralised Procedures-Human (CMD(h)) and the decision making process in relation to the MRP and DCP is also discussed.

[Errors and dangers in intestinal sutures and anastomoses using stapler suture instruments (EEA, TA55, TA90, GIA)].

PubMed

Gögler, E

1985-01-01

In different tables the most important faults with enteral sutures and anastomoses in general and at special operations are demonstrated: end-to-end anastomoses with congruent diameter, anastomoses with different diameters, B I, B II, low anterior resection, esophago-jejunostomy. Only if the surgeon has experience in standard technique, faults and risks with mechanical staplers and manual sutures, the advantage-progress of staplers will be effective avoiding special risks. Surgeons without experience may produce real catastrophes which may turn out hopeless without training in manual suture technique.

A Bibliography with Abstracts of U. S. Army Engineer Waterways Experiment Station Publications Related to Vehicle Mobility

DTIC Science & Technology

1976-08-01

Schreiner M-75-5 Jun 1975 Comparison of the Ride Qualities of Standard AD A013 986 M6OAI and Hybrid (Tube-Over-Bar) M60A1E3 Tanks, by N. R. Murphy Jr. and J...toingandowedtes snowere ondced wth eea hee trakedmiltar veicls a a arityof ficegaiiy ned alnd weet coirstaryapici snowconitins. he 955test wee...T- or ORIPOAT & PEltOC COVERED COMPARISON OF THE RIDE QUJALITIE.S OF STANL)AKII) Final report M60AI AN!) HYBRID (TUbE-OVER-BAR) M6OAlE3 TANKS

Antioxidant and Antiproliferative Activities of Leaf Extracts from Plukenetia volubilis Linneo (Euphorbiaceae)

PubMed Central

Nascimento, Ana Karina Lima; Melo-Silveira, Raniere Fagundes; Dantas-Santos, Nednaldo; Fernandes, Júlia Morais; Zucolotto, Silvana Maria; Rocha, Hugo Alexandre Oliveira; Scortecci, Katia Castanho

2013-01-01

Plukenetia volubilis Linneo, or Sacha inca, is an oleaginous plant from the Euphorbiaceae family. The aim of this work was to perform a chemical and biological analysis of different leaf extracts from P. volubilis such as aqueous extract (AEL), methanol (MEL), ethanol (EEL), chloroform (CEL), and hexane (HEL). Thin layer chromatography analysis revealed the presence of phenolic compounds, steroids, and/or terpenoídes. Furthermore, the antioxidant activities were analyzed by in vitro assays and their effects on cell lineages by in vivo assays. The Total Antioxidant Capacity (TCA) was expressed as equivalent ascorbic acid (EEA/g) and it was observed that the extracts showed values ranging from 59.31 to 97.76 EAA/g. Furthermore, the DPPH assay values ranged from 62.8% to 88.3%. The cell viability assay showed that the extracts were able to reduce viability from cancer cells such as HeLa and A549 cells. The extracts MEL and HEL (250 µg/mL) were able to reduce the proliferation of HeLa cells up to 54.3% and 48.5%, respectively. The flow cytometer results showed that these extracts induce cell death via the apoptosis pathway. On the other hand, the extracts HEL and AEL were able to induce cell proliferation of normal fibroblast 3T3 cells. PMID:24159355

Unfolding and inactivation during thermal denaturation of an enzyme that exhibits phytase and acid phosphatase activities.

PubMed

Wang, Xiao-Yun; Meng, Fan-Guo; Zhou, Hai-Meng

2004-03-01

The thermostability of an enzyme that exhibits phytase and acid phosphatase activities was studied. Kinetics of inactivation and unfolding during thermal denaturation of the enzyme were compared. The loss of phytase activity on thermal denaturation is most suggestive of a reversible process. As for acid phosphatase activities, an interesting phenomenon was observed; there are two phases in thermal inactivation: when the temperature was between 45 and 50 degrees C, the thermal inactivation could be characterized as an irreversible inactivation which had some residual activity and when the temperature was above 55 degrees C, the thermal inactivation could be characterized as an irreversible process which had no residual activity. The microscopic rate constants for the free enzyme and substrate-enzyme complex were determined by Tsou's method [Adv. Enzymol. Relat. Areas Mol. Biol. 61 (1988) 381]. Fluorescence analyses indicate that when the enzyme was treated at temperatures below 60 degrees C for 60 min, the conformation of the enzyme had no detectable change; when the temperatures were above 60 degrees C, some fluorescence red-shift could be observed with a decrease in emission intensity. The inactivation rates (k(+0)) of free enzymes were faster than those of conformational changes during thermal denaturation at the same temperature. The rapid inactivation and slow conformational changes of phytase during thermal denaturation suggest that inactivation occurs before significant conformational changes of the enzyme, and the active site of this enzyme is situated in a relatively fragile region which makes the active site more flexible than the molecule as a whole.

Drought and water scarcity indicators: experience and operational applications in italian basins

NASA Astrophysics Data System (ADS)

Mazzanti, Bernardo; Checcucci, Gaia; Monacelli, Giuseppina; Puma, Francesco; Vezzani, Claudia

2013-04-01

In the framework of River Basin Managment Plans (RBMPs), according to the Water Framework Directive, prevention and mitigation of water scarcity and droughts are some of the most challenging tasks. In the last ten years Italy experienced the highest ever observed frequency of occurrence of drought/water scarcity events. As an example, the damages for the latest, country-wide drought event of summer 2012 exceeded one billion euros. On the other hand, according to the more recent reports on the risks of extreme events, there is evidence, providing a basis for medium confidence, that droughts will intensify over the coming century in southern Europe and in the Mediterranean region (IPCC 2012). Monitoring actions are necessary and extremely effective to "feel the pulse of the situation" about both natural availability and anthropic use of freshwater resources. In this context, referring to the Programmes of Measures of RBMPs, italian River Basin Authorities (RBA) are tackling the issue at different spatial scales, planning an operational use of different indicators, between theme the Water Exploitation Index (EEA, 2009) and some statistical indicators. In this context, Po and Arno River Basin authorities, with the support of ISPRA, are directly involved in the experimental application of some significant indicators combining climatic, hydrological and anthropic factors affecting water availability. Planning and operational experiences for the two main basins (Po and Arno) and for a list of smaller scale subbasins are presented, with a detailed description of data needs, range of application, spatial and temporal scale issues, and threshold definition. For each indicator, a critical analysis of strenghts and weaknesses (at data and response level) is reported, with particular regard to the feasibility of its use within water management and water planning actions at the river basin and district scale. Tests were carried out for the whole Po River and Northern Appennines districts, and for the basin of Arno, Trebbia and Oglio rivers, and included the following indicators: modified Water Exploitation Index (WEI+) calculated at annual and monthly scale (ETC/ICM, 2012); Standardized Precipitation Index (SPI); Standardized Runoff Index (SRI). The results confirmed the need for the analysis of the indicators different spatial and temporal scales to understand their meaning in the local regional context considered. It was clearly shown that sub-annual patterns of water availability and water use play a fundamental role in the indicators meaning, and should be considered in their operational application. References: IPCC, 2012 - Special Report on Managing the Risks of Extreme Events and Disasters to Advance Climate Change Adaptation, 2012 ETC/ICM, 2012 - Vulnerability to Water Scarcity and Drought in Europe - Background thematic report for EEA water 2012 report, EEA/NSV/10/002

Developmental and hormone-induced changes of mitochondrial electron transport chain enzyme activities during the last instar larval development of maize stem borer, Chilo partellus (Lepidoptera: Crambidae).

PubMed

VenkatRao, V; Chaitanya, R K; Naresh Kumar, D; Bramhaiah, M; Dutta-Gupta, A

2016-12-01

The energy demand for structural remodelling in holometabolous insects is met by cellular mitochondria. Developmental and hormone-induced changes in the mitochondrial respiratory activity during insect metamorphosis are not well documented. The present study investigates activities of enzymes of mitochondrial electron transport chain (ETC) namely, NADH:ubiquinone oxidoreductase or complex I, Succinate: ubiquinone oxidoreductase or complex II, Ubiquinol:ferricytochrome c oxidoreductase or complex III, cytochrome c oxidase or complex IV and F 1 F 0 ATPase (ATPase), during Chilo partellus development. Further, the effect of juvenile hormone (JH) analog, methoprene, and brain and corpora-allata-corpora-cardiaca (CC-CA) homogenates that represent neurohormones, on the ETC enzyme activities was monitored. The enzymatic activities increased from penultimate to last larval stage and thereafter declined during pupal development with an exception of ATPase which showed high enzyme activity during last larval and pupal stages compared to the penultimate stage. JH analog, methoprene differentially modulated ETC enzyme activities. It stimulated complex I and IV enzyme activities, but did not alter the activities of complex II, III and ATPase. On the other hand, brain homogenate declined the ATPase activity while the injected CC-CA homogenate stimulated complex I and IV enzyme activities. Cumulatively, the present study is the first to show that mitochondrial ETC enzyme system is under hormone control, particularly of JH and neurohormones during insect development. Copyright © 2015 Elsevier Inc. All rights reserved.

LECITHINASE AND LYSOLECITHINASE ACTIVITY OF RAT INTESTINAL MUCOSA AFTER WHOLE-BODY X-IRRADIATION

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

Ottolenghi, A.; Bernheim, F.

1961-11-01

Twenty-four hours after whole-body x irradiation the lecithinase activity of rat intestinal mucosa has markedly decreased and the lysolecithinase activity has decreasecp to a lesser extent. Addition of normal mucosa or chyi'otrypsin to the irradiated mucosa restores the activity of both enzymes. This indicates that irradiation eithei produces an inhibitor or inactivates a mechanism necessarly to convert pro-enzymes into active enzymes. Since chymo trypsin can increase to some extent the activity of the enzymes in normal mucosa, the second possibility seems more probable. (auth)

Regulation of Hydrolytic Enzyme Activity in Aquatic Microbial Communities Hosted by Carnivorous Pitcher Plants.

PubMed

Young, Erica B; Sielicki, Jessica; Grothjan, Jacob J

2018-04-20

Carnivorous pitcher plants Sarracenia purpurea host diverse eukaryotic and bacterial communities which aid in insect prey digestion, but little is known about the functional processes mediated by the microbial communities. This study aimed to connect pitcher community diversity with functional nutrient transformation processes, identifying bacterial taxa, and measuring regulation of hydrolytic enzyme activity in response to prey and alternative nutrient sources. Genetic analysis identified diverse bacterial taxa known to produce hydrolytic enzyme activities. Chitinase, protease, and phosphatase activities were measured using fluorometric assays. Enzyme activity in field pitchers was positively correlated with bacterial abundance, and activity was suppressed by antibiotics suggesting predominantly bacterial sources of chitinase and protease activity. Fungi, algae, and rotifers observed could also contribute enzyme activity, but fresh insect prey released minimal chitinase activity. Activity of chitinase and proteases was upregulated in response to insect additions, and phosphatase activity was suppressed by phosphate additions. Particulate organic P in prey was broken down, appearing as increasing dissolved organic and inorganic P pools within 14 days. Chitinase and protease were not significantly suppressed by availability of dissolved organic substrates, though organic C and N stimulated bacterial growth, resulting in elevated enzyme activity. This comprehensive field and experimental study show that pitcher plant microbial communities dynamically regulate hydrolytic enzyme activity, to digest prey nutrients to simpler forms, mediating biogeochemical nutrient transformations and release of nutrients for microbial and host plant uptake.

Isolated gene encoding an enzyme with UDP-glucose pyrophosphorylase and phosphoglucomutase activities from Cyclotella cryptica

DOEpatents

Jarvis, Eric E.; Roessler, Paul G.

1999-01-01

The present invention relates to a cloned gene which encodes an enzyme, the purified enzyme, and the applications and products resulting from the use of the gene and enzyme. The gene, isolated from Cyclotella cryptica, encodes a multifunctional enzyme that has both UDP-glucose pyrophosphorylase and phosphoglucomutase activities.

Regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity in murine epidermis. Modulation of enzyme content and activation state by barrier requirements.

PubMed Central

Proksch, E; Elias, P M; Feingold, K R

1990-01-01

Epidermal cholesterol biosynthesis is regulated by barrier function. We quantitated the amount and activation state (phosphorylation-dephosphorylation) of the rate-limiting enzyme, 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase, in epidermis before and after barrier disruption. In murine epidermis we found high enzyme activity (1.75 +/- 0.02 nmol/min per mg protein). After acute barrier disruption, enzyme activity began to increase after 1.5 h, reaching a maximum increase by 2.5 h, and returned to normal by 15 h. Chronic barrier disruption increased total enzyme activity by 83%. In normal epidermis, measurement of HMG CoA reductase activity in microsomes isolated in NaF- vs. NaCl-containing buffers demonstrated that 46 +/- 2% of the enzyme was in the active form. After acute or chronic barrier disruption, a marked increase in the percentage of HMG CoA reductase in the active form was observed. Acute disruption increased enzyme activation state as early as 15 min, reaching a maximum after 2.5 h, with an increase still present at 15 h, indicating that changes in activation state had a close temporal relationship with barrier function. Increases in total HMG CoA reductase activity occurred only after profound barrier disruption, whereas changes in activation state occur with lesser degrees of barrier disruption. Artificial correction of barrier function prevented the increase in total HMG CoA reductase activity, and partially prevented the increase in enzyme activation. These results show that barrier requirements regulate epidermal cholesterol synthesis by modulating both the HMG CoA reductase amount and activation state. Images PMID:2312730

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

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.

Effects of protease and non-starch polysaccharide enzyme on performance, digestive function, activity and gene expression of endogenous enzyme of broilers

PubMed Central

Wang, Mingfa; Zhang, Xiaotu; Wang, Zhixiang

2017-01-01

Three hundred one-day-old male broiler chickens (Ross-308) were fed corn-soybean basal diets containing non-starch polysaccharide (NSP) enzyme and different levels of acid protease from 1 to 42 days of age to investigate the effects of exogenous enzymes on growth performance, digestive function, activity of endogenous digestive enzymes in the pancreas and mRNA expression of pancreatic digestive enzymes. For days 1-42, compared to the control chickens, average daily feed intake (ADFI) and average daily gain (ADG) were significantly enhanced by the addition of NSP enzyme in combination with protease supplementation at 40 or 80 mg/kg (p<0.05). Feed-to-gain ratio (FGR) was significantly improved by supplementation with NSP enzymes or NSP enzyme combined with 40 or 80 mg/kg protease compared to the control diet (p<0.05). Apparent digestibility of crude protein (ADCP) was significantly enhanced by the addition of NSP enzyme or NSP enzyme combined with 40 or 80 mg/kg protease (p<0.05). Cholecystokinin (CCK) level in serum was reduced by 31.39% with NSP enzyme combined with protease supplementation at 160 mg/kg (p<0.05), but the CCK level in serum was increased by 26.51% with NSP enzyme supplementation alone. After 21 days, supplementation with NSP enzyme and NSP enzyme combined with 40 or 80 mg/kg protease increased the activity of pancreatic trypsin by 74.13%, 70.66% and 42.59% (p<0.05), respectively. After 42 days, supplementation with NSP enzyme and NSP enzyme combined with 40 mg/kg protease increased the activity of pancreatic trypsin by 32.45% and 27.41%, respectively (p<0.05). However, supplementation with NSP enzyme and 80 or 160 mg/kg protease decreased the activity of pancreatic trypsin by 10.75% and 25.88%, respectively (p<0.05). The activities of pancreatic lipase and amylase were significantly higher in treated animals than they were in the control group (p<0.05). Supplementation with NSP enzyme, NSP enzyme combined with 40 or 80 mg/kg protease increased pancreatic trypsin mRNA levels by 40%, 44% and 28%, respectively. Supplementation with NSP enzyme and 160 mg/kg protease decreased pancreatic trypsin mRNA levels by 13%. Pancreatic lipase and amylase mRNA expression were significantly elevated in treated animals compared to the control group (p<0.05). These results suggest that the amount of NSP enzyme and acid protease in the diet significantly affects digestive function, endogenous digestive-enzyme activity and mRNA expression in broilers. PMID:28323908

Visualization of enzyme activities inside earthworm biopores by in situ soil zymography

NASA Astrophysics Data System (ADS)

Thu Duyen Hoang, Thi; Razavi, Bahar. S.; Blagodatskaya, Evgenia; Kuzyakov, Yakov

2015-04-01

Earthworms can strongly activate microorganisms, increase microbial and enzyme activities and consequently the turnover of native soil organic matter. In extremely dynamic microhabitats and hotspots as biopores made by earthworms, the in situ enzyme activities are a footprint of complex biotic interactions. The effect of earthworms on the alteration of enzyme activities inside biopores and the difference between bio-pores and earthworm-free soil was visualized by in situ soil zymography (Spohn and Kuzyakov, 2014). For the first time, we prepared quantitative imaging of enzyme activities in biopores. Furthermore, we developed the zymography technique by direct application of a substrate saturated membrane to the soil to obtain better spatial resolution. Lumbricus terrestris L. was placed into transparent box (15×20×15cm). Simultaneously, maize seed was sown in the soil. Control soil box with maize and without earthworm was prepared in the same way. After two weeks when bio-pore systems were formed by earthworm, we visualized in situ enzyme activities of five hydrolytic enzymes (β-glucosidase, cellobiohydrolase, chitinase, xylanase, leucine aminopeptidase) and phosphatase. Followed by non-destructive zymography, biopore samples and control soil were destructively collected to assay enzyme kinetics by fluorogenically labeled substrates method. Zymography showed higher activity of β-glucosidase, chitinase, xylanase and phosphatase in biopores comparing to bulk soil. These differences were further confirmed by fluorimetric microplate enzyme assay detected significant difference of Vmax in four above mentioned enzymes. Vmax of β-glucosidase, chitinase, xylanase and phosphatase in biopores is 68%, 108%, 50% and 49% higher than that of control soil. However, no difference in cellobiohydrolase and leucine aminopeptidase kinetics between biopores and control soil were detected. This indicated little effect of earthworms on protein and cellulose transformation in soil. In conclusion, earthworms contribute to the decomposition of carbohydrates through promoting enzyme activities involved in the C-cycle except for leucine aminopeptidase and cellobiohydrolase. References Spohn M, Kuzyakov Y. (2014) Spatial and temporal dynamics of hotspots of enzyme activity in soil as affected by living and dead roots - a soil zymography analysis, Plant Soil 379: 67-77

Optimization of Enzyme Co-Immobilization with Sodium Alginate and Glutaraldehyde-Activated Chitosan Beads.

PubMed

Gür, Sinem Diken; İdil, Neslihan; Aksöz, Nilüfer

2018-02-01

In this study, two different materials-alginate and glutaraldehyde-activated chitosan beads-were used for the co-immobilization of α-amylase, protease, and pectinase. Firstly, optimization of multienzyme immobilization with Na alginate beads was carried out. Optimum Na alginate and CaCl 2 concentration were found to be 2.5% and 0.1 M, respectively, and optimal enzyme loading ratio was determined as 2:1:0.02 for pectinase, protease, and α-amylase, respectively. Next, the immobilization of multiple enzymes on glutaraldehyde-activated chitosan beads was optimized (3% chitosan concentration, 0.25% glutaraldehyde with 3 h of activation and 3 h of coupling time). While co-immobilization was successfully performed with both materials, the specific activities of enzymes were found to be higher for the enzymes co-immobilized with glutaraldehyde-activated chitosan beads. In this process, glutaraldehyde was acting as a spacer arm. SEM and FTIR were used for the characterization of activated chitosan beads. Moreover, pectinase and α-amylase enzymes immobilized with chitosan beads were also found to have higher activity than their free forms. Three different enzymes were co-immobilized with these two materials for the first time in this study.

Degradation of ureidoglycolate in French bean (Phaseolus vulgaris) is catalysed by a ubiquitous ureidoglycolate urea-lyase.

PubMed

Muñoz, Alfonso; Raso, María José; Pineda, Manuel; Piedras, Pedro

2006-06-01

A ureidoglycolate-degrading activity was analysed in different tissues of French bean (Phaseolus vulgaris L.) plants during development. Activity was detected in all the tissues analysed, although values were very low in seeds before germination and in cotyledons. After radicle emergence, the activity increased due to high activity present in the axes. The highest levels of specific activity were found in developing fruits, from which the enzyme was purified and characterised. This is the first ureidoglycolate-degrading activity that has been purified to homogeneity from a ureide legume. The enzyme was purified 280 fold, and the specific activity for the pure enzyme was 4.4 units mg(-1), which corresponds to a turnover number of 1,055 min(-1). The native enzyme has a molecular mass of 240 kDa and consists of six identical or similar-sized subunits each of 38 kDa. The activity of the purified enzyme was completely dependent on manganese and asparagine. The enzyme exhibited hyperbolic, Michaelian kinetics for ureidoglycolate with a K(m) value of 3.9 mM. This enzyme has been characterised as a ureidoglycolate urea-lyase (EC 4.3.2.3).

Enzyme activity screening of thermophilic bacteria isolated from Dusun Tua Hot Spring, Malaysia

NASA Astrophysics Data System (ADS)

Msarah, Marwan; Ibrahim, Izyanti; Aqma, Wan Syaidatul

2018-04-01

Thermophilic bacteria have biotechnological importance due to the availability of unique enzymes which are stable in extreme circumstances. The aim of this study includes to isolate thermophilic bacteria from hot spring and screen for important enzyme activities. Water samples from the Dusun Tua Hot Spring were collected and the physiochemical characterisation of water was measured. Eight thermophilic bacteria were isolated and determined to have at least three strong enzyme activity including protease, lipase, amylase, cellulase, pectinase and xylanase. The results showed that HuluC2 displayed all the enzyme activities and can be further studied.

The Catalytic Function of Enzymes.

ERIC Educational Resources Information Center

Splittgerber, Allan G.

1985-01-01

Discusses: structure of the enzyme molecule; active site; reaction mechanism; transition state; factors affecting enzyme reaction rates, concentration of enzyme; concentration of substrate; product concentration; temperature effects and pH effects; factors causing a lowering of activation energy; proximity and orientation effects; substrate strain…

Structural prediction and comparative docking studies of psychrophilic β- Galactosidase with lactose, ONPG and PNPG against its counter parts of mesophilic and thermophilic enzymes.

PubMed

Kumar, Ponnada Suresh; Pulicherla, Kk; Ghosh, Mrinmoy; Kumar, Anmol; Rao, Krs Sambasiva

2011-01-01

Enzymes from psychrophiles catalyze the reactions at low temperatures with higher specific activity. Among all the psychrophilic enzymes produced, cold active β-galactosidase from marine psychrophiles revalorizes a new arena in numerous areas at industrial level. The hydrolysis of lactose in to glucose and galactose by cold active β-galactosidase offers a new promising approach in removal of lactose from milk to overcome the problem of lactose intolerance. Herein we propose, a 3D structure of cold active β-galactosidase enzyme sourced from Pseudoalteromonas haloplanktis by using Modeler 9v8 and best model was developed having 88% of favourable region in ramachandran plot. Modelling was followed by docking studies with the help of Auto dock 4.0 against the three substrates lactose, ONPG and PNPG. In addition, comparative docking studies were also performed for the 3D model of psychrophilic β-galactosidase with mesophilic and thermophilic enzymes. Docking studies revealed that binding affinity of enzyme towards the three different substrates is more for psychrophilic enzyme when compared with mesophilic and thermophilic enzymes. It indicates that the enzyme has high specific activity at low temperature when compared with mesophilic and thermophilic enzymes.

Profiling the orphan enzymes

PubMed Central

2014-01-01

The emergence of Next Generation Sequencing generates an incredible amount of sequence and great potential for new enzyme discovery. Despite this huge amount of data and the profusion of bioinformatic methods for function prediction, a large part of known enzyme activities is still lacking an associated protein sequence. These particular activities are called “orphan enzymes”. The present review proposes an update of previous surveys on orphan enzymes by mining the current content of public databases. While the percentage of orphan enzyme activities has decreased from 38% to 22% in ten years, there are still more than 1,000 orphans among the 5,000 entries of the Enzyme Commission (EC) classification. Taking into account all the reactions present in metabolic databases, this proportion dramatically increases to reach nearly 50% of orphans and many of them are not associated to a known pathway. We extended our survey to “local orphan enzymes” that are activities which have no representative sequence in a given clade, but have at least one in organisms belonging to other clades. We observe an important bias in Archaea and find that in general more than 30% of the EC activities have incomplete sequence information in at least one superkingdom. To estimate if candidate proteins for local orphans could be retrieved by homology search, we applied a simple strategy based on the PRIAM software and noticed that candidates may be proposed for an important fraction of local orphan enzymes. Finally, by studying relation between protein domains and catalyzed activities, it appears that newly discovered enzymes are mostly associated with already known enzyme domains. Thus, the exploration of the promiscuity and the multifunctional aspect of known enzyme families may solve part of the orphan enzyme issue. We conclude this review with a presentation of recent initiatives in finding proteins for orphan enzymes and in extending the enzyme world by the discovery of new activities. Reviewers This article was reviewed by Michael Galperin, Daniel Haft and Daniel Kahn. PMID:24906382

Development of in vivo biotransformation enzyme assays for ecotoxicity screening: In vivo measurement of phases I and II enzyme activities in freshwater planarians.

PubMed

Li, Mei-Hui

2016-08-01

The development of a high-throughput tool is required for screening of environmental pollutants and assessing their impacts on aquatic animals. Freshwater planarians can be used in rapid and sensitive toxicity bioassays. Planarians are known for their remarkable regeneration ability but much less known for their metabolic and xenobiotic biotransformation abilities. In this study, the activities of different phase I and II enzymes were determined in vivo by directly measuring fluorescent enzyme substrate disappearance or fluorescent enzyme metabolite production in planarian culture media. For phase I enzyme activity, O-deethylation activities with alkoxyresorufin could not be detected in planarian culture media. By contrast, O-deethylation activities with alkoxycoumarin were detected in planarian culture media. Increases in 7-ethoxycoumarin O-deethylase (ECOD) activities was only observed in planarians exposed to 1μM, but not 10μM, β-naphthoflavone for 24h. ECOD activity was inhibited in planarians exposed to 10 and 100μM rifampicin or carbamazepine for 24h. For phase II enzyme activity, DT-diaphorase, arylsulfatases, uridine 5'-diphospho (UDP)-glucuronosyltransferase or catechol-O-methyltransferase activity was determined in culture media containing planarians. The results of this study indicate that freshwater planarians are a promising model organism to monitor exposure to environmental pollutants or assess their impacts through the in vivo measurement of phase I and II enzyme activities. Copyright © 2016. Published by Elsevier Inc.

Immobilization of an enzyme from a Fusarium fungus WZ-I for chlorpyrifos degradation.

PubMed

Xie, Hui; Zhu, Lusheng; Ma, Tingting; Wang, Jun; Wang, Jinhua; Su, Jun; Shao, Bo

2010-01-01

The free enzyme extracted from WZ-I, which was identified as Fusarium LK. ex Fx, could effectively degrade chlorpyrifos, an organophosphate insecticide. The methods of immobilizing this free enzyme and determined its degradation-related characteristics were investigated. The properties of the immobilized enzyme were compared with those of the free enzyme. The optimal immobilization of the enzyme was achieved in a solution of 30 g/L sodium alginate at 4 degrees C for 4-12 hr. The immobilized enzyme showed the maximal activity at pH 8.0, 45 degrees C. The maximum initial rate and the substrate concentration of the immobilized enzyme were less than that of the free enzyme. The immobilized enzyme, therefore, had a higher capacity to withstand a broader range of temperatures and pH conditions than the free enzyme. With varying pH and temperatures, the immobilized enzyme was more active than the free enzyme in the degradation reaction. In addition, the immobilized enzyme exhibited only a slight loss in its initial activity, even after three repeated uses. The results showed that the immobilized enzyme was more resistant to different environmental conditions, suggesting that it was viable for future practical use.

Lignocellulolytic enzyme production of Pleurotus ostreatus growth in agroindustrial wastes

PubMed Central

da Luz, José Maria Rodrigues; Nunes, Mateus Dias; Paes, Sirlaine Albino; Torres, Denise Pereira; de Cássia Soares da Silva, Marliane; Kasuya, Maria Catarina Megumi

2012-01-01

The mushroom Pleurotus ostreatus has nutritional and medicinal characteristics that depend on the growth substrate. In nature, this fungus grows on dead wood, but it can be artificially cultivated on agricultural wastes (coffee husks, eucalyptus sawdust, corncobs and sugar cane bagasse). The degradation of agricultural wastes involves some enzyme complexes made up of oxidative (laccase, manganese peroxidase and lignin peroxidase) and hydrolytic enzymes (cellulases, xylanases and tanases). Understanding how these enzymes work will help to improve the productivity of mushroom cultures and decrease the potential pollution that can be caused by inadequate discharge of the agroindustrial residues. The objective of this work was to assess the activity of the lignocellulolytic enzymes produced by two P. ostreatus strains (PLO 2 and PLO 6). These strains were used to inoculate samples of coffee husks, eucalyptus sawdust or eucalyptus bark add with or without 20 % rice bran. Every five days after substrate inoculation, the enzyme activity and soluble protein concentration were evaluated. The maximum activity of oxidative enzymes was observed at day 10 after inoculation, and the activity of the hydrolytic enzymes increased during the entire period of the experiment. The results show that substrate composition and colonization time influenced the activity of the lignocellulolytic enzymes. PMID:24031982

Application of carbohydrate arrays coupled with mass spectrometry to detect activity of plant-polysaccharide degradative enzymes from the fungus Aspergillus niger.

PubMed

van Munster, Jolanda M; Thomas, Baptiste; Riese, Michel; Davis, Adrienne L; Gray, Christopher J; Archer, David B; Flitsch, Sabine L

2017-02-21

Renewables-based biotechnology depends on enzymes to degrade plant lignocellulose to simple sugars that are converted to fuels or high-value products. Identification and characterization of such lignocellulose degradative enzymes could be fast-tracked by availability of an enzyme activity measurement method that is fast, label-free, uses minimal resources and allows direct identification of generated products. We developed such a method by applying carbohydrate arrays coupled with MALDI-ToF mass spectrometry to identify reaction products of carbohydrate active enzymes (CAZymes) of the filamentous fungus Aspergillus niger. We describe the production and characterization of plant polysaccharide-derived oligosaccharides and their attachment to hydrophobic self-assembling monolayers on a gold target. We verify effectiveness of this array for detecting exo- and endo-acting glycoside hydrolase activity using commercial enzymes, and demonstrate how this platform is suitable for detection of enzyme activity in relevant biological samples, the culture filtrate of A. niger grown on wheat straw. In conclusion, this versatile method is broadly applicable in screening and characterisation of activity of CAZymes, such as fungal enzymes for plant lignocellulose degradation with relevance to biotechnological applications as biofuel production, the food and animal feed industry.

Function-based classification of carbohydrate-active enzymes by recognition of short, conserved peptide motifs.

PubMed

Busk, Peter Kamp; Lange, Lene

2013-06-01

Functional prediction of carbohydrate-active enzymes is difficult due to low sequence identity. However, similar enzymes often share a few short motifs, e.g., around the active site, even when the overall sequences are very different. To exploit this notion for functional prediction of carbohydrate-active enzymes, we developed a simple algorithm, peptide pattern recognition (PPR), that can divide proteins into groups of sequences that share a set of short conserved sequences. When this method was used on 118 glycoside hydrolase 5 proteins with 9% average pairwise identity and representing four characterized enzymatic functions, 97% of the proteins were sorted into groups correlating with their enzymatic activity. Furthermore, we analyzed 8,138 glycoside hydrolase 13 proteins including 204 experimentally characterized enzymes with 28 different functions. There was a 91% correlation between group and enzyme activity. These results indicate that the function of carbohydrate-active enzymes can be predicted with high precision by finding short, conserved motifs in their sequences. The glycoside hydrolase 61 family is important for fungal biomass conversion, but only a few proteins of this family have been functionally characterized. Interestingly, PPR divided 743 glycoside hydrolase 61 proteins into 16 subfamilies useful for targeted investigation of the function of these proteins and pinpointed three conserved motifs with putative importance for enzyme activity. Furthermore, the conserved sequences were useful for cloning of new, subfamily-specific glycoside hydrolase 61 proteins from 14 fungi. In conclusion, identification of conserved sequence motifs is a new approach to sequence analysis that can predict carbohydrate-active enzyme functions with high precision.

Value of bilirubin oxidase and its mutants in the diagnosis of hyperbilirubinemia.

PubMed

Zhang, Lei; Zhang, Xiao; Luo, Zhi-Ying

2005-11-01

To elucidate the significance of the coordination amino acid residues in bilirubin oxidase (BO) and their kinetic characteristics, and evaluate whether BO mutants may serve as better diagnostic agent for hyperbilirubinemia. The BO mutants I402G and C457S were obtained by site-directed mutagenesis and confirmed by amino acid sequence analysis. Ru-incorporated C457S mutant was obtained by direct incubation of ruthenium compounds with the mutant. The electron paramagnetic resonance (EPR) spectra of the recombinant BO and the mutants were investigated, and the enzyme kinetics of the recombinant BO and I402G mutant were measured with bilirubin as the substrate at 25 degrees C. The BO mutants were expressed and purified successfully. The mutant I402G showed low enzyme activity, and had C457S virtually no enzyme activity. Nevertheless Ru-incorporation conferred higher enzyme activity to C457S mutant. The enzyme kinetic investigations revealed that the kinetic parameter k(cat) of the recombinant BO and I402G mutant was 235.8 min(-1) and 6.9 min(-1), respectively, suggesting higher enzyme activity of the recombinant BO. The coordinating amino acids have important significance in maintaining the integrity of active centers and enzyme activities of recombinant BO and its mutants. The enzyme activities of the mutants I402G and C457S are much lower than those of recombinant BO, therefore they are not appropriate for diagnostic purpose. Ru-incorporation facilitates the formation of a new intact active center in C457S mutant, which therefore acquires enzyme activity.

A meta-analysis of soil exoenzyme responses to simulated climate change

NASA Astrophysics Data System (ADS)

Gebhardt, M.; Espinosa, N. J.; Blankinship, J. C.; Gallery, R. E.

2017-12-01

Microorganisms produce extracellular enzymes to decompose plant matter and drive biogeochemical transformations in soils. Climate change factors, such as warming and altered precipitation patterns, can impact enzyme activity through both direct and indirect mechanisms. Although many individual studies have examined how soil exoenzyme activities respond to climate change manipulations, there is disagreement surrounding the direction of these responses. We performed a synthesis of published studies to examine the influence of warming and altered precipitation on microbial exoenzyme activity. We found that warming increased enzyme activity with a more pronounced effect for oxidative relative to hydrolytic enzymes. Reduced precipitation consistently decreased exoenzyme activity. These responses, however, varied by season, biome, and enzyme type. The majority of studies fitting our criteria (e.g., experiments lasting a minimum of one growing season, paired treatments and controls) were located in North America and Europe. Inferences from this analysis therefore exclude many important ecosystems such as hyper-arid, wetlands, and artic systems. Carbon degrading enzyme activities were less sensitive to climate change manipulations when compared to phosphorus and nitrogen degrading enzyme activities. Linking enzyme activity to biogeochemical processes requires concomitant measurements of organic and inorganic carbon pools, mineralogy, nutrients, microbial biomass and community structure, and heterotrophic respiration within individual studies. Furthermore, linking these parameters to climate and environmental factors will require a comprehensive and consistent inclusion of biotic and abiotic variables among researchers and experiments. Globally, soils contain the largest carbon pools. Understanding the impacts of large-scale perturbations on soil enzyme activity will help to constrain predictions on the fate of biogeochemical transformations and improve model projections.

Novel epoxy activated hydrogels for solving lactose intolerance.

PubMed

Elnashar, Magdy M M; Hassan, Mohamed E

2014-01-01

"Lactose intolerance" is a medical problem for almost 70% of the world population. Milk and dairy products contain 5-10% w/v lactose. Hydrolysis of lactose by immobilized lactase is an industrial solution. In this work, we succeeded to increase the lactase loading capacity to more than 3-fold to 36.3 U/g gel using epoxy activated hydrogels compared to 11 U/g gel using aldehyde activated carrageenan. The hydrogel's mode of interaction was proven by FTIR, DSC, and TGA. The high activity of the epoxy group was regarded to its ability to attach to the enzyme's -SH, -NH, and -OH groups, whereas the aldehyde group could only bind to the enzyme's -NH2 group. The optimum conditions for immobilization such as epoxy chain length and enzyme concentration have been studied. Furthermore, the optimum enzyme conditions were also deliberated and showed better stability for the immobilized enzyme and the Michaelis constants, K m and V max, were doubled. Results revealed also that both free and immobilized enzymes reached their maximum rate of lactose conversion after 2 h, albeit, the aldehyde activated hydrogel could only reach 63% of the free enzyme. In brief, the epoxy activated hydrogels are more efficient in immobilizing more enzymes than the aldehyde activated hydrogel.

An appraisal of the enzyme stability-activity trade-off.

PubMed

Miller, Scott R

2017-07-01

A longstanding idea in evolutionary physiology is that an enzyme cannot jointly optimize performance at both high and low temperatures due to a trade-off between stability and activity. Although a stability-activity trade-off has been observed for well-characterized examples, such a trade-off is not imposed by any physical chemical constraint. To better understand the pervasiveness of this trade-off, I investigated the stability-activity relationship for comparative biochemical studies of purified orthologous enzymes identified by a literature search. The nature of this relationship varied greatly among studies. Notably, studies of enzymes with low mean synonymous nucleotide sequence divergence were less likely to exhibit the predicted negative correlation between stability and activity. Similarly, a survey of directed evolution investigations of the stability-activity relationship indicated that these traits are often uncoupled among nearly identical yet phenotypically divergent enzymes. This suggests that the presumptive trade-off often reported for investigations of enzymes with high mean sequence divergence may in some cases instead be a consequence of the degeneration over time of enzyme function in unselected environments, rather than a direct effect of thermal adaptation. The results caution against the general assertion of a stability-activity trade-off during enzyme adaptation. © 2017 The Author(s). Evolution © 2017 The Society for the Study of Evolution.

Primordial-like enzymes from bacteria with reduced genomes.

PubMed

Ferla, Matteo P; Brewster, Jodi L; Hall, Kelsi R; Evans, Gary B; Patrick, Wayne M

2017-08-01

The first cells probably possessed rudimentary metabolic networks, built using a handful of multifunctional enzymes. The promiscuous activities of modern enzymes are often assumed to be relics of this primordial era; however, by definition these activities are no longer physiological. There are many fewer examples of enzymes using a single active site to catalyze multiple physiologically-relevant reactions. Previously, we characterized the promiscuous alanine racemase (ALR) activity of Escherichia coli cystathionine β-lyase (CBL). Now we have discovered that several bacteria with reduced genomes lack alr, but contain metC (encoding CBL). We characterized the CBL enzymes from three of these: Pelagibacter ubique, the Wolbachia endosymbiont of Drosophila melanogaster (wMel) and Thermotoga maritima. Each is a multifunctional CBL/ALR. However, we also show that CBL activity is no longer required in these bacteria. Instead, the wMel and T. maritima enzymes are physiologically bi-functional alanine/glutamate racemases. They are not highly active, but they are clearly sufficient. Given the abundance of the microorganisms using them, we suggest that much of the planet's biochemistry is carried out by enzymes that are quite different from the highly-active exemplars usually found in textbooks. Instead, primordial-like enzymes may be an essential part of the adaptive strategy associated with streamlining. © 2017 The Authors. Molecular Microbiology Published by John Wiley & Sons Ltd.

Modeling nitrous oxide production and reduction in soil through explicit representation of denitrification enzyme kinetics.

PubMed

Zheng, Jianqiu; Doskey, Paul V

2015-02-17

An enzyme-explicit denitrification model with representations for pre- and de novo synthesized enzymes was developed to improve predictions of nitrous oxide (N2O) accumulations in soil and emissions from the surface. The metabolic model of denitrification is based on dual-substrate utilization and Monod growth kinetics. Enzyme synthesis/activation was incorporated into each sequential reduction step of denitrification to regulate dynamics of the denitrifier population and the active enzyme pool, which controlled the rate function. Parameterizations were developed from observations of the dynamics of N2O production and reduction in soil incubation experiments. The model successfully reproduced the dynamics of N2O and N2 accumulation in the incubations and revealed an important regulatory effect of denitrification enzyme kinetics on the accumulation of denitrification products. Pre-synthesized denitrification enzymes contributed 20, 13, 43, and 62% of N2O that accumulated in 48 h incubations of soil collected from depths of 0-5, 5-10, 10-15, and 15-25 cm, respectively. An enzyme activity function (E) was defined to estimate the relative concentration of active enzymes and variation in response to environmental conditions. The value of E allows for activities of pre-synthesized denitrification enzymes to be differentiated from de novo synthesized enzymes. Incorporating explicit representations of denitrification enzyme kinetics into biogeochemical models is a promising approach for accurately simulating dynamics of the production and reduction of N2O in soils.

Development of Activity-based Cost Functions for Cellulase, Invertase, and Other Enzymes

NASA Astrophysics Data System (ADS)

Stowers, Chris C.; Ferguson, Elizabeth M.; Tanner, Robert D.

As enzyme chemistry plays an increasingly important role in the chemical industry, cost analysis of these enzymes becomes a necessity. In this paper, we examine the aspects that affect the cost of enzymes based upon enzyme activity. The basis for this study stems from a previously developed objective function that quantifies the tradeoffs in enzyme purification via the foam fractionation process (Cherry et al., Braz J Chem Eng 17:233-238, 2000). A generalized cost function is developed from our results that could be used to aid in both industrial and lab scale chemical processing. The generalized cost function shows several nonobvious results that could lead to significant savings. Additionally, the parameters involved in the operation and scaling up of enzyme processing could be optimized to minimize costs. We show that there are typically three regimes in the enzyme cost analysis function: the low activity prelinear region, the moderate activity linear region, and high activity power-law region. The overall form of the cost analysis function appears to robustly fit the power law form.

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

Isolated gene encoding an enzyme with UDP-glucose pyrophosphorylase and phosphoglucomutase activities from Cyclotella cryptica

DOEpatents

Jarvis, E.E.; Roessler, P.G.

1999-07-27

The present invention relates to a cloned gene which encodes an enzyme, the purified enzyme, and the applications and products resulting from the use of the gene and enzyme. The gene, isolated from Cyclotella cryptica, encodes a multifunctional enzyme that has both UDP-glucose pyrophosphorylase and phosphoglucomutase activities. 8 figs.

Ectomycorrhizal Fungal Communities and Enzymatic Activities Vary across an Ecotone between a Forest and Field

PubMed Central

Rúa, Megan A.; Moore, Becky; Hergott, Nicole; Van, Lily; Jackson, Colin R.; Hoeksema, Jason D.

2015-01-01

Extracellular enzymes degrade macromolecules into soluble substrates and are important for nutrient cycling in soils, where microorganisms, such as ectomycorrhizal (ECM) fungi, produce these enzymes to obtain nutrients. Ecotones between forests and fields represent intriguing arenas for examining the effect of the environment on ECM community structure and enzyme activity because tree maturity, ECM composition, and environmental variables may all be changing simultaneously. We studied the composition and enzymatic activity of ECM associated with loblolly pine (Pinus taeda) across an ecotone between a forest where P. taeda is established and an old field where P. taeda saplings had been growing for <5 years. ECM community and environmental characteristics influenced enzyme activity in the field, indicating that controls on enzyme activity may be intricately linked to the ECM community, but this was not true in the forest. Members of the Russulaceae were associated with increased phenol oxidase activity and decreased peroxidase activity in the field. Members of the Atheliaceae were particularly susceptible to changes in their abiotic environment, but this did not mediate differences in enzyme activity. These results emphasize the complex nature of factors that dictate the distribution of ECM and activity of their enzymes across a habitat boundary. PMID:29376908

Redox Switch for the Inhibited State of Yeast Glycogen Synthase Mimics Regulation by Phosphorylation

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

Mahalingan, Krishna K.; Baskaran, Sulochanadevi; DePaoli-Roach, Anna A.

Glycogen synthase (GS) is the rate limiting enzyme in the synthesis of glycogen. Eukaryotic GS is negatively regulated by covalent phosphorylation and allosterically activated by glucose-6-phosphate (G-6-P). To gain structural insights into the inhibited state of the enzyme, we solved the crystal structure of yGsy2-R589A/R592A to a resolution of 3.3 Å. The double mutant has an activity ratio similar to the phosphorylated enzyme and also retains the ability to be activated by G-6-P. When compared to the 2.88 Å structure of the wild-type G-6-P activated enzyme, the crystal structure of the low-activity mutant showed that the N-terminal domain of themore » inhibited state is tightly held against the dimer-related interface thereby hindering acceptor access to the catalytic cleft. On the basis of these two structural observations, we developed a reversible redox regulatory feature in yeast GS by substituting cysteine residues for two highly conserved arginine residues. When oxidized, the cysteine mutant enzyme exhibits activity levels similar to the phosphorylated enzyme but cannot be activated by G-6-P. Upon reduction, the cysteine mutant enzyme regains normal activity levels and regulatory response to G-6-P activation.« less

Dramatic enhancement of enzymatic activity in organic solvents by lyoprotectants

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

Dabulis, K.; Klibanov, A.M.

1993-03-05

When seven different hydrolytic enzymes (four proteases and three lipases) were lyophilized from aqueous solution containing a ligand, N-Ac-L-Phe-NH[sub 2], their catalytic activity in anhydrous solvents was far greater (one to two orders of magnitude) than that of the enzymes lyophilized without the ligand. This ligand-induced activation was expressed regardless of whether the substrate employed in organic solvents structurally resembled the ligand. Furthermore, nonligand lyoprotectants [sorbitol, other sugars, and poly(ethylene glycol)] also dramatically enhanced enzymatic activity in anhydrous solvents when present in enzyme aqueous solution prior to lyophilization. The effects of the ligand and of the lyoprotectants were nonadditive, suggestingmore » the same mechanism of action. Excipient-activated and nonactivated enzymes exhibited identical activities in water. Also, addition of the excipients directly to suspensions of nonactivated enzymes in organic solvents had no appreciable effect on catalytic activity. These observations indicate that the mechanism of the excipient-induced activation is based on the ability of the excipients to alleviate reversible denaturation of enzymes upon lyophilization. Activity enhancement induced by the excipients is displayed even after their removal by washing enzymes with anhydrous solvents. Subtilisin Carlsberg, lyophilized with sorbitol, was found to be a much more efficient practical catalyst than its regular' counterpart.« less

Sphingolipid Long-Chain Base Synthesis in Plants (Characterization of Serine Palmitoyltransferase Activity in Squash Fruit Microsomes).

PubMed

Lynch, D. V.; Fairfield, S. R.

1993-12-01

The activity of serine palmitoyltransferase (palmitoyl-coenzyme A [CoA]:L-serine [Ser]-C-palmitoyltransferase [decarboxylating], EC 2.3.1.50), the enzyme catalyzing the first step in the synthesis of the long-chain base required for sphingolipid assembly, has been characterized in a plant system. Enzyme activity in a microsomal membrane fraction from summer squash fruit (Cucurbita pepo L. cv Early Prolific Straightneck) was assayed by monitoring the incorporation of L-[3H]Ser into the chloroform-soluble product, 3-ketosphinganine. Addition of NADPH to the assay system resulted in the conversion of 3-ketosphinganine to sphinganine. The apparent Km for Ser was approximately 1.8 mM. The enzyme exhibited a strong preference for palmitoyl-CoA, with optimal activity at a substrate concentration of 200 [mu]M. Pyridoxal 5[prime]-phosphate was required as a coenzyme. The pH optimum was 7.6, and the temperature optimum was 36 to 40[deg]C. Enzyme activity was greatest in the microsomal fraction obtained by differential centrifugation and was localized to the endoplasmic reticulum using marker enzymes. Two known mechanism-based inhibitors of the mammalian enzyme, L-cycloserine and [beta]-chloro-L-alanine, were effective inhibitors of enzyme activity in squash microsomes. Changes in enzyme activity with size (age) of squash fruit were observed. The results from this study suggest that the properties and catalytic mechanism of Ser palmitoyltransferase from squash are similar to those of the animal, fungal, and bacterial enzyme in most respects. The specific activity of the enzyme in squash microsomes ranged from 0.57 to 0.84 nmol min-1 mg-1 of protein, values 2- to 20-fold higher than those previously reported for preparations from animal tissues.

Life-history evolution and the microevolution of intermediary metabolism: activities of lipid-metabolizing enzymes in life-history morphs of a wing-dimorphic cricket.

PubMed

Zera, Anthony J; Zhao, Zhangwu

2003-03-01

Although a considerable amount of information is available on the ecology, genetics, and physiology of life-history traits, much more limited data are available on the biochemical and genetic correlates of life-history variation within species. Specific activities of five enzymes of lipid biosynthesis and two enzymes of amino acid catabolism were compared among lines selected for flight-capable (LW[f]) versus flightless (SW) morphs of the cricket Gryllus firmus. These morphs, which exist in natural populations, differ genetically in ovarian growth (100-400% higher in SW) and aspects of flight capability including the size of wings and flight muscles, and the concentration of triglyceride flight fuel (40% greater in LW[f]). Consistently higher activity of each enzyme in LW(f) versus SW-selected lines, and strong co-segregation between morph and enzyme activity, demonstrated genetically based co-variance between wing morph and enzyme activity. Developmental profiles of enzyme activities strongly paralleled profiles of triglyceride accumulation during adulthood and previous measures of in vivo lipid biosynthesis. These data strongly imply that genetically based elevation in activities of lipogenic enzymes, and enzymes controlling the conversion of amino acids into lipids, is an important cause underlying the elevated accumulation of triglyceride in the LW(f) morph, a key biochemical component of the trade-off between elevated early fecundity and flight capability. Global changes in lipid and amino-acid metabolism appear to have resulted from microevolutionary alteration of regulators of metabolism. Finally, strong genotype x environment (diet) interactions were observed for most enzyme activities. Future progress in understanding the functional causes of life-history evolution requires a more detailed synthesis of the fields of life-history evolution and metabolic biochemistry. Wing polymorphism is a powerful experimental model in such integrative studies.

Variation in Its C-Terminal Amino Acids Determines Whether Endo-β-Mannanase Is Active or Inactive in Ripening Tomato Fruits of Different Cultivars1

PubMed Central

Bourgault, Richard; Bewley, J. Derek

2002-01-01

Endo-β-mannanase cDNAs were cloned and characterized from ripening tomato (Lycopersicon esculentum Mill. cv Trust) fruit, which produces an active enzyme, and from the tomato cv Walter, which produces an inactive enzyme. There is a two-nucleotide deletion in the gene from tomato cv Walter, which results in a frame shift and the deletion of four amino acids at the C terminus of the full-length protein. Other cultivars that produce either active or inactive enzyme show the same absence or presence of the two-nucleotide deletion. The endo-β-mannanase enzyme protein was purified and characterized from ripe fruit to ensure that cDNA codes for the enzyme from fruit. Immunoblot analysis demonstrated that non-ripening mutants, which also fail to exhibit endo-β-mannanase activity, do so because they fail to express the protein. In a two-way genetic cross between tomato cvs Walter and Trust, all F1 progeny from both crosses produced fruit with active enzyme, suggesting that this form is dominant and homozygous in tomato cv Trust. Self-pollination of a plant from the heterozygous F1 generation yielded F2 plants that bear fruit with and without active enzyme at a ratio appropriate to Mendelian genetic segregation of alleles. Heterologous expression of the two endo-β-mannanase genes in Escherichia coli resulted in active enzyme being produced from cultures containing the tomato cv Trust gene and inactive enzyme being produced from those containing the tomato cv Walter gene. Site-directed mutagenesis was used to establish key elements in the C terminus of the endo-β-mannanase protein that are essential for full enzyme activity. PMID:12427992

Enzymes immobilized in mesoporous silica: a physical-chemical perspective.

PubMed

Carlsson, Nils; Gustafsson, Hanna; Thörn, Christian; Olsson, Lisbeth; Holmberg, Krister; Åkerman, Björn

2014-03-01

Mesoporous materials as support for immobilized enzymes have been explored extensively during the last two decades, primarily not only for biocatalysis applications, but also for biosensing, biofuels and enzyme-controlled drug delivery. The activity of the immobilized enzymes inside the pores is often different compared to that of the free enzymes, and an important challenge is to understand how the immobilization affects the enzymes in order to design immobilization conditions that lead to optimal enzyme activity. This review summarizes methods that can be used to understand how material properties can be linked to changes in enzyme activity. Real-time monitoring of the immobilization process and techniques that demonstrate that the enzymes are located inside the pores is discussed by contrasting them to the common practice of indirectly measuring the depletion of the protein concentration or enzyme activity in the surrounding bulk phase. We propose that pore filling (pore volume fraction occupied by proteins) is the best standard for comparing the amount of immobilized enzymes at the molecular level, and present equations to calculate pore filling from the more commonly reported immobilized mass. Methods to detect changes in enzyme structure upon immobilization and to study the microenvironment inside the pores are discussed in detail. Combining the knowledge generated from these methodologies should aid in rationally designing biocatalyst based on enzymes immobilized in mesoporous materials. © 2013 Elsevier B.V. All rights reserved.

Designing Allosteric Control into Enzymes by Chemical Rescue of Structure

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

Deckert, Katelyn; Budiardjo, S. Jimmy; Brunner, Luke C.

2012-08-07

Ligand-dependent activity has been engineered into enzymes for purposes ranging from controlling cell morphology to reprogramming cellular signaling pathways. Where these successes have typically fused a naturally allosteric domain to the enzyme of interest, here we instead demonstrate an approach for designing a de novo allosteric effector site directly into the catalytic domain of an enzyme. This approach is distinct from traditional chemical rescue of enzymes in that it relies on disruption and restoration of structure, rather than active site chemistry, as a means to achieve modulate function. We present two examples, W33G in a {beta}-glycosidase enzyme ({beta}-gly) and W492Gmore » in a {beta}-glucuronidase enzyme ({beta}-gluc), in which we engineer indole-dependent activity into enzymes by removing a buried tryptophan side chain that serves as a buttress for the active site architecture. In both cases, we observe a loss of function, and in both cases we find that the subsequent addition of indole can be used to restore activity. Through a detailed analysis of {beta}-gly W33G kinetics, we demonstrate that this rescued enzyme is fully functionally equivalent to the corresponding wild-type enzyme. We then present the apo and indole-bound crystal structures of {beta}-gly W33G, which together establish the structural basis for enzyme inactivation and rescue. Finally, we use this designed switch to modulate {beta}-glycosidase activity in living cells using indole. Disruption and recovery of protein structure may represent a general technique for introducing allosteric control into enzymes, and thus may serve as a starting point for building a variety of bioswitches and sensors.« less

Immobilization of Chloroperoxidase on Aminopropyl-Glass

PubMed Central

Kadima, Tenshuk A.; Pickard, Michael A.

1990-01-01

Chloroperoxidase (CPO) purified from Caldariomyces fumago CMI 89362 was covalently bound to aminopropyl-glass by using a modification of an established method. Acid-washed glass was derivatized by using aminopropyltriethoxysilane, and the enzyme was ionically bound at low ionic strength. Further treatment with glutaraldehyde covalently linked the enzyme to the glass beads in an active form. No elution of bound activity from glass beads could be detected with a variety of washings. The loading of enzyme protein to the glass beads was highest, 100 mg of CPO per g of glass, at high reaction ratios of CPO to glass, but the specific activity of the immobilized enzyme was highest, 36% of theoretical, at low enzyme-to-carrier ratios. No differences in the properties of the soluble and immobilized enzymes could be detected by a number of criteria: their pH-activity and pH-stability profiles were similar, as were their thermal stabilities. After five uses, the immobilized enzyme retained full activity between pH 6.0 and 6.7. PMID:16348352

Enzyme Prodrug Therapy Engineered into Electrospun Fibers with Embedded Liposomes for Controlled, Localized Synthesis of Therapeutics.

PubMed

Chandrawati, Rona; Olesen, Morten T J; Marini, Thatiane C C; Bisra, Gurpal; Guex, Anne Géraldine; de Oliveira, Marcelo G; Zelikin, Alexander N; Stevens, Molly M

2017-09-01

Enzyme prodrug therapy (EPT) enables localized conversion of inert prodrugs to active drugs by enzymes. Performance of EPT necessitates that the enzyme remains active throughout the time frame of the envisioned therapeutic application. β-glucuronidase is an enzyme with historically validated performance in EPT, however it retains its activity in biomaterials for an insufficiently long period of time, typically not exceeding 7 d. Herein, the encapsulation of β-glucuronidase in liposomal subcompartments within poly(vinyl alcohol) electrospun fibers is reported, leading to the assembly of biocatalytically active materials with activity of the enzyme sustained over at least seven weeks. It is further shown that liposomes provide the highly beneficial stabilization of the enzyme when incubated in cell culture media. The assembled biocatalytic materials successfully produce antiproliferative drugs (SN-38) using externally administered prodrugs (SN-38-glucuronide) and effectively suppress cell proliferation, with envisioned utility in the design of cardiovascular grafts. © 2017 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Loss in photosynthesis during senescence is accompanied by an increase in the activity of β-galactosidase in leaves of Arabidopsis thaliana: modulation of the enzyme activity by water stress.

PubMed

Pandey, Jitendra Kumar; Dash, Sidhartha Kumar; Biswal, Basanti

2017-07-01

The precise nature of the developmental modulation of the activity of cell wall hydrolases that breakdown the wall polysaccharides to maintain cellular sugar homeostasis under sugar starvation environment still remains unclear. In this work, the activity of β-galactosidase (EC 3.2.1.23), a cell-wall-bound enzyme known to degrade the wall polysaccharides, has been demonstrated to remarkably enhance during senescence-induced loss in photosynthesis in Arabidopsis thaliana. The enhancement in the enzyme activity reaches a peak at the terminal phase of senescence when the rate of photosynthesis is at its minimum. Although the precise nature of chemistry of the interface between the decline in photosynthesis and enhancement in the activity of the enzyme could not be fully resolved, the enhancement in its activity in dark and its suppression in light or with exogenous sugars may indicate the involvement of loss of photosynthetic production of sugars as a key factor that initiates and stimulates the activity of the enzyme. The hydrolase possibly participates in the catabolic network of cell wall polysaccharides to produce sugars for execution of energy-dependant senescence program in the background of loss of photosynthesis. Drought stress experienced by the senescing leaves accelerates the decline in photosynthesis with further stimulation in the activity of the enzyme. The stress recovery of photosynthesis and suppression of the enzyme activity on withdrawal of stress support the proposition of photosynthetic modulation of the cell-wall-bound enzyme activity.

Purification and characterization of endo-beta-1,4 mannanase from Aspergillus niger gr for application in food processing industry.

PubMed

Naganagouda, K; Salimath, P V; Mulimani, V H

2009-10-01

A thermostable extracellular beta-mannanase from the culture supernatant of a fungus Aspergillus niger gr was purified to homogeneity. SDS-PAGE of the purified enzyme showed a single protein band of molecular mass 66 kDa. The beta- mannanase exhibited optimum catalytic activity at pH 5.5 and 55 degrees C. It was thermostable at 55 degrees C, and retained 50% activity after 6 h at 55 degrees C. The enzyme was stable at a pH range of 3.0 to 7.0. The metal ions Hg(2+), Cu(2+), and Ag(2+) inhibited complete enzyme activity. The inhibitors tested, EDTA, PMSF, and 1,10-phenanthroline, did not inhibit the enzyme activity. N-Bromosuccinimide completely inhibited enzyme activity. The relative substrate specificity of enzyme towards the various mannans is in the order of locust bean gum>guar gum>copra mannan, with K(m) of 0.11, 0.28, and 0.33 mg/ml, respectively. Since the enzyme is active over a wide range of pH and temperature, it could find potential use in the food-processing industry.

Remarkable Reproducibility of Enzyme Activity Profiles in Tomato Fruits Grown under Contrasting Environments Provides a Roadmap for Studies of Fruit Metabolism1[W][OPEN

PubMed Central

Biais, Benoît; Bénard, Camille; Beauvoit, Bertrand; Colombié, Sophie; Prodhomme, Duyên; Ménard, Guillaume; Bernillon, Stéphane; Gehl, Bernadette; Gautier, Hélène; Ballias, Patricia; Mazat, Jean-Pierre; Sweetlove, Lee; Génard, Michel; Gibon, Yves

2014-01-01

To assess the influence of the environment on fruit metabolism, tomato (Solanum lycopersicum ‘Moneymaker’) plants were grown under contrasting conditions (optimal for commercial, water limited, or shaded production) and locations. Samples were harvested at nine stages of development, and 36 enzyme activities of central metabolism were measured as well as protein, starch, and major metabolites, such as hexoses, sucrose, organic acids, and amino acids. The most remarkable result was the high reproducibility of enzyme activities throughout development, irrespective of conditions or location. Hierarchical clustering of enzyme activities also revealed tight relationships between metabolic pathways and phases of development. Thus, cell division was characterized by high activities of fructokinase, glucokinase, pyruvate kinase, and tricarboxylic acid cycle enzymes, indicating ATP production as a priority, whereas cell expansion was characterized by enzymes involved in the lower part of glycolysis, suggesting a metabolic reprogramming to anaplerosis. As expected, enzymes involved in the accumulation of sugars, citrate, and glutamate were strongly increased during ripening. However, a group of enzymes involved in ATP production, which is probably fueled by starch degradation, was also increased. Metabolites levels seemed more sensitive than enzymes to the environment, although such differences tended to decrease at ripening. The integration of enzyme and metabolite data obtained under contrasting growth conditions using principal component analysis suggests that, with the exceptions of alanine amino transferase and glutamate and malate dehydrogenase and malate, there are no links between single enzyme activities and metabolite time courses or levels. PMID:24474652

Simultaneously and separately immobilizing incompatible dual-enzymes on polymer substrate via visible light induced graft polymerization

NASA Astrophysics Data System (ADS)

Zhu, Xing; He, Bin; Zhao, Changwen; Ma, Yuhong; Yang, Wantai

2018-04-01

Developing facile and mild strategy to construct multi-enzymes immobilization system has attracted considerable attentions in recent years. Here a simple immobilization strategy called visible light induced graft polymerization that can simultaneously and separately encapsulate two kinds of enzymes on one polymer film was proposed. Two incompatible enzymes, trypsin and transglutaminase (TGase) were selected as model dual-enzymes system and simultaneously immobilized on two sides of low-density polyethylene (LDPE) film. After immobilization, it was found that more than 90% of the enzymes can be embedded into dual-enzymes loaded film without leakage. And the activities of both separately immobilized enzymes were higher than the activities of mixed co-immobilized enzymes or the sequential immobilized ones. This dual-enzymes loaded film (DEL film) showed excellent recyclability and can retain >87% activities of both enzymes after 4 cycles of utilization. As an example, this DEL film was used to conjugate a prodrug of cytarabine with a target peptide. The successful preparation of expected product demonstrated that the separately immobilized two enzymes can worked well together to catalyze a two-step reaction.

Kinetic study of Candida antarctica lipase B immobilization using poly(methyl methacrylate) nanoparticles obtained by miniemulsion polymerization as support.

PubMed

Valério, Alexsandra; Nicoletti, Gabrieli; Cipolatti, Eliane P; Ninow, Jorge L; Araújo, Pedro H H; Sayer, Cláudia; de Oliveira, Débora

2015-03-01

With the objective to obtain immobilized Candida antarctica lipase B (CalB) with good activity and improved utilization rate, this study evaluated the influence of enzyme and crodamol concentrations and initiator type on the CalB enzyme immobilization in nanoparticles consisting of poly(methyl methacrylate) (PMMA) obtained by miniemulsion polymerization. The kinetic study of immobilized CalB enzyme in PMMA nanoparticles was evaluated in terms of monomer conversion, particle size, zeta potential, and relative activity. The optimum immobilization condition for CalB was compared with free enzyme in the p-NPL hydrolysis activity measurement. Results showed a higher CalB enzyme stability after 20 hydrolysis cycles compared with free CalB enzyme; in particular, the relative immobilized enzyme activity was maintained up to 40%. In conclusion, PMMA nanoparticles proved to be a good support for the CalB enzyme immobilization and may be used as a feasible alternative catalyst in industrial processes.

Function and biotechnology of extremophilic enzymes in low water activity

PubMed Central

2012-01-01

Enzymes from extremophilic microorganisms usually catalyze chemical reactions in non-standard conditions. Such conditions promote aggregation, precipitation, and denaturation, reducing the activity of most non-extremophilic enzymes, frequently due to the absence of sufficient hydration. Some extremophilic enzymes maintain a tight hydration shell and remain active in solution even when liquid water is limiting, e.g. in the presence of high ionic concentrations, or at cold temperature when water is close to the freezing point. Extremophilic enzymes are able to compete for hydration via alterations especially to their surface through greater surface charges and increased molecular motion. These properties have enabled some extremophilic enzymes to function in the presence of non-aqueous organic solvents, with potential for design of useful catalysts. In this review, we summarize the current state of knowledge of extremophilic enzymes functioning in high salinity and cold temperatures, focusing on their strategy for function at low water activity. We discuss how the understanding of extremophilic enzyme function is leading to the design of a new generation of enzyme catalysts and their applications to biotechnology. PMID:22480329

Neofunctionalization of a duplicate hatching enzyme gene during the evolution of teleost fishes.

PubMed

Sano, Kaori; Kawaguchi, Mari; Watanabe, Satoshi; Yasumasu, Shigeki

2014-10-19

Duplication and subsequent neofunctionalization of the teleostean hatching enzyme gene occurred in the common ancestor of Euteleostei and Otocephala, producing two genes belonging to different phylogenetic clades (clade I and II). In euteleosts, the clade I enzyme inherited the activity of the ancestral enzyme of swelling the egg envelope by cleavage of the N-terminal region of egg envelope proteins. The clade II enzyme gained two specific cleavage sites, N-ZPd and mid-ZPd but lost the ancestral activity. Thus, euteleostean clade II enzymes assumed a new function; solubilization of the egg envelope by the cooperative action with clade I enzyme. However, in Otocephala, the clade II gene was lost during evolution. Consequently, in a late group of Otocephala, only the clade I enzyme is present to swell the egg envelope. We evaluated the egg envelope digestion properties of clade I and II enzymes in Gonorynchiformes, an early diverging group of Otocephala, using milkfish, and compared their digestion with those of other fishes. Finally, we propose a hypothesis of the neofunctionalization process. The milkfish clade II enzyme cleaved N-ZPd but not mid-ZPd, and did not cause solubilization of the egg envelope. We conclude that neofunctionalization is incomplete in the otocephalan clade II enzymes. Comparison of clade I and clade II enzyme characteristics implies that the specificity of the clade II enzymes gradually changed during evolution after the duplication event, and that a change in substrate was required for the addition of the mid-ZPd site and loss of activity at the N-terminal region. We infer the process of neofunctionalization of the clade II enzyme after duplication of the gene. The ancestral clade II gene gained N-ZPd cleavage activity in the common ancestral lineage of the Euteleostei and Otocephala. Subsequently, acquisition of cleavage activity at the mid-ZPd site and loss of cleavage activity in the N-terminal region occurred during the evolution of Euteleostei, but not of Otocephala. The clade II enzyme provides an example of the development of a neofunctional gene for which the substrate, the egg envelope protein, has adapted to a gradual change in the specificity of the corresponding enzyme.

[Hepatic allopurinol oxidizing enzyme in mice].

PubMed

Huh, K; Iwata, H; Yamamoto, I

1975-03-01

The relationship between allopurinol oxidizing enzyme and aldehyde oxidase was investaged in mice. The oxidation of both N-methylnicotinamide and allopurinol appears to be catalized by a single enzyme, aldehyde oxidase (aldehyde-oxygen oxidoreductase EC, 1.2.3.1.). This conclusion is based on the following evidence; The postnatal changes of allopurinol and N-methylnicotinamide oxidizing activities were similar during growth and the levels of both activities increased in a parallel fashion upon the attainment of sexual maturity. The rates of loss of the activities of both enzymes by heat denaturation as well as dexamethasone administration were similar. The inhibitors of allopurinol oxidizing enzyme also suppressed N-methylnicotinamide oxidation. Competition of N-methylnicotineamide and allopurinol for oxidation was demonstrated. The rate of increase of the activities in both enzymes was almost parallel during each step of the purification from mouse liver supernatant. It was ascertained that xanthine oxidase in the enzyme preparation does not influence allopurinol oxidation.

Kinase Activity Studied in Living Cells Using an Immunoassay

ERIC Educational Resources Information Center

Bavec, Aljos?a

2014-01-01

This laboratory exercise demonstrates the use of an immunoassay for studying kinase enzyme activity in living cells. The advantage over the classical method, in which students have to isolate the enzyme from cell material and measure its activity in vitro, is that enzyme activity is modulated and measured in living cells, providing a more…

Effects of dietary lead acetate on hepatic detoxication enzyme activity

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

Wagstaff, D.J.

1979-12-01

Lead-containing compounds usually inhibit enzymic and metabolic processes. This inhibition is presumed to be the mechanism of intoxication by these compounds. Inhibition of detoxication activities of liver microsomal enzymes could be particularly detrimental because the toxicity of many different substances would be increased. Exposure of experimental animals to lead compounds in several studies has been associated with depressed activity of hepatic microsomal enzymes, reduced levels of hepatic cytochrome P-450, reduced levels of hepatic microsomal protein, and prolonged hexobarbital sleep times. The present report contains observations that under certain experimental conditions there is stimulated hepatic meicrosomal enzyme activity in rats fedmore » lead acetate.« less

The kinetics of the oxidation of cytochrome c by Paracoccus cytochrome c peroxidase.

PubMed

Gilmour, R; Goodhew, C F; Pettigrew, G W; Prazeres, S; Moura, J J; Moura, I

1994-06-15

In work that is complementary to our investigation of the spectroscopic features of the cytochrome c peroxidase from Paracoccus denitrificans [Gilmour, Goodhew, Pettigrew, Prazeres, Moura and Moura (1993) Biochem. J. 294, 745-752], we have studied the kinetics of oxidation of cytochrome c by this enzyme. The enzyme, as isolated, is in the fully oxidized form and is relatively inactive. Reduction of the high-potential haem at pH 6 with ascorbate results in partial activation of the enzyme. Full activation is achieved by addition of 1 mM CaCl2. Enzyme activation is associated with formation of a high-spin state at the oxidized low-potential haem. EGTA treatment of the oxidized enzyme prevents activation after reduction with ascorbate, while treatment with EGTA of the reduced, partially activated, form abolishes the activity. We conclude that the active enzyme is a mixed-valence form with the low-potential haem in a high-spin state that is stabilized by Ca2+. Dilution of the enzyme results in a progressive loss of activity, the extent of which depends on the degree of dilution. Most of the activity lost upon dilution can be recovered after reconcentration. The M(r) of the enzyme on molecular-exclusion chromatography is concentration-dependent, with a shift to lower values at lower concentrations. Values of M(r) obtained are intermediate between those of a monomer (39,565) and a dimer. We propose that the active form of the enzyme is a dimer which dissociates at high dilution to give inactive monomers. From the activity of the enzyme at different dilutions, a KD of 0.8 microM can be calculated for the monomerdimer equilibrium. The cytochrome c peroxidase oxidizes horse ferrocytochrome c with first-order kinetics, even at high ferrocytochrome c concentrations. The maximal catalytic-centre activity ('turnover number') under the assay conditions used is 62,000 min-1, with a half-saturating ferrocytochrome c concentration of 3.3 microM. The corresponding values for the Paracoccus cytochrome c-550 (presumed to be the physiological substrate) are 85,000 min-1 and 13 microM. However, in this case, the kinetics deviate from first-order progress curves at all ferrocytochrome c concentrations. Consideration of the periplasmic environment in Paracoccus denitrificans leads us to propose that the enzyme will be present as the fully active dimer supplied with saturating ferrocytochrome c-550.

Effect of vitamin A deprivation on the cholesterol side-chain cleavage enzyme activity of testes and ovaries of rats (Short Communication)

PubMed Central

Jayaram, M.; Murthy, S. K.; Ganguly, J.

1973-01-01

The cholesterol side-chain cleavage enzyme activity is decreased considerably at the mild stage of vitamin A deficiency in rat testes and ovaries and the decrease in activity becomes more pronounced with progress of deficiency. Supplementation of the deficient rats with retinyl acetate, but not retinoic acid, restores the enzyme activity to normal values. The cholesterol side-chain cleavage enzyme of adrenals is not affected by any of the above treatments. PMID:4772624

Accuracy of tissue Doppler echocardiography in the diagnosis of new-onset congestive heart failure in patients with levels of B-type natriuretic peptide in the midrange and normal left ventricular ejection fraction.

PubMed

Arques, Stephane; Roux, Emmanuel; Sbragia, Pascal; Pieri, Bertrand; Gelisse, Richard; Ambrosi, Pierre; Luccioni, Roger

2006-09-01

Based on the hypothesis that it reflects left ventricular (LV) diastolic pressures, B-type natriuretic peptide (BNP) is largely utilized as first-line diagnostic complement in the emergency diagnosis of congestive heart failure (HF). The incremental diagnostic value of tissue Doppler echocardiography, a reliable noninvasive estimate of LV filling pressures, has been reported in patients with preserved LV ejection fraction and discrepancy between BNP levels and the clinical judgment, however, its clinical validity in such patients in the presence of BNP concentrations in the midrange, which may reflect intermediate, nondiagnostic levels of LV filling pressures, is unknown. 34 patients without history of HF, presenting with acute dyspnea at rest, BNP levels of 100-400 pg/ml and normal LV ejection fraction were prospectively enrolled (17 with congestive HF and 17 with noncardiac cause). Tissue Doppler echocardiography was performed within 3 hours after admission. Unlike BNP (P = 0.78), Boston criteria (P = 0.0129), radiographic pulmonary edema (P = 0.0036) and average E/Ea ratio (P = 0.0032) were predictive of congestive HF by logistic regression analysis. In this clinical setting, radiographic pulmonary edema had a positive predictive value of 80% in the diagnosis of congestive HF. In patients without evidence of radiographic pulmonary edema, average E/Ea > 10 was a powerful predictor of congestive HF (area under the ROC curve of 0.886, P < 0.001, sensitivity 100% and specificity 78.6%). By better reflecting LV filling pressures, bedside tissue Doppler echocardiography accurately differentiates congestive HF from noncardiac cause in dyspneic patients with intermediate, nondiagnostic BNP levels and normal LV ejection fraction.

Rabies virus co-localizes with early (Rab5) and late (Rab7) endosomal proteins in neuronal and SH-SY5Y cells.

PubMed

Ahmad, Waqas; Li, Yingying; Guo, Yidi; Wang, Xinyu; Duan, Ming; Guan, Zhenhong; Liu, Zengshan; Zhang, Maolin

2017-06-01

Rabies virus (RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and pH-dependent pathway for trafficking and invasion into endothelial cells. Early (Rab5, EEA1) and late (Rab7, LAMP1) endosomal proteins play critical roles in endosomal sorting, maturity and targeting various molecular cargoes, but their precise functions in the early stage of RABV neuronal infection remain elusive. In this study, the relationship between enigmatic entry of RABV with these endosomal proteins into neuronal and SH-SY5Y cells was investigated. Immunofluorescence, TCID 50 titers, electron microscopy and western blotting were carried out to determine the molecular interaction of the nucleoprotein (N) of RABV with early or late endosomal proteins in these cell lines. The expression of N was also determined by down-regulating Rab5 and Rab7 in both cell lines through RNA interference. The results were indicative that N proficiently colocalized with Rab5/EEA1 and Rab7/LAMP1 in both cell lines at 24 and 48 h post-infection, while N titers significantly decreased in early infection of RABV. Down-regulation of Rab5 and Rab7 did not inhibit N expression, but it prevented productive infection via blocking the normal trafficking of RABV in a low pH environment. Ultrathin sections of cells studied by electron microscope also verified the close association of RABV with Rab5 and Rab7 in neurons. From the data it was concluded that primary entry of RABV strongly correlates with the kinetics of Rab-proteins present on early and late vesicles, which provides helpful clues to explain the early events of RABV in nerve cells.

The effect of furosemide on left atrial pressure in dogs with mitral valve regurgitation.

PubMed

Suzuki, S; Ishikawa, T; Hamabe, L; Aytemiz, D; Huai-Che, H; Fukushima, R; Machida, N; Tanaka, R

2011-01-01

The effects of furosemide on left atrial pressure (LAP) in dogs with mitral regurgitation (MR) have not been documented in a quantitative manner and between different routes of administration. To document LAP and echocardiographic parameters in MR dogs administered furosemide IV or PO, in order to document changes in LAP after furosemide treatment. Five healthy Beagle dogs (3 males and 2 females; aged 2 years) were used. Experimental, cross-over, and interventional study. LAP was measured before the administration of furosemide, and 30 minutes, 1, 1.5, 2, 3, 4, 5, 6, 8, 12, and 24 hours after administration. Furosemide 1, 2, or 4 mg/kg IV, PO or placebo was administered. LAP was significantly decreased with all administrations of furosemide but not after placebo (P < .05, respectively). The max reduction was observed 1 hour (1 mg/kg IV, 15.04 ± 7.02 mmHg), 3 hours (2, 4 mg/kg IV, 13.28 ± 8.01, 9.23 ± 4.92 mmHg), 4 hours (1 mg/kg PO, 14.68 ± 11.51 mmHg), and 5 hours (2, 4 mg/kg PO, 13.19 ± 10.52, 10.70 ± 7.69 mmHg). E wave and E/Ea were significantly decreased corresponding to the reduction of LAP after administration of 2 and 4 mg/kg (P < .05, respectively). LAP was decreased in proportion to the dosage of furosemide, which did not significantly differ between IV and PO of the same dosages. E wave and E/Ea might be useful for the treatment evaluation of furosemide. Copyright © 2011 by the American College of Veterinary Internal Medicine.

Effects of treatment on respiratory rate, serum natriuretic peptide concentration, and Doppler echocardiographic indices of left ventricular filling pressure in dogs with congestive heart failure secondary to degenerative mitral valve disease and dilated cardiomyopathy.

PubMed

Schober, Karsten E; Hart, Taye M; Stern, Joshua A; Li, Xiaobai; Samii, Valerie F; Zekas, Lisa J; Scansen, Brian A; Bonagura, John D

2011-08-15

To evaluate the effects of treatment on respiratory rate, serum natriuretic peptide concentrations, and Doppler echocardiographic indices of left ventricular filling pressure in dogs with congestive heart failure (CHF) secondary to degenerative mitral valve disease (MVD) and dilated cardiomyopathy (DCM). Prospective cohort study. 63 client-owned dogs. Physical examination, thoracic radiography, analysis of natriuretic peptide concentrations, and Doppler echocardiography were performed twice, at baseline (examination 1) and 5 to 14 days later (examination 2). Home monitoring of respiratory rate was performed by the owners between examinations. In dogs with MVD, resolution of CHF was associated with a decrease in respiratory rate, serum N-terminal probrain natriuretic peptide (NT-proBNP) concentration, and diastolic functional class and an increase of the ratio of peak velocity of early diastolic transmitral flow to peak velocity of early diastolic lateral mitral annulus motion (E:Ea Lat). In dogs with DCM, resolution of CHF was associated with a decrease in respiratory rate and serum NT-proBNP concentration and significant changes in 7 Doppler echocardiographic variables, including a decrease of E:Ea Lat and the ratio of peak velocity of early diastolic transmitral flow to isovolumic relaxation time. Only respiratory rate predicted the presence of CHF at examination 2 with high accuracy. Resolution of CHF was associated with predictable changes in respiratory rate, serum NT-proBNP concentration, and selected Doppler echocardiographic variables in dogs with DCM and MVD. Home monitoring of respiratory rate was simple and was the most useful in the assessment of successful treatment of CHF.

Modeling urban air pollution with optimized hierarchical fuzzy inference system.

PubMed

Tashayo, Behnam; Alimohammadi, Abbas

2016-10-01

Environmental exposure assessments (EEA) and epidemiological studies require urban air pollution models with appropriate spatial and temporal resolutions. Uncertain available data and inflexible models can limit air pollution modeling techniques, particularly in under developing countries. This paper develops a hierarchical fuzzy inference system (HFIS) to model air pollution under different land use, transportation, and meteorological conditions. To improve performance, the system treats the issue as a large-scale and high-dimensional problem and develops the proposed model using a three-step approach. In the first step, a geospatial information system (GIS) and probabilistic methods are used to preprocess the data. In the second step, a hierarchical structure is generated based on the problem. In the third step, the accuracy and complexity of the model are simultaneously optimized with a multiple objective particle swarm optimization (MOPSO) algorithm. We examine the capabilities of the proposed model for predicting daily and annual mean PM2.5 and NO2 and compare the accuracy of the results with representative models from existing literature. The benefits provided by the model features, including probabilistic preprocessing, multi-objective optimization, and hierarchical structure, are precisely evaluated by comparing five different consecutive models in terms of accuracy and complexity criteria. Fivefold cross validation is used to assess the performance of the generated models. The respective average RMSEs and coefficients of determination (R (2)) for the test datasets using proposed model are as follows: daily PM2.5 = (8.13, 0.78), annual mean PM2.5 = (4.96, 0.80), daily NO2 = (5.63, 0.79), and annual mean NO2 = (2.89, 0.83). The obtained results demonstrate that the developed hierarchical fuzzy inference system can be utilized for modeling air pollution in EEA and epidemiological studies.

Methods for determining enzymatic activity comprising heating and agitation of closed volumes

DOEpatents

Thompson, David Neil; Henriksen, Emily DeCrescenzo; Reed, David William; Jensen, Jill Renee

2016-03-15

Methods for determining thermophilic enzymatic activity include heating a substrate solution in a plurality of closed volumes to a predetermined reaction temperature. Without opening the closed volumes, at least one enzyme is added, substantially simultaneously, to the closed volumes. At the predetermined reaction temperature, the closed volumes are agitated and then the activity of the at least one enzyme is determined. The methods are conducive for characterizing enzymes of high-temperature reactions, with insoluble substrates, with substrates and enzymes that do not readily intermix, and with low volumes of substrate and enzyme. Systems for characterizing the enzymes are also disclosed.

A biochemical and physicochemical comparison of two recombinant enzymes used for enzyme replacement therapies of hunter syndrome.

PubMed

Chung, Yo Kyung; Sohn, Young Bae; Sohn, Jong Mun; Lee, Jieun; Chang, Mi Sun; Kwun, Younghee; Kim, Chi Hwa; Lee, Jin Young; Yook, Yeon Joo; Ko, Ah-Ra; Jin, Dong-Kyu

2014-05-01

Mucopolysaccharidosis II (MPS II, Hunter syndrome; OMIM 309900) is an X-linked lysosomal storage disease caused by a deficiency in the enzyme iduronate-2-sulfatase (IDS), leading to accumulation of glycosaminoglycans (GAGs). For enzyme replacement therapy (ERT) of Hunter syndrome, two recombinant enzymes, idursulfase (Elaprase(®), Shire Human Genetic Therapies, Lexington, MA) and idursulfase beta (Hunterase(®), Green Cross Corporation, Yongin, Korea), are currently available in Korea. To compare the biochemical and physicochemical differences between idursulfase and idursulfase beta, we examined the formylglycine (FGly) content, specific enzyme activity, mannose-6-phosphate (M6P) content, sialic acid content, and in vitro cell uptake activity of normal human fibroblasts of these two enzymes.The FGly content, which determines the enzyme activity, of idursulfase beta was significantly higher than that of idursulfase (79.4 ± 0.9 vs. 68.1 ± 2.2 %, P < 0.001). In accordance with the FGly content, the specific enzyme activity of idursulfase beta was significantly higher than that of idursulfase (42.6 ± 1.1 vs. 27.8 ± 0.9 nmol/min/μg protein, P < 0.001). The levels of M6P and sialic acid were not significantly different (2.4 ± 0.1 vs 2.4 ± 0.3 mol/mol protein for M6P and 12.3 ± 0.7 vs. 12.4 ± 0.4 mol/mol protein for sialic acid). However, the cellular uptake activity of the normal human fibroblasts in vitro showed a significant difference (Kuptake, 5.09 ± 0.96 vs. 6.50 ± 1.28 nM protein, P = 0.017).In conclusion, idursulfase beta exhibited significantly higher specific enzyme activity than idursulfase, resulting from higher FGly content. These biochemical differences may be partly attributed to clinical efficacy. However, long-term clinical evaluations of Hunter syndrome patients treated with these two enzymes will be needed to demonstrate the clinical implications of significant difference of the enzyme activity and the FGly content.

Determining Enzyme Activity by Radial Diffusion

ERIC Educational Resources Information Center

Davis, Bill D.

1977-01-01

Discusses advantages of radial diffusion assay in determining presence of enzyme and/or rough approximation of amount of enzyme activities. Procedures are included for the preparation of starch-agar plates, and the application and determination of enzyme. Techniques using plant materials (homogenates, tissues, ungerminated embryos, and seedlings)…

Towards Understanding the Catalytic Mechanism of Human Paraoxonase 1: Experimental and In Silico Mutagenesis Studies.

PubMed

Tripathy, Rajan K; Aggarwal, Geetika; Bajaj, Priyanka; Kathuria, Deepika; Bharatam, Prasad V; Pande, Abhay H

2017-08-01

Human paraoxonase 1 (h-PON1) is a ~45-kDa serum enzyme that can hydrolyze a variety of substrates, including organophosphate (OP) compounds. It is a potential candidate for the development of antidote against OP poisoning in humans. However, insufficient OP-hydrolyzing activity of native enzyme affirms the urgent need to develop improved variant(s) having enhanced OP-hydrolyzing activity. The crystal structure of h-PON1 remains unsolved, and the molecular details of how the enzyme catalyses hydrolysis of different types of substrates are also not clear. Understanding the molecular details of the catalytic mechanism of h-PON1 is essential to engineer better variant(s) of enzyme. In this study, we have used a random mutagenesis approach to increase the OP-hydrolyzing activity of recombinant h-PON1. The mutants not only showed a 10-340-fold increased OP-hydrolyzing activity against different OP substrates but also exhibited differential lactonase and arylesterase activities. In order to investigate the mechanistic details of the effect of observed mutations on the hydrolytic activities of enzyme, molecular docking studies were performed with selected mutants. The results suggested that the observed mutations permit differential binding of substrate/inhibitor into the enzyme's active site. This may explain differential hydrolytic activities of the enzyme towards different substrates.

Microbial Enzyme Activity and Carbon Cycling in Grassland Soil Fractions

NASA Astrophysics Data System (ADS)

Allison, S. D.; Jastrow, J. D.

2004-12-01

Extracellular enzymes are necessary to degrade complex organic compounds present in soils. Using physical fractionation procedures, we tested whether old soil carbon is spatially isolated from degradative enzymes across a prairie restoration chronosequence in Illinois, USA. We found that carbon-degrading enzymes were abundant in all soil fractions, including macroaggregates, microaggregates, and the clay fraction, which contains carbon with a mean residence time of ~200 years. The activities of two cellulose-degrading enzymes and a chitin-degrading enzyme were 2-10 times greater in organic matter fractions than in bulk soil, consistent with the rapid turnover of these fractions. Polyphenol oxidase activity was 3 times greater in the clay fraction than in the bulk soil, despite very slow carbon turnover in this fraction. Changes in enzyme activity across the restoration chronosequence were small once adjusted for increases in soil carbon concentration, although polyphenol oxidase activity per unit carbon declined by 50% in native prairie versus cultivated soil. These results are consistent with a `two-pool' model of enzyme and carbon turnover in grassland soils. In light organic matter fractions, enzyme production and carbon turnover both occur rapidly. However, in mineral-dominated fractions, both enzymes and their carbon substrates are immobilized on mineral surfaces, leading to slow turnover. Soil carbon accumulation in the clay fraction and across the prairie restoration chronosequence probably reflects increasing physical isolation of enzymes and substrates on the molecular scale, rather than the micron to millimeter scale.

Partial purification of saccharifying and cell wall-hydrolyzing enzymes from malt in waste from beer fermentation broth.

PubMed

Khattak, Waleed Ahmad; Kang, Minkyung; Ul-Islam, Mazhar; Park, Joong Kon

2013-06-01

A number of hydrolyzing enzymes that are secreted from malt during brewing, including cell wall-hydrolyzing, saccharide-hydrolyzing, protein-degrading, lipid-hydrolyzing, and polyphenol and thiol-hydrolyzing enzymes, are expected to exist in an active form in waste from beer fermentation broth (WBFB). In this study, the existence of these enzymes was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis, after which enzyme extract was partially purified through a series of purification steps. The hydrolyzing enzyme activity was then measured under various conditions at each purification step using carboxymethyl cellulose as a substrate. The best hydrolyzing activities of partially purified enzymes were found at pH 4.5 and 50 °C in a citrate buffer system. The enzymes showed highest thermal stability at 30 °C when exposed for prolonged time. As the temperature increased gradually from 25 to 70 °C, yeast cells in the chemically defined medium with enzyme extract lost their cell wall and viability earlier than those without enzyme extract. Cell wall degradation and the release of cell matrix into the culture media at elevated temperature (45-70 °C) in the presence of enzyme extract were monitored through microscopic pictures. Saccharification enzymes from malt were relatively more active in the original WBFB than supernatant and diluted sediments. The presence of hydrolyzing enzymes from malt in WBFB is expected to play a role in bioethanol production using simultaneous saccharification and fermentation without the need for additional enzymes, nutrients, or microbial cells via a cell-free enzyme system.

Should we test TPMT enzyme levels before starting azathioprine?

PubMed

Richard, Vijay Samuel; Al-Ismail, Deana; Salamat, Ahmed

2007-08-01

Thiopurine methyltransferase (TPMT) is the main enzyme responsible for inactivating toxic products of azathioprine (AZA) metabolism. Patients with homozygous deficiency of this enzyme have no enzyme activity and ideally should not be given AZA. Patients with heterozygous deficiency have 50% of enzyme activity and have been shown to respond well and tolerate half a standard dose. We describe a patient with homozygous deficiency of TPMT who developed life threatening neutropenic sepsis, and advocate that all patients should be tested for TPMT activity prior to starting AZA therapy.

Application of carbohydrate arrays coupled with mass spectrometry to detect activity of plant-polysaccharide degradative enzymes from the fungus Aspergillus niger

PubMed Central

van Munster, Jolanda M.; Thomas, Baptiste; Riese, Michel; Davis, Adrienne L.; Gray, Christopher J.; Archer, David B.; Flitsch, Sabine L.

2017-01-01

Renewables-based biotechnology depends on enzymes to degrade plant lignocellulose to simple sugars that are converted to fuels or high-value products. Identification and characterization of such lignocellulose degradative enzymes could be fast-tracked by availability of an enzyme activity measurement method that is fast, label-free, uses minimal resources and allows direct identification of generated products. We developed such a method by applying carbohydrate arrays coupled with MALDI-ToF mass spectrometry to identify reaction products of carbohydrate active enzymes (CAZymes) of the filamentous fungus Aspergillus niger. We describe the production and characterization of plant polysaccharide-derived oligosaccharides and their attachment to hydrophobic self-assembling monolayers on a gold target. We verify effectiveness of this array for detecting exo- and endo-acting glycoside hydrolase activity using commercial enzymes, and demonstrate how this platform is suitable for detection of enzyme activity in relevant biological samples, the culture filtrate of A. niger grown on wheat straw. In conclusion, this versatile method is broadly applicable in screening and characterisation of activity of CAZymes, such as fungal enzymes for plant lignocellulose degradation with relevance to biotechnological applications as biofuel production, the food and animal feed industry. PMID:28220903

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

Velletri, P.A.; Aquilano, D.R.; Bruckwick, E.

Hypophysectomy of prepubescent (3-week-old) rats prevented the pubertal development of testicular, but not pulmonary, angiotensin-converting enzyme (EC 3.4.15.1). Additionally, hypophysectomy resulted in a loss of testicular converting enzyme activity in 10-week-old rats that had achieved puberty and had developed enzyme activity. Hormone regimens consisting of FSH/LH (7.5 U/rat X day), hCG (10 U/rat X day), or testosterone (1 mg/rat X day) were employed to ascertain their ability to maintain activity in hypophysectomized rats. All three of the above hormone regimens, if initiated on the first day after hypophysectomy of 10-week-old rats, were capable of maintaining testicular converting enzyme activity. Centrifugalmore » elutriation of dispersed testicular cells indicated that the majority of enzyme activity in mature rats was associated with the germinal cells, a result consistent with the data accumulated from the hormonal studies. Lastly, (/sup 3/H)captopril bound specifically to cellular fractions enriched in germinal cells. The above studies suggest that the pituitary gland is required for the development and maintenance of testicular angiotensin-converting enzyme in the rat by stimulating steroidogenesis in the testes. Furthermore, the sensitivity of converting enzyme activity to androgen coupled with the centrifugal elutriation and (/sup 3/H) captopril binding studies strongly support the notion that testicular converting enzyme is associated with germinal cells.« less

Expression, purification and immobilization of tannase from Staphylococcus lugdunensis MTCC 3614.

PubMed

Chaitanyakumar, Amballa; Anbalagan, M

2016-12-01

Enzymes find their applications in various industries, due to their error free conversion of substrate into product. Tannase is an enzyme used by various industries for degradation of tannin. Biochemical characterization of a specific enzyme from one organism to other is one of the ways to search for enzymes with better traits for industrial applications. Here, tannase encoding gene from Staphylococcus lugdunensis was cloned and suitability of the enzyme in various conditions was analysed to find its application in various industry. The recombinant protein was expressed with 6× His tag and purified using nickel affinity beads. The enzyme was purified up to homogeneity, with approximate molecular weight of 66 kDa. Purified tannase exhibited specific activity of about 716 U/mg. Optimum enzyme activity was found to be 40 °C at pH 7.0. Biochemical characterization revealed; metal ions such as Zn 2+ , Fe 2+ , Fe 3+ and Mn 2+ inhibited tannase activity, and SDS at lower concentration, increased tannase activity. Non polar organic solvents increased the tannase activity and polar solvents inhibited the tannase activity. Tannase immobilization studies show protection of the enzyme under wide range of pH and temperature. Also in this study we report a method for recovery and repeated use of the tannase.

A Chaperone Enhances Blood α-Glucosidase Activity in Pompe Disease Patients Treated With Enzyme Replacement Therapy

PubMed Central

Parenti, Giancarlo; Fecarotta, Simona; la Marca, Giancarlo; Rossi, Barbara; Ascione, Serena; Donati, Maria Alice; Morandi, Lucia Ovidia; Ravaglia, Sabrina; Pichiecchio, Anna; Ombrone, Daniela; Sacchini, Michele; Pasanisi, Maria Barbara; De Filippi, Paola; Danesino, Cesare; Della Casa, Roberto; Romano, Alfonso; Mollica, Carmine; Rosa, Margherita; Agovino, Teresa; Nusco, Edoardo; Porto, Caterina; Andria, Generoso

2014-01-01

Enzyme replacement therapy is currently the only approved treatment for Pompe disease, due to acid α-glucosidase deficiency. Clinical efficacy of this approach is variable, and more effective therapies are needed. We showed in preclinical studies that chaperones stabilize the recombinant enzyme used for enzyme replacement therapy. Here, we evaluated the effects of a combination of enzyme therapy and a chaperone on α-glucosidase activity in Pompe disease patients. α-Glucosidase activity was analyzed by tandem-mass spectrometry in dried blood spots from patients treated with enzyme replacement therapy, either alone or in combination with the chaperone N-butyldeoxynojirimycin given at the time of the enzyme infusion. Thirteen patients with different presentations (3 infantile-onset, 10 late-onset) were enrolled. In 11 patients, the combination treatment resulted in α-glucosidase activities greater than 1.85-fold the activities with enzyme replacement therapy alone. In the whole patient population, α-glucosidase activity was significantly increased at 12 hours (2.19-fold, P = 0.002), 24 hours (6.07-fold, P = 0.001), and 36 hours (3.95-fold, P = 0.003). The areas under the curve were also significantly increased (6.78-fold, P = 0.002). These results suggest improved stability of recombinant α-glucosidase in blood in the presence of the chaperone. PMID:25052852

Dihydroquercetin Does Not Affect Age-Dependent Increase in Blood Pressure and Angiotensin-Converting Enzyme Activity in the Aorta of Hypertensive Rats.

PubMed

Slashcheva, G A; Rykov, V A; Lobanov, A V; Murashev, A N; Kim, Yu A; Arutyunyan, T V; Korystova, A F; Kublik, L N; Levitman, M Kh; Shaposhnikona, V V; Korystov, Yu N

2016-09-01

We analyzed changes in angiotensin-converting enzyme activity in the aorta of hypertensive SHR rats against the background of age-related BP increase (from week 7 to 14) and the effect of dihydroquercetin on BP rise and angiotensin-converting enzyme activity. Normotensive WKY rats of the same age were used as the control. BP and activity of angiotensin-converting enzyme in the aorta of SHR rats increased with age. Dihydroquercetin in doses of 100 and 300 μg/kg per day had no effect on the increase of these parameters; dihydroquercetin administered to 14-week-old WKY rats in a dose of 300 μg/kg reduced activity of the angiotensin-converting enzyme. Thus, the early (7-14 weeks) increase in BP and angiotensin-converting enzyme activity in the aorta of SHR rats was not modified by flavonoids (dihydroquercetin) in contrast to other rat strains and humans, which is indicative of specificity of hypertension mechanism in SHR rats.

Enhanced stability and catalytic activity of immobilized α-amylase on modified Fe3O4 nanoparticles for potential application in food industries

NASA Astrophysics Data System (ADS)

Hosseinipour, Seyyedeh Leila; Khiabani, Mahmoud Sowti; Hamishehkar, Hamed; Salehi, Roya

2015-09-01

Enzymes play an essential role in catalyzing various reactions. However, their instability upon repetitive/prolonged use, elevated temperature, acidic or alkaline pH remains an area of concern. α-Amylase, a widely used enzyme in food industries for starch hydrolysis, was covalently immobilized on the surface of two developed matrices, amino-functionalized silica-coated magnetite nanoparticles (AFSMNPs) alone and covered with chitosan. The synthesis steps and characterizations of NPs were examined by FT-IR, VSM, and SEM. Modified nanoparticles with average diameters of 20-80 nm were obtained. Enzyme immobilization efficiencies of 89 and 74 were obtained for AFSMNPs and chitosan-coated AFSMNPs, respectively. The optimum pH obtained was 6.5 and 8.0 for the enzyme immobilized on AFSMNPs and chitosan-coated AFSMNPs, respectively. Optimum temperature for the immobilized enzyme shifted toward higher temperatures. Considerable enhancements in thermal stabilities were observed for the immobilized enzyme at elevated temperatures up to 80 °C. A frequent use experiment demonstrated that the immobilized enzyme retained 74 and 85 % of its original activity even after 20 times of repeated use in AFSMNPs and chitosan-coated AFSMNPs, respectively. Storage stability demonstrated that free enzyme lost its activity completely within 30 days. But, immobilized enzyme on AFSMNPs and chitosan-coated AFSMNPs preserved 65.73 and 78.63 % of its initial activity, respectively, after 80 days of incubation. In conclusion, a substantial improvement in the performance of the immobilized enzyme with reference to the free enzyme was obtained. Furthermore, the relative activities of immobilized enzyme are superior than free enzyme over the broader pH and temperature ranges.

Comparison of Free and Immobilized L-asparaginase Synthesized by Gamma-Irradiated Penicillium cyclopium.

PubMed

El-Refai, Heba A; Shafei, Mona S; Mostafa, Hanan; El-Refai, Abdel-Monem H; Araby, Eman M; El-Beih, Fawkia M; Easa, Saadia M; Gomaa, Sanaa K

2016-01-01

Gamma irradiation is used on Penicillium cyclopium in order to obtain mutant cells of high L-asparaginase productivity. Using gamma irradiation dose of 4 KGy, P. cyclopium cells yielded L-asparaginase with extracellular enzyme activity of 210.8 ± 3 U/ml, and specific activity of 752.5 ± 1.5 U/mg protein, which are 1.75 and 1.53 times, respectively, the activity of the wild strain. The enzyme was partially purified by 40-60% acetone precipitation. L-asparaginase was immobilized onto Amberlite IR-120 by ionic binding. Both free and immobilized enzymes exhibited maximum activity at pH 8 and 40 degrees C. The immobilization process improved the enzyme thermal stability significantly. The immobilized enzyme remained 100% active at temperatures up to 60 degrees C, while the free asparaginase was less tolerant to high temperatures. The immobilized enzyme was more stable at pH 9.0 for 50 min, retaining 70% of its relative activity. The maximum reaction rate (V(max)) and Michaelis-Menten constant (K(m)) of the free form were significantly changed after immobilization. The K(m) value for immobilized L-asparaginase was about 1.3 times higher than that of free enzyme. The ions K+, Ba2+ and Na+ showed stimulatory effect on enzyme activity with percentages of 110%, 109% and 106% respectively.

Catalytic power of enzymes decreases with temperature: New insights for understanding soil C cycling and microbial ecology under warming.

PubMed

Alvarez, Gaël; Shahzad, Tanvir; Andanson, Laurence; Bahn, Michael; Wallenstein, Matthew D; Fontaine, Sébastien

2018-04-23

Most current models of soil C dynamics predict that climate warming will accelerate soil C mineralization, resulting in a long-term CO 2 release and positive feedback to global warming. However, ecosystem warming experiments show that CO 2 loss from warmed soils declines to control levels within a few years. Here, we explore the temperature dependence of enzymatic conversion of polymerized soil organic C (SOC) into assimilable compounds, which is presumed the rate-limiting step of SOC mineralization. Combining literature review, modelling and enzyme assays, we studied the effect of temperature on activity of enzymes considering their thermal inactivation and catalytic activity. We defined the catalytic power of enzymes (E power ) as the cumulative amount of degraded substrate by one unit of enzyme until its complete inactivation. We show a universal pattern of enzyme's thermodynamic properties: activation energy of catalytic activity (EA cat ) < activation energy of thermal inactivation (EA inact ). By investing in stable enzymes (high EA inact ) having high catalytic activity (low EA cat ), microorganisms may maximize the E power of their enzymes. The counterpart of such EAs' hierarchical pattern is the higher relative temperature sensitivity of enzyme inactivation than catalysis, resulting in a reduction in E power under warming. Our findings could explain the decrease with temperature in soil enzyme pools, microbial biomass (MB) and carbon use efficiency (CUE) reported in some warming experiments and studies monitoring the seasonal variation in soil enzymes. They also suggest that a decrease in soil enzyme pools due to their faster inactivation under warming contributes to the observed attenuation of warming effect on soil C mineralization. This testable theory predicts that the ultimate response of SOC degradation to warming can be positive or negative depending on the relative temperature response of E power and microbial production of enzymes. © 2018 John Wiley & Sons Ltd.

A Simulation Game for the Study of Enzyme Kinetics and Inhibition.

ERIC Educational Resources Information Center

Chayoth, Reuben; Cohen, Annette

1996-01-01

Presents a simulation game that facilitates understanding of the concepts of enzyme kinetics and inhibition. The first part of the game deals with the relationship between enzyme activity and substrate concentration while the second part deals with characterization of competitive and noncompetitive inhibition of enzyme activity. (JRH)

A Hereditary Spastic Paraplegia Mouse Model Supports a Role of ZFYVE26/SPASTIZIN for the Endolysosomal System

PubMed Central

Khundadze, Mukhran; Kollmann, Katrin; Koch, Nicole; Biskup, Christoph; Nietzsche, Sandor; Zimmer, Geraldine; Hennings, J. Christopher; Huebner, Antje K.; Symmank, Judit; Jahic, Amir; Ilina, Elena I.; Karle, Kathrin; Schöls, Ludger; Kessels, Michael; Braulke, Thomas; Qualmann, Britta; Kurth, Ingo; Beetz, Christian; Hübner, Christian A.

2013-01-01

Hereditary spastic paraplegias (HSPs) are characterized by progressive weakness and spasticity of the legs because of the degeneration of cortical motoneuron axons. SPG15 is a recessively inherited HSP variant caused by mutations in the ZFYVE26 gene and is additionally characterized by cerebellar ataxia, mental decline, and progressive thinning of the corpus callosum. ZFYVE26 encodes the FYVE domain-containing protein ZFYVE26/SPASTIZIN, which has been suggested to be associated with the newly discovered adaptor protein 5 (AP5) complex. We show that Zfyve26 is broadly expressed in neurons, associates with intracellular vesicles immunopositive for the early endosomal marker EEA1, and co-fractionates with a component of the AP5 complex. As the function of ZFYVE26 in neurons was largely unknown, we disrupted Zfyve26 in mice. Zfyve26 knockout mice do not show developmental defects but develop late-onset spastic paraplegia with cerebellar ataxia confirming that SPG15 is caused by ZFYVE26 deficiency. The morphological analysis reveals axon degeneration and progressive loss of both cortical motoneurons and Purkinje cells in the cerebellum. Importantly, neuron loss is preceded by accumulation of large intraneuronal deposits of membrane-surrounded material, which co-stains with the lysosomal marker Lamp1. A density gradient analysis of brain lysates shows an increase of Lamp1-positive membrane compartments with higher densities in Zfyve26 knockout mice. Increased levels of lysosomal enzymes in brains of aged knockout mice further support an alteration of the lysosomal compartment upon disruption of Zfyve26. We propose that SPG15 is caused by an endolysosomal membrane trafficking defect, which results in endolysosomal dysfunction. This appears to be particularly relevant in neurons with highly specialized neurites such as cortical motoneurons and Purkinje cells. PMID:24367272

A hereditary spastic paraplegia mouse model supports a role of ZFYVE26/SPASTIZIN for the endolysosomal system.

PubMed

Khundadze, Mukhran; Kollmann, Katrin; Koch, Nicole; Biskup, Christoph; Nietzsche, Sandor; Zimmer, Geraldine; Hennings, J Christopher; Huebner, Antje K; Symmank, Judit; Jahic, Amir; Ilina, Elena I; Karle, Kathrin; Schöls, Ludger; Kessels, Michael; Braulke, Thomas; Qualmann, Britta; Kurth, Ingo; Beetz, Christian; Hübner, Christian A

2013-01-01

Hereditary spastic paraplegias (HSPs) are characterized by progressive weakness and spasticity of the legs because of the degeneration of cortical motoneuron axons. SPG15 is a recessively inherited HSP variant caused by mutations in the ZFYVE26 gene and is additionally characterized by cerebellar ataxia, mental decline, and progressive thinning of the corpus callosum. ZFYVE26 encodes the FYVE domain-containing protein ZFYVE26/SPASTIZIN, which has been suggested to be associated with the newly discovered adaptor protein 5 (AP5) complex. We show that Zfyve26 is broadly expressed in neurons, associates with intracellular vesicles immunopositive for the early endosomal marker EEA1, and co-fractionates with a component of the AP5 complex. As the function of ZFYVE26 in neurons was largely unknown, we disrupted Zfyve26 in mice. Zfyve26 knockout mice do not show developmental defects but develop late-onset spastic paraplegia with cerebellar ataxia confirming that SPG15 is caused by ZFYVE26 deficiency. The morphological analysis reveals axon degeneration and progressive loss of both cortical motoneurons and Purkinje cells in the cerebellum. Importantly, neuron loss is preceded by accumulation of large intraneuronal deposits of membrane-surrounded material, which co-stains with the lysosomal marker Lamp1. A density gradient analysis of brain lysates shows an increase of Lamp1-positive membrane compartments with higher densities in Zfyve26 knockout mice. Increased levels of lysosomal enzymes in brains of aged knockout mice further support an alteration of the lysosomal compartment upon disruption of Zfyve26. We propose that SPG15 is caused by an endolysosomal membrane trafficking defect, which results in endolysosomal dysfunction. This appears to be particularly relevant in neurons with highly specialized neurites such as cortical motoneurons and Purkinje cells.

[Effects of snow pack on soil nitrogen transformation enzyme activities in a subalpine Abies faxioniana forest of western Sichuan, China].

PubMed

Xiong, Li; Xu, Zhen-Feng; Wu, Fu-Zhong; Yang, Wan-Qin; Yin, Rui; Li, Zhi-Ping; Gou, Xiao-Lin; Tang, Shi-Shan

2014-05-01

This study characterized the dynamics of the activities of urease, nitrate reductase and nitrite reductase in both soil organic layer and mineral soil layer under three depths of snow pack (deep snowpack, moderate snowpack and shallow snowpack) over the three critical periods (snow formed period, snow stable period, and snow melt period) in the subalpine Abies faxoniana forest of western Sichuan in the winter of 2012 and 2013. Throughout the winter, soil temperature under deep snowpack increased by 46.2% and 26.2%, respectively in comparison with moderate snowpack and shallow snowpack. In general, the three nitrogen-related soil enzyme activities under shallow snowpack were 0.8 to 3.9 times of those under deep snowpack during the winter. In the beginning and thawing periods of seasonal snow pack, shallow snowpack significantly increased the activities of urease, nitrate and nitrite reductase enzyme in both soil organic layer and mineral soil layer. Although the activities of the studied enzymes in soil organic layer and mineral soil layer were observed to be higher than those under deep- and moderate snowpacks in deep winter, no significant difference was found under the three snow packs. Meanwhile, the effects of snowpack on the activities of the measured enzymes were related with season, soil layer and enzyme type. Significant variations of the activities of nitrogen-related enzymes were found in three critical periods over the winter, and the three measured soil enzymes were significantly higher in organic layer than in mineral layer. In addition, the activities of the three measured soil enzymes were closely related with temperature and moisture in soils. In conclusion, the decrease of snow pack induced by winter warming might increase the activities of soil enzymes related with nitrogen transformation and further stimulate the process of wintertime nitrogen transformation in soils of the subalpine forest.

Enzyme activity in dialkyl phosphate ionic liquids

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

Thomas, M.F.; Dunn, J.; Li, L.-L.

2011-12-01

The activity of four metagenomic enzymes and an enzyme cloned from the straw mushroom, Volvariellavolvacea were studied in the following ionic liquids, 1,3-dimethylimidazolium dimethyl phosphate, [mmim][dmp], 1-ethyl-3-methylimidazolium dimethyl phosphate, [emim][dmp], 1-ethyl-3-methylimidazolium diethyl phosphate, [emim][dep] and 1-ethyl-3-methylimidazolium acetate, [emim][OAc]. Activity was determined by analyzing the hydrolysis of para-nitrobenzene carbohydrate derivatives. In general, the enzymes were most active in the dimethyl phosphate ionic liquids, followed by acetate. Generally speaking, activity decreased sharply for concentrations of [emim][dep] above 10% v/v, while the other ionic liquids showed less impact on activity up to 20% v/v.

Some enzymes of carbohydrate metabolism in Mesocestoides corti and Heterakis spumosa.

PubMed

Dubinský, P; Ruscinová, B; Hetmanski, S L; Arme, C; Turceková, L; Rybos, M

1991-09-01

The activities of selected enzymes of carbohydrate metabolism were measured in tetrathyridia of Mesocestoides corti and in adult females and males of Heterakis spumosa. When the species were compared, only lactate dehydrogenase and phosphoenolpyruvate carboxykinase activities were considerably higher in M. corti. Activities of other enzymes were higher in H. spumosa, with malate dehydrogenase activity being considerably so. In H. spumosa, enzyme activity was higher, and succinate dehydrogenase markedly so in males, when compared with females. Tetrathyridia aged 170 and 210 days show relatively stable malate and lactate dehydrogenase activities, and mice of ICR and BALB/c strains are suitable for the maintenance of tetrathyridia.

In-vitro engineering of novel bioactivity in the natural enzymes

NASA Astrophysics Data System (ADS)

Tiwari, Vishvanath

2016-10-01

Enzymes catalyze various biochemical functions with high efficiency and specificity. In-vitro design of the enzyme leads to novel bioactivity in this natural biomolecule that give answers of some vital questions like crucial residues in binding with substrate, molecular evolution, cofactor specificity etc. Enzyme engineering technology involves directed evolution, rational designing, semi-rational designing and structure-based designing using chemical modifications. Similarly, combined computational and in-vitro evolution approaches together help in artificial designing of novel bioactivity in the natural enzyme. DNA shuffling, error prone PCR and staggered extension process are used to artificially redesign active site of enzyme, which can alter its efficiency and specificity. Modifications of the enzyme can lead to the discovery of new path of molecular evolution, designing of efficient enzymes, locating active sites and crucial residues, shift in substrate and cofactor specificity. The methods and thermodynamics of in-vitro designing of the enzyme are also discussed. Similarly, engineered thermophilic and psychrophilic enzymes attain substrate specificity and activity of mesophilic enzymes that may also be beneficial for industry and therapeutics.

Dried blood spots for the enzymatic diagnosis of lysosomal storage diseases in dogs and cats.

PubMed

Sewell, Adrian C; Haskins, Mark E; Giger, Urs

2012-12-01

In people, lysosomal storage diseases (LSD) can be diagnosed by assaying enzyme activities in dried blood spots (DBS). The aim of this study was to evaluate the feasibility of using DBS samples from dogs and cats to measure lysosomal enzymatic activities and diagnose LSD. Drops of fresh whole blood collected in EDTA from dogs and cats with known or suspected LSD and from clinically healthy dogs and cats were placed on neonatal screening cards, dried, and mailed to the Metabolic Laboratory, University Children's Hospital, Frankfurt, Germany. Activities of selected lysosomal enzymes were measured using fluorescent substrates in a 2-mm diameter disk (~2.6 μL blood) punched from the DBS. Results were expressed as nmol substrate hydrolyzed per mL of blood per minute or hour. Reference values were established for several lysosomal enzyme activities in DBS from dogs and cats; for most enzymes, activities were higher than those published for human samples. Activities of β-glucuronidase, N-acetylglucosamine-4-sulfatase (arylsulfatase B), α-mannosidase, α-galactosidase, α-fucosidase, and hexosaminidase A were measureable in DBS from healthy cats and dogs; α-iduronidase activity was measureable only in cats. In samples from animals with LSD, markedly reduced activity of a specific enzyme was found. In contrast, in samples from cats affected with mucolipidosis II, activities of lysosomal enzymes were markedly increased. Measurement of lysosomal enzyme activities in DBS provides an inexpensive, simple, and convenient method to screen animals for suspected LSD and requires only a small sample volume. For diseases in which the relevant enzyme activity can be measured in DBS, a specific diagnosis can be made. © 2012 American Society for Veterinary Clinical Pathology.

Activities of five enzymes following soil disturbance and weed control in a Missouri forest

Treesearch

Felix, Jr. Ponder; Frieda Eivazi

2008-01-01

Forest disturbances associated with harvesting activities can affect soil properties including enzyme activity and overall soil quality. The activities of five enzymes (acid and alkaline phosphatases, betaglucosidase, aryl-sulfatase, and beta-glucosominidase) were measured after 8 years in soil from clearcut and uncut control plots of a Missouri oak-hickory (...

Purification and properties of a novel ferricyanide-linked xanthine dehydrogenase from Pseudomonas putida 40.

PubMed Central

Woolfolk, C A

1985-01-01

The isolation of a xanthine dehydrogenase from Pseudomonas putida 40 which utilizes ferricyanide as an electron acceptor at high efficiency is presented. The new activity is separate from the NAD+ and oxygen-utilizing activities of the same organism but displays a broad pattern for reducing substrates typical of those of previously studied xanthine-oxidizing enzymes. Unlike the previously studied enzymes, the new enzyme appears to lack flavin but possess heme and is resistant to cyanide treatment. However, sensitivity of the purified enzyme to methanol and the selective elimination of the activity when tungstate is added to certain growth media suggest a role for molybdenum. The enzyme is subject to a selective proteolytic action during processing which is not accompanied by denaturation or loss of activity and which is minimized by the continuous exposure of the activity to EDTA and phenylmethylsulfonyl fluoride. Electrophoresis of the denatured enzyme in the presence of sodium dodecyl sulfate suggests that the enzyme is constructed of subunits with a molecular weight of approximately 72,000. Electrophoresis under native conditions of a purified enzyme previously exposed to magnesium ion reveals a series of major and minor activity bands which display some selectivity toward both electron donors and acceptors. An analysis of the effect of gel concentration on this pattern suggests that the enzyme forms a series of charge and size isomers with a pair of trimeric forms predominating. Comparison of the rate of sedimentation of the enzyme in sucrose gradients with its elution profile from standardized Sepharose 6B columns suggests a molecular weight of 255,000 for the major form of the native enzyme. Images PMID:3860496

Gene expression and activity of digestive enzymes of Daphnia pulex in response to food quality differences.

PubMed

Schwarzenberger, Anke; Fink, Patrick

2018-04-01

Food quality is an important factor influencing organisms' well-being. In freshwater ecosystems, food quality has been studied extensively for the keystone herbivore genus Daphnia, as they form the critical trophic link between primary producers and higher order consumers such as fish. For Daphnia, the edible fraction of phytoplankton in lakes (consisting mostly of unicellular algae and cyanobacteria) is extraordinarily diverse. To be able to digest different food particles, Daphnia possess a set of digestive enzymes that metabolize carbohydrates, lipids and proteins. Recent studies have found a connection between gene expression and activity of single digestive enzyme types of Daphnia, i.e. lipases and proteases, and transcriptome studies have shown that a variety of genes coding for gut enzymes are differentially expressed in response to different food algae. However, never before has a set of digestive enzymes been studied simultaneously both on the gene expression and the enzyme activity level in Daphnia. Here, we investigated several digestive enzymes of Daphnia pulex in a comparison between a high-quality (green algal) and a low-quality (cyanobacterial) diet. Diet significantly affected the expression of all investigated digestive enzyme genes and enzyme activity was altered between treatments. Furthermore, we found that gene expression and enzyme activity were significantly correlated in cellulase, triacylglycerol lipase and β-glucosidase when switched from high to low-quality food. We conclude that one of the factors causing the often observed low biomass and energy transfer efficiency from cyanobacteria to Daphnia is probably the switch to a cost-effective overall increase of gene expression and activity of digestive enzymes of this herbivore. Copyright © 2018 Elsevier Inc. All rights reserved.

Activity loss by H46A mutation in Mycobacterium tuberculosis isocitrate lyase is due to decrease in structural plasticity and collective motions of the active site.

PubMed

Shukla, Rohit; Shukla, Harish; Tripathi, Timir

2018-01-01

Mycobacterium tuberculosis isocitrate lyase (MtbICL) is a crucial enzyme of the glyoxylate cycle and is a validated anti-tuberculosis drug target. Structurally distant, non-active site mutation (H46A) in MtbICL has been found to cause loss of enzyme activity. The aim of the present work was to explore the structural alterations induced by H46A mutation that caused the loss of enzyme activity. The structural and dynamic consequences of H46A mutation were studied using multiple computational methods such as docking, molecular dynamics simulation and residue interaction network analysis (RIN). Principal component analysis and cross correlation analysis revealed the difference in conformational flexibility and collective modes of motions between the wild-type and mutant enzyme, particularly in the active site region. RIN analysis revealed that the active site geometry was disturbed in the mutant enzyme. Thus, the dynamic perturbation of the active site led to enzyme transition from its active form to inactive form upon mutation. The computational analyses elucidated the mutant-specific conformational alterations, differential dominant motions, and anomalous residue level interactions that contributed to the abrogated function of mutant MtbICL. An understanding of interactions of mutant enzymes may help in modifying the existing drugs and designing improved drugs for successful control of tuberculosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

Ecotoxicological effects of copper and selenium combined pollution on soil enzyme activities in planted and unplanted soils.

PubMed

Hu, Bin; Liang, Dongli; Liu, Juanjuan; Xie, Junyu

2013-04-01

The present study explored the joint effects of Cu and Se pollution mechanisms on soil enzymes to provide references for the phytoremediation of contaminated areas and agricultural environmental protection. Pot experiments and laboratory analyses were carried out to study the individual and combined influences of Cu and Se on soil enzyme activities. The activities of four soil enzymes (urease, catalase, alkaline phosphatase, and nitrate reductase) were chosen. All soil enzyme activities tested were inhibited by Cu and Se pollution, either individually or combined, in varying degrees, following the order nitrate reductase>urease>catalase>alkaline phosphatase. Growing plants stimulated soil enzyme activity in a similar trend compared with treatments without plants. The joint effects of Cu and Se on catalase activity showed synergism at low concentrations and antagonism at high concentrations, whereas the opposite was observed for urease activity. However, nitrate reductase activity showed synergism both with and without plant treatments. The half maximal effective concentration (EC50) of exchangeable fractions had a similar trend with the EC50 of total content and was lower than that of total content. The EC50 values of nitrate reductase and urease activities were significantly lower for both Se and Cu (p<0.05), which indicated that they were more sensitive than the other two enzymes. Copyright © 2013 SETAC.

Comparative kinetic studies of Mn2+-activated and fructose-1,6-P-modified Mg2+-activated pyruvate kinase from Concholepas concholepas.

PubMed

Carvajal, N; González, R; Morán, A; Oyarce, A M

1985-01-01

Initial velocity and product inhibition studies of Mn2+-activated and FDP-modified Mg2+-activated pyruvate kinase from Concholepas concholepas, were performed. Evidence is presented to show that the Mn2+-enzyme catalyzes an ordered sequential mechanism, with ADP being the first substrate and pyruvate the last product. The results presented are consistent with a random combination of reactants with the FDP-modified Mg2+-activated enzyme and the formation of the dead-end complexes enzyme ADP-ATP and enzyme-PEP-ATP.

The Molecular Basis of Dominance

PubMed Central

Kacser, Henrik; Burns, James A.

1981-01-01

The best known genes of microbes, mice and men are those that specify enzymes. Wild type, mutant and heterozygote for variants of such genes differ in the catalytic activity at the step in the enzyme network specified by the gene in question. The effect on the respective phenotypes of such changes in catalytic activity, however, is not defined by the enzyme change as estimated by in vitro determination of the activities obtained from the extracts of the three types. In vivo enzymes do not act in isolation, but are kinetically linked to other enzymes via their substrates and products. These interactions modify the effect of enzyme variation on the phenotype, depending on the nature and quantity of the other enzymes present. An output of such a system, say a flux, is therefore a systemic property, and its response to variation at one locus must be measured in the whole system. This response is best described by the sensitivity coefficient, Z, which is defined by the fractional change in flux over the fractional change in enzyme activity.(see PDF)Its magnitude determines the extent to which a particular enzyme "controls" a particular flux or phenotype and, implicitly, determines the values that the three phenotypes will have. There are as many sensitivity coefficients for a given flux as there are enzymes in the system. It can be shown that the sum of all such coefficients equals unity.(see PDF)Since n, the number of enzymes, is large, this summation property results in the individual coefficients being small. The effect of making a large change in enzyme activity therefore usually results in only a negligible change in flux. A reduction to 50% activity in the heterozygote, a common feature for many mutants, is therefore not expected to be detectable in the phenotype. The mutant would therefore be described as "recessive". The widespread occurrence of recessive mutants is thus seen to be the inevitable consequence of the kinetic structure of enzyme networks. The ad hoc hypothesis of "modifiers" selected to maximize the fitness of the heterozygote, as proposed by Fisher, is therefore unnecessary. It is based on the false general expectation of an intermediate phenotype in the heterozygote. Wright's analysis, substantially sound in its approach, proposed selection of a "safety factor" in enzyme activity. The derivation of the summation property explains why such safety factors are automatically present in almost all enzymes without selection. PMID:7297851

Microbial dynamics and enzyme activities in tropical Andosols depending on land use and nutrient inputs

NASA Astrophysics Data System (ADS)

Mganga, Kevin; Razavi, Bahar; Kuzyakov, Yakov

2015-04-01

Microbial decomposition of soil organic matter is mediated by enzymes and is a key source of terrestrial CO2 emissions. Microbial and enzyme activities are necessary to understand soil biochemical functioning and identify changes in soil quality. However, little is known about land use and nutrients availability effects on enzyme activities and microbial processes, especially in tropical soils of Africa. This study was conducted to examine how microbial and enzyme activities differ between different land uses and nutrient availability. As Andosols of Mt. Kilimanjaro are limited by nutrient concentrations, we hypothesize that N and P additions will stimulate enzyme activity. N and P were added to soil samples (0-20 cm) representing common land use types in East Africa: (1) savannah, (2) maize fields, (3) lower montane forest, (4) coffee plantation, (5) grasslands and (6) traditional Chagga homegardens. Total CO2 efflux from soil, microbial biomass and activities of β-glucosidase, cellobiohydrolase, chitinase and phosphatase involved in C, N and P cycling, respectively was monitored for 60 days. Total CO2 production, microbial biomass and enzyme activities varied in the order forest soils > grassland soils > arable soils. Increased β-glucosidase and cellobiohydrolase activities after N addition of grassland soils suggest that microorganisms increased N uptake and utilization to produce C-acquiring enzymes. Low N concentration in all soils inhibited chitinase activity. Depending on land use, N and P addition had an inhibitory or neutral effect on phosphatase activity. We attribute this to the high P retention of Andosols and low impact of N and P on the labile P fractions. Enhanced CO2 production after P addition suggests that increased P availability could stimulate soil organic matter biodegradation in Andosols. In conclusion, land use and nutrients influenced soil enzyme activities and microbial dynamics and demonstrated the decline in soil quality after landuse change. Key words: Andosols, β-glucosidase, Cellobiohydrolase, Chitinase, Phosphatase, Mt. Kilimanjaro

Enzyme Activity Experiments Using a Simple Spectrophotometer

ERIC Educational Resources Information Center

Hurlbut, Jeffrey A.; And Others

1977-01-01

Experimental procedures for studying enzyme activity using a Spectronic 20 spectrophotometer are described. The experiments demonstrate the effect of pH, temperature, and inhibitors on enzyme activity and allow the determination of Km, Vmax, and Kcat. These procedures are designed for teaching large lower-level biochemistry classes. (MR)

[Effect of space flight on the Kosmos-1129 biosatellite on enzyme activity of the rat liver].

PubMed

Nemeth, S; Tigranian, R A

1983-01-01

After the 18.5 day Cosmos-1129 flight the activity of 7 glucocorticoid-stimulated enzymes of the rat liver was measured. Immediately postflight the activity of tyrosine aminotransferase, tryptophan pyrolase and serine dehydrogenase increased. These enzymes rapidly (within several hours) react to increased glucocorticoids. The activity of aspartate and alanine aminotransferases also increased. These enzymes require many days of a continuous effect of glucocorticoids. The glycogen concentration in the rat liver also grew. At R + 6 the activity of tryptophan pyrolase and serine dehydrogenase decreased and that of the other enzymes returned to normal. The immobilization stress applied postflight led to an increased activity of tyrosine aminotransferase and tryptophan pyrolase. This study gives evidence that after space flight rats are in an acute stress state, evidently, produced by the biosatellite recovery.

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

Small heat shock protein AgsA: an effective stabilizer of enzyme activities.

PubMed

Tomoyasu, Toshifumi; Tabata, Atsushi; Ishikawa, Yoko; Whiley, Robert A; Nagamune, Hideaki

2013-01-01

A small heat shock protein, AgsA, possesses chaperone activity that can reduce the amount of heat-aggregated protein in vivo, and suppress the aggregation of chemical- and heat-denatured proteins in vitro. Therefore, we examined the ability of AgsA to stabilize the activity of several enzymes by using this chaperone activity. We observed that AgsA can stabilize the enzymatic activities of Renilla (Renilla reniformis) luciferase, firefly (Photinus pyralis) luciferase, and β-galactosidase, and showed comparable or greater stabilization of these enzymes than bovine serum albumin (BSA), a well-known stabilizer of enzyme activities. In particular, AgsA revealed better stabilization of Renilla luciferase and β-galactosidase than BSA under disulfide bond-reducing conditions with dithiothreitol. In addition, AgsA also increased the enzymatic performance of β-galactosidase and various restriction enzymes to a comparable or greater extent than BSA. These data indicate that AgsA may be useful as a general stabilizer of enzyme activities. Copyright © 2012 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

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

Enzyme-polymer composites with high biocatalytic activity and stability

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

Kim, Jungbae; Kosto, Timothy J.; Manimala, Joseph C.

2004-08-22

We have applied vacuum-spraying and electrospinning to incorporate an enzyme into a polymer matrix, creating a novel and highly active biocatalytic composite. As a unique technical approach, enzymes were co-dissolved in toluene with polymers, and the solvent was then rapidly removed by injecting the mixture into a vacuum chamber or by electrospinning. Subsequent crosslinking of the enzyme with glutaraldehyde resulted in stable entrapped enzyme within the polymeric matrices. For example, an amorphous composite of alpha-chymotrypsin and polyethylene showed no significant loss of enzymatic activity in aqueous buffer for one month. Nanofibers of alpha-chymotrypsin and polystyrene also showed no decrease inmore » activity for more than two weeks. The normalized activity of amorphous composite in organic solvents was 3-13 times higher than that of native alpha-chymotrypsin. The activity of nanofibers was 5-7 times higher than that of amorphous composite in aqueous buffer solution. The composites of alpha-chymotrypsin and polymers demonstrate the feasibility of obtaining a wide variety of active and stable biocatalytic materials with many combinations of enzymes and polymers.« less

In vitro antibody-enzyme conjugates with specific bactericidal activity.

PubMed

Knowles, D M; Sulivan, T J; Parker, C W; Williams, R C

1973-06-01

IgG with antibacterial antibody opsonic activity was isolated from rabbit antisera produced by intravenous hyperimmunization with several test strains of pneumococci, Group A beta-hemolytic streptococci, Staphylococcus aureus, Proteus mirabilis, Pseudomonas aeruginosa, and Escherichia coli. Antibody-enzyme conjugates were prepared, using diethylmalonimidate to couple glucose oxidase to IgG antibacterial antibody preparations. Opsonic human IgG obtained from serum of patients with subacute bacterial endocarditis was also conjugated to glucose oxidase. Antibody-enzyme conjugates retained combining specificity for test bacteria as demonstrated by indirect immunofluorescence. In vitro test for bactericidal activity of antibody-enzyme conjugates utilized potassium iodide, lactoperoxidase, and glucose as cofactors. Under these conditions glucose oxidase conjugated to antibody generates hydrogen peroxide, and lactoperoxidase enzyme catalyzes the reduction of hydrogen peroxide with simultaneous oxidation of I(-) and halogenation and killing of test bacteria. Potent in vitro bactericidal activity of this system was repeatedly demonstrated for antibody-enzyme conjugates against pneumococci, streptococci, S. aureus, P. mirabilis, and E. coli. However, no bactericidal effect was demonstrable with antibody-enzyme conjugates and two test strains of P. aeruginosa. Bactericidal activity of antibody-enzyme conjugates appeared to parallel original opsonic potency of unconjugated IgG preparations. Antibody-enzyme conjugates at concentrations as low as 0.01 mg/ml were capable of intense bactericidal activity producing substantial drops in surviving bacterial counts within 30-60 min after initiation of assay. These in vitro bactericidal systems indicate that the concept of antibacterial antibody-enzyme conjugates may possibly be adaptable as a mechanism for treatment of patients with leukocyte dysfunction or fulminant bacteremia.

Enzymes in Commercial Cellulase Preparations Bind Differently to Dioxane Extracted Lignins

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

Yarbrough, John M.; Mittal, Ashutosh; Katahira, Rui

Commercial fungal cellulases used in biomass-to-biofuels processes can be grouped into three general classes: native, augmented, and engineered. To evaluate lignin binding affinities of different enzyme activities in various commercial cellulase formulations in order to determine if enzyme losses due to lignin binding can be modulated by using different enzymes of the same activity We used water:dioxane (1:9) to extract lignin from pretreated corn stover. Commercial cellulases were incubated with lignin and the unbound supernatants were evaluated for individual enzyme loss by SDS=PAGE and these were correlated with activity loss using various pNP-sugar substrates. Colorimetric assays for general glycosyl hydrolasemore » activities showed distinct differences in enzyme binding to lignin for each enzyme activity. Native systems demonstrated low binding of endo- and exo-cellulases, high binding of xylanase, and moderate ..beta..-glucosidase binding. Engineered cellulase mixtures exhibited low binding of exo-cellulases, very strong binding of endocellulases and ..beta..- glucosidase, and mixed binding of xylanase activity. The augmented cellulase had low binding of exocellulase, high binding of endocellulase and xylanase, and moderate binding of ..beta..-glucosidase activities. Bound and unbound activities were correlated with general molecular weight ranges of proteins as measured by loss of proteins bands in bound fractions on SDS-PAGE gels. Lignin-bound high molecular weight bands correlated with binding of ..beta..-glucosidase activity. While ..beta..-glucosidases demonstrated high binding in many cases, they have been shown to remain active. Bound low molecular weight bands correlated with xylanase activity binding. Contrary to other literature, exocellulase activity did not show strong lignin binding. The variation in enzyme activity binding between the three classes of cellulases preparations indicate that it is certainly possible to alter the binding of specific glycosyl hydrolase activities. It remains unclear whether loss of endocellulase activity to lignin binding is problematic for biomass conversion.« less

Functional and structural characterization of the pentapeptide insertion of Theileria annulata lactate dehydrogenase by site-directed mutagenesis, comparative modeling and molecular dynamics simulations.

PubMed

Erdemir, Aysegul; Mutlu, Ozal

2017-06-01

Lactate dehydrogenase (LDH) is an important metabolic enzyme in glycolysis and it has been considered as the main energy source in many organisms including apicomplexan parasites. Differences at the active site loop of the host and parasite LDH's makes this enzyme an attractive target for drug inhibitors. In this study, five amino acid insertions in the active site pocket of Theileria annulata LDH (TaLDH) were deleted by PCR-based site-directed mutagenesis, expression and activity analysis of mutant and wild type TaLDH enzymes were performed. Removal of the insertion at the active site loop caused production of an inactive enzyme. Furthermore, structures of wild and mutant enzymes were predicted by comparative modeling and the importance of the insertions at the active site loop were also assigned by molecular docking and dynamics simulations in order to evaluate essential role of this loop for the enzymatic activity. Pentapeptide insertion removal resulted in loss of LDH activity due to deletion of Trp96 and conformational change of Arg98 because of loop instability. Analysis of wild type and mutant enzymes with comparative molecular dynamics simulations showed that the fluctuations of the loop residues increase in mutant enzyme. Together with in silico studies, in vitro results revealed that active site loop has a vital role in the enzyme activity and our findings promise hope for the further drug design studies against theileriosis and other apicomplexan parasite diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Improved production of an enzyme that hydrolyses raw yam starch by Penicillium sp. S-22 using fed-batch fermentation.

PubMed

Sun, Hai-Yan; Ge, Xiang-Yang; Zhang, Wei-Guo

2006-11-01

A newly isolated strain, Penicillium sp. S-22, was used to produce an enzyme that hydrolyses raw yam starch [raw yam starch digesting enzyme (RYSDE)]. The enzyme activity and overall enzyme productivity were respectively 16 U/ml and 0.19 U/ml h in the batch culture. The enzyme activity increased to 85 U/ml by feeding of partially hydrolyzed raw yam starch. When a mixture containing partially hydrolyzed raw yam starch and peptone was fed by a pH-stat strategy, the enzyme activity reached 366 U/ml, 23-fold of that obtained in the batch culture, and the overall productivity reached 3.4 U/ml h, which was 18-fold of that in the batch culture.

Effect of monensin on the levels of tachykinins and their processing enzyme activity in rat dorsal root ganglia.

PubMed

Chikuma, Toshiyuki; Inomata, Yuji; Tsuchida, Ken; Hojo, Hiroshi; Kato, Takeshi

2002-06-28

Th effect of monensin, which inhibits trans-Golgi function, on the levels of tachykinins and their processing enzyme activity was examined in organ-cultured rat dorsal root ganglia (DRG). Using an enzyme immunoassay method, we measured neurokinin A and substance P immunoreactivity in the DRG cultured for 72 h with and without 0.1 microM monensin. Both tachykinins were reduced in the DRG treated with monensin. Treatment with monensin also reduced the activity of carboxypeptidase E, which is one of the proteolytic processing enzymes of neuropeptides. These data suggest that proteolytic processing enzymes may in part modulate the biological activity of neuropeptides within a trans-Golgi apparatus.

The effect of preparation, storage and shipping of dried blood spots on the activity of five lysosomal enzymes.

PubMed

Elbin, Carole S; Olivova, Petra; Marashio, Carla A; Cooper, Samantha K; Cullen, Emmaline; Keutzer, Joan M; Zhang, X Kate

2011-06-11

Fluorometric and tandem mass spectrometry assays can be used to measure lysosomal enzyme activities in dried blood spots (DBS). The effect of DBS preparation, storage and shipping was evaluated on the activities of acid α-glucosidase, acid α-galactosidase, acid β-glucocerebrosidase, acid sphingomyelinase, and galactocerebrosidase. Whole blood from normal donors was used to prepare DBS following Clinical and Laboratory Standards Institute guidelines and by several deviations. Some DBS were subjected to various treatments, storage and shipping conditions. The activity of 5 lysosomal enzymes (GAA, GLA, GBA, ASM, and GALC) was measured using tandem mass spectrometric and fluorometric (GAA only) assays with 2 distinct and commonly used synthetic substrates. Enzyme activities were strongly affected by the way DBS were prepared and stored. Exposure of DBS to elevated heat and humidity can destroy enzyme functions rapidly. DBS prepared from poorly mixed blood caused significant variation on enzyme activities. EDTA, but not heparin, as an anti-coagulant gave more precise results. The study confirmed the importance of proper and consistent DBS preparation and storage when screening for deficiencies of lysosomal enzymes. Copyright © 2011 Elsevier B.V. All rights reserved.

Arylsulfatase Activity in Salt Marsh Soils †

PubMed Central

Oshrain, R. L.; Wiebe, W. J.

1979-01-01

The presence of arylsulfatase(s) was confirmed in salt marsh soils. The temperatures of maximum activity and inactivation, the pH range over which the enzyme was active, and the Km values were similar to those of soil enzymes. Unlike soil arylsulfatases, however, the salt marsh enzymes do not appear to be repressed by sulfate. It is postulated that these enzymes may be necessary for the initiation of arylsulfate ester metabolism. PMID:16345425

Perfusion chromatography separation of the tomato fruit-specific pectin methylesterase from a semipurified commercial enzyme preparation.

PubMed

Savary, B J

2001-08-01

A rapid and simple method was developed, using perfusion chromatography media, to separate the fruit-specific pectin methylesterase (PME) isoform from the depolymerizing enzyme polygalacturonase (PG) and other contaminating pectinases present in a commercial tomato enzyme preparation. Pectinase activities were adsorbed onto a Poros HS (a strong cation exchanger) column in 20 M HEPES buffer at pH 7.5. The fruit-specific PME was eluted from the column with 80 mM NaCl, followed by a step to 300 mM NaCl to elute PG activity. Rechromatography of the PME activity peak with a linear gradient further resolved two PME isoenzymes and removed residual traces of PG activity. The PG activity peak was further treated with lectin affinity chromatography to provide purified PG enzyme, which was separated from a salt-dependent PME (tentatively identified as a "ubiquitous-type" isoform), and a pectin acetylesterase. The later enzyme has not been reported previously in tomato. This method provides monocomponent enzymes that will be useful for studying enzyme mechanisms and for modifying pectin structure and functional properties.

Celluloytic enzymes, nucleic acids encoding them and methods for making and using them

DOEpatents

Gray, Kevin A; Zhao, Lishan; Cayouette, Michelle H

2015-11-04

The invention is directed to polypeptides having any cellulolytic activity, e.g., a cellulase activity, e.g., endoglucanase, cellobiohydrolase, beta-glucosidase, xylanase, mannanse, .beta.-xylosidase, arabinofuranosidase, and/or oligomerase activity, including thermostable and thermotolerant activity, and polynucleotides encoding these enzymes, and making and using these polynucleotides and polypeptides. The polypeptides of the invention can be used in a variety of pharmaceutical, agricultural, food and feed processing and industrial contexts. The invention also provides compositions or products of manufacture comprising mixtures of enzymes comprising at least one enzyme of this invention.

Celluloytic enzymes, nucleic acids encoding them and methods for making and using them

DOEpatents

Gray, Kevin A.; Zhao, Lishan; Cayouette, Michelle H.

2015-09-08

The invention is directed to polypeptides having any cellulolytic activity, e.g., a cellulase activity, e.g., endoglucanase, cellobiohydrolase, beta-glucosidase, xylanase, mannanse, .beta.-xylosidase, arabinofuranosidase, and/or oligomerase activity, including thermostable and thermotolerant activity, and polynucleotides encoding these enzymes, and making and using these polynucleotides and polypeptides. The polypeptides of the invention can be used in a variety of pharmaceutical, agricultural, food and feed processing and industrial contexts. The invention also provides compositions or products of manufacture comprising mixtures of enzymes comprising at least one enzyme of this invention.

L-asparaginase activity in Aeromonas sp. isolated from freshwater mussel.

PubMed

Pattnaik, S; Kabi, R; Janaki Ram, K; Bhanot, K K

2000-11-01

Aeromonas sp. from Lamellidens marginalis produced L-asparaginase when grown at 37 degrees C. The optimum enzyme activity was at pH 9 when temperature was 45 degrees C. Half-life of partially purified enzyme at 50 degrees C and 55 degrees C was 35 and 20 min, respectively. Activation and deactivation energies of partially purified enzyme were 17.48 and 24.86 kcal mol-1 respectively. The enzyme exhibited a Km (L-asparagine) value of 4.9 x 10(-6) mol l-1 and a Vmax of 9.803 IU ml-1. Three metal ions inhibited the enzyme activity at 10-20 mumol l-1 concentrations. Catalytic activity was also inhibited by EDTA, iodoacetic acid, parachloromercuribenzoic acid and phenylmethylsulphonyl fluoride at 0.1 mumol l-1.

Sodium and Potassium Ions in Proteins and Enzyme Catalysis.

PubMed

Vašák, Milan; Schnabl, Joachim

2016-01-01

The group I alkali metal ions Na(+) and K(+) are ubiquitous components of biological fluids that surround biological macromolecules. They play important roles other than being nonspecific ionic buffering agents or mediators of solute exchange and transport. Molecular evolution and regulated high intracellular and extracellular M(+) concentrations led to incorporation of selective Na(+) and K(+) binding sites into enzymes to stabilize catalytic intermediates or to provide optimal positioning of substrates. The mechanism of M(+) activation, as derived from kinetic studies along with structural analysis, has led to the classification of cofactor-like (type I) or allosteric effector (type II) activated enzymes. In the type I mechanism substrate anchoring to the enzyme active site is mediated by M(+), often acting in tandem with a divalent cation like Mg(2+), Mn(2+) or Zn(2+). In the allosteric type II mechanism, M(+) binding enhances enzyme activity through conformational transitions triggered upon binding to a distant site. In this chapter, following the discussion of the coordination chemistry of Na(+) and K(+) ions and the structural features responsible for the metal binding site selectivity in M(+)-activated enzymes, well-defined examples of M(+)-activated enzymes are used to illustrate the structural basis for type I and type II activation by Na(+) and K(+).

Standardized Assay Medium To Measure Lactococcus lactis Enzyme Activities while Mimicking Intracellular Conditions

PubMed Central

Goel, Anisha; Santos, Filipe; de Vos, Willem M.; Teusink, Bas

2012-01-01

Knowledge of how the activity of enzymes is affected under in vivo conditions is essential for analyzing their regulation and constructing models that yield an integrated understanding of cell behavior. Current kinetic parameters for Lactococcus lactis are scattered through different studies and performed under different assay conditions. Furthermore, assay conditions often diverge from conditions prevailing in the intracellular environment. To establish uniform assay conditions that resemble intracellular conditions, we analyzed the intracellular composition of anaerobic glucose-limited chemostat cultures of L. lactis subsp. cremoris MG 1363. Based on this, we designed a new assay medium for enzyme activity measurements of growing cells of L. lactis, mimicking as closely as practically possible its intracellular environment. Procedures were optimized to be carried out in 96-well plates, and the reproducibility and dynamic range were checked for all enzyme activity measurements. The effects of freezing and the carryover of ammonium sulfate from the addition of coupling enzymes were also established. Activities of all 10 glycolytic and 4 fermentative enzymes were measured. Remarkably, most in vivo-like activities were lower than previously published data. Yet, the ratios of Vmax over measured in vivo fluxes were above 1. With this work, we have developed and extensively validated standard protocols for enzyme activity measurements for L. lactis. PMID:22020503

Positron emitter labeled enzyme inhibitors

DOEpatents

Fowler, J.S.; MacGregor, R.R.; Wolf, A.P.

1987-05-22

This invention involved a new strategy for imaging and mapping enzyme activity in the living human and animal body using positron emitter-labeled suicide enzyme inactivators or inhibitors which become covalently bound to the enzyme as a result of enzymatic catalysis. Two such suicide in activators for monoamine oxidase have been labeled with carbon-11 and used to map the enzyme subtypes in the living human and animal body using PET. By using positron emission tomography to image the distribution of radioactivity produced by the body penetrating radiation emitted by carbon-11, a map of functionally active monoamine oxidase activity is obtained. Clorgyline and L-deprenyl are suicide enzyme inhibitors and irreversibly inhibit monoamine oxidase. When these inhibitors are labeled with carbon-11 they provide selective probes for monoamine oxidase localization and reactivity in vivo using positron emission tomography. 2 figs.

Positron emitter labeled enzyme inhibitors

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

Fowler, J.S.; MacGregor, R.R.; Wolf, A.P.

This invention involved a new strategy for imaging and mapping enzyme activity in the living human and animal body using positron emitter-labeled suicide enzyme inactivators or inhibitors which become covalently bound to the enzyme as a result of enzymatic catalysis. Two such suicide in activators for monoamine oxidase have been labeled with carbon-11 and used to map the enzyme subtypes in the living human and animal body using PET. By using positron emission tomography to image the distribution of radioactivity produced by the body penetrating radiation emitted by carbon-11, a map of functionally active monoamine oxidase activity is obtained. Clorgylinemore » and L-deprenyl are suicide enzyme inhibitors and irreversibly inhibit monoamine oxidase. When these inhibitors are labeled with carbon-11 they provide selective probes for monoamine oxidase localization and reactivity in vivo using positron emission tomography. 2 figs.« less

Multifunctional enzymes from reduced genomes - model proteins for simple primordial metabolism?

PubMed

Seelig, Burckhard

2017-08-01

Billions of years of evolution have yielded today's complex metabolic networks driven by efficient and highly specialized enzymes. In contrast, the metabolism of the earliest cellular life forms was likely much simpler with only a few enzymes of comparatively low activity. It has been speculated that these early enzymes had low specificities and in turn were able to perform multiple functions. In this issue of Molecular Microbiology, Ferla et al. describe examples of enzymes that catalyze chemically distinct reactions while using the same active site. Most importantly, the authors demonstrated that the comparatively weak activities of these multifunctional enzymes are each physiologically relevant. These findings contrast with simply promiscuous enzyme activities, which have been described numerous times but are not physiologically relevant. Ferla et al. elegantly combined initial bioinformatics searches for enzyme candidates with sound kinetic measurements, evolutionary considerations and even structural discussions. The phenomenon of multifunctionality appears to be a mechanism for bacteria with reduced genomes to compensate for their lack of certain enzymes. In the broader context of evolution, these organisms could be considered living model systems to study features of long-extinct early cellular life. © 2017 John Wiley & Sons Ltd.

Immobilization of fungal beta-glucosidase on silica gel and kaolin carriers.

PubMed

Karagulyan, Hakob K; Gasparyan, Vardan K; Decker, Stephen R

2008-03-01

Beta-glucosidase is a key enzyme in the hydrolysis of cellulose for producing feedstock glucose for various industrial processes. Reuse of enzyme through immobilization can significantly improve the economic characteristics of the process. Immobilization of the fungal beta-glucosidase by covalent binding and physical adsorption on silica gel and kaolin was conducted for consequent application of these procedures in large-scale industrial processes. Different immobilization parameters (incubation time, ionic strength, pH, enzyme/support ratio, glutaric aldehyde concentration, etc.) were evaluated for their effect on the thermal stability of the immobilized enzyme. It was shown that the immobilized enzyme activity is stable at 50 degrees C over 8 days. It has also been shown that in the case of immobilization on kaolin, approximately 95% of the initial enzyme was immobilized onto support, and loss of activity was not observed. However, covalent binding of the enzyme to silica gel brings significant loss of enzyme activity, and only 35% of activity was preserved. In the case of physical adsorption on kaolin, gradual desorption of enzyme takes place. To prevent this process, we have carried out chemical modification of the protein. As a result, after repeated washings, enzyme desorption from kaolin has been reduced from 75 to 20-25% loss.

The Holo-Transcriptome of the Zoantharian Protopalythoa variabilis (Cnidaria: Anthozoa): A Plentiful Source of Enzymes for Potential Application in Green Chemistry, Industrial and Pharmaceutical Biotechnology.

PubMed

R L Morlighem, Jean-Étienne; Huang, Chen; Liao, Qiwen; Braga Gomes, Paula; Daniel Pérez, Carlos; de Brandão Prieto-da-Silva, Álvaro Rossan; Ming-Yuen Lee, Simon; Rádis-Baptista, Gandhi

2018-06-13

Marine invertebrates, such as sponges, tunicates and cnidarians (zoantharians and scleractinian corals), form functional assemblages, known as holobionts, with numerous microbes. This type of species-specific symbiotic association can be a repository of myriad valuable low molecular weight organic compounds, bioactive peptides and enzymes. The zoantharian Protopalythoa variabilis (Cnidaria: Anthozoa) is one such example of a marine holobiont that inhabits the coastal reefs of the tropical Atlantic coast and is an interesting source of secondary metabolites and biologically active polypeptides. In the present study, we analyzed the entire holo-transcriptome of P. variabilis , looking for enzyme precursors expressed in the zoantharian-microbiota assemblage that are potentially useful as industrial biocatalysts and biopharmaceuticals. In addition to hundreds of predicted enzymes that fit into the classes of hydrolases, oxidoreductases and transferases that were found, novel enzyme precursors with multiple activities in single structures and enzymes with incomplete Enzyme Commission numbers were revealed. Our results indicated the predictive expression of thirteen multifunctional enzymes and 694 enzyme sequences with partially characterized activities, distributed in 23 sub-subclasses. These predicted enzyme structures and activities can prospectively be harnessed for applications in diverse areas of industrial and pharmaceutical biotechnology.

Aβ-degrading enzymes: potential for treatment of Alzheimer disease.

PubMed

Miners, James Scott; Barua, Neil; Kehoe, Patrick Gavin; Gill, Steven; Love, Seth

2011-11-01

There is increasing evidence that deficient clearance of β-amyloid (Aβ) contributes to its accumulation in late-onset Alzheimer disease (AD). Several Aβ-degrading enzymes, including neprilysin (NEP), insulin-degrading enzyme, and endothelin-converting enzyme reduce Aβ levels and protect against cognitive impairment in mouse models of AD. The activity of several Aβ-degrading enzymes rises with age and increases still further in AD, perhaps as a physiological response to minimize the buildup of Aβ. The age- and disease-related changes in expression of more recently recognized Aβ-degrading enzymes (e.g. NEP-2 and cathepsin B) remain to be investigated, and there is strong evidence that reduced NEP activity contributes to the development of cerebral amyloid angiopathy. Regardless of the role of Aβ-degrading enzymes in the development of AD, experimental data indicate that increasing the activity of these enzymes (NEP in particular) has therapeutic potential in AD, although targeting their delivery to the brain remains a major challenge. The most promising current approaches include the peripheral administration of agents that enhance the activity of Aβ-degrading enzymes and the direct intracerebral delivery of NEP by convection-enhanced delivery. In the longer term, genetic approaches to increasing the intracerebral expression of NEP or other Aβ-degrading enzymes may offer advantages.

[Importance of the 11β-hydroxysteroid dehydrogenase enzyme in clinical disorders].

PubMed

Feldman, Karolina; Likó, István; Nagy, Zsolt; Szappanos, Agnes; Grolmusz, Vince Kornél; Tóth, Miklós; Rácz, Károly; Patócs, Attila

2013-02-24

Glucocorticoids play an important role in the regulation of carbohydrate and amino acid metabolism, they modulate the function of the immune system, and contribute to stress response. Increased and decreased production of glucocorticoids causes specific diseases. In addition to systemic hypo- or hypercortisolism, alteration of local synthesis and metabolism of cortisol may result in tissue-specific hypo- or hypercortisolism. One of the key enzymes participating in the local synthesis and metabolism of cortisol is the 11β-hydroxysteroid dehydrogenase enzyme. Two isoforms, type 1 and type 2 enzymes are located in the endoplasmic reticulum and catalyze the interconversion of hormonally active cortisol and inactive cortisone. The type 1 enzyme mainly works as an activator, and it is responsible for the generation of cortisol from cortisone in liver, adipose tissue, brain and bone. The gene encoding this enzyme is located on chromosome 1. The authors review the physiological and pathophysiological processes related to the function of the type 1 11β-hydroxysteroid dehydrogenase enzyme. They summarize the potential significance of polymorphic variants of the enzyme in clinical diseases as well as knowledge related to inhibitors of enzyme activity. Although further studies are still needed, inhibition of the enzyme activity may prove to be an effective tool for the treatment of several diseases such as obesity, osteoporosis and type 2 diabetes.

Immobilization of Fungal β-Glucosidase on Silica Gel and Kaolin Carriers

NASA Astrophysics Data System (ADS)

Karagulyan, Hakob K.; Gasparyan, Vardan K.; Decker, Stephen R.

β-Glucosidase is a key enzyme in the hydrolysis of cellulose for producing feedstock glucose for various industrial processes. Reuse of enzyme through immobilization can significantly improve the economic characteristics of the process. Immobilization of the fungal β-glucosidase by covalent binding and physical adsorption on silica gel and kaolin was conducted for consequent application of these procedures in large-scale industrial processes. Different immobilization parameters (incubation time, ionic strength, pH, enzyme/support ratio, glutaric aldehyde concentration, etc.) were evaluated for their effect on the thermal stability of the immobilized enzyme. It was shown that the immobilized enzyme activity is stable at 50 °C over 8 days. It has also been shown that in the case of immobilization on kaolin, approximately 95% of the initial enzyme was immobilized onto support, and loss of activity was not observed. However, covalent binding of the enzyme to silica gel brings significant loss of enzyme activity, and only 35% of activity was preserved. In the case of physical adsorption on kaolin, gradual desorption of enzyme takes place. To prevent this process, we have carried out chemical modification of the protein. As a result, after repeated washings, enzyme desorption from kaolin has been reduced from 75 to 20-25% loss.

Stability of the anti-oxidative enzymes in aqueous and detergent solution.

PubMed

Mailer, K; Del Maestro, R F

1991-09-18

Activities of the anti-oxidative enzymes, superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase were studied in rat tissues to determine the ability of detergents both to solubilize the enzymes and also to stabilize enzyme activity. Rat brain, heart and liver were homogenized in 0.1M KCl, 0.1% sodium dodecyl sulfate, 0.1% lubrol, or 0.1% cetyl-trimethylammonium bromide. In general lubrol was more effective than the other solutions in solubilizing GPx and catalase. Lubrol and 0.1M KCl were equally effective in solubilizing SOD. The highest enzyme activities were (1) SOD: 2484 ng/mg (brain), 2501 ng/mg (heart), and 5586 ng/mg (liver); (2) GPx: 224 mU/mg (brain), 1870 mU/mg (heart), and 7332 mU/mg (liver); (3) catalase: 2.8 mU/mg (brain), 10.6 mU/mg (heart), and 309 mU/mg (liver). While cetyl trimethylammonium bromide is marginally better than sodium dodecyl sulfate in solubilizing active enzyme, neither ionic detergent has any advantage over lubrol or 0.1M KCl. For catalase and GPx, enzyme activity loss with time is biphasic. After initial, rapid activity loss (1-5 days for GPx and 7-10 days for catalase) the differences noted among the homogenizing solutions disappear and very little if any activity loss is noted over the next 2-3 weeks. For catalase and GPx, only baseline enzyme activity from t = 0-3 weeks is found in the most chaotropic solution, 0.1% sodium dodecyl sulfate while biphasic activity loss is most pronounced in 0.1% lubrol. These results may indicate active GPx and catalase species stabilized by a lipid-like environment. Correlating in vitro catalase or GPx measurements with in vivo anti-oxidative protection may underestimate tissue defences.

Partial purification and some properties of a latent CO2 reductase from green potato tuber chloroplasts.

PubMed

Arora, S; Ramaswamy, N K; Nair, P M

1985-12-16

We have partially purified the CO2 reductase, present in green potato tuber chloroplasts, as a latent form. Illumination of the chloroplasts in the absence of substrate, bicarbonate, activated the enzyme, which could then be obtained in soluble forms. Purification of the enzyme was achieved by (NH4)2SO4 fractionation (0-30%) and adsorption and elution from a DEAE-Sephadex A-50 column. The final preparation showed 15-fold purification and 50% recovery of the activity. The pH optimum for CO2 reductase was 8.0. Hepes and Tricine buffers showed maximum activity whereas Tris/phosphate or borate failed to show any activity. The enzyme reaction was sensitive to the presence of metal ions like Fe3+, Hg2+, Cu2+, Mo6+ and Zn2+, however, a threefold activation was observed with Fe2+. The metal requirement for CO2 reductase was evident from the observed inhibition by metal chelators like o-phenanthroline, alpha, alpha'-dipyridyl, bathocuproine, 8-hydroxyquinoline etc. Out of these o-phenanthroline was the strongest inhibitor and its concentration for 50% inhibition was 40 microM. The presence of Fe2+ ions in the reaction mixture protected the enzyme from heat denaturation upto 50 degrees C. Maximum enzyme activity was observed at 15 degrees C. The enzyme activity showed a 30-s lag period and the maximum was reached in 90 s. Supplementation of sodium dithionite in the reaction activated enzyme activity threefold, suggesting involvement of dithiol groups in the catalytic activity. There was strong inhibition by -SH inhibitors like 5,5'-dithiobis(2-nitrobenzoic acid) and N-ethylmaleimide and -SH reagents like dithiothreitol, 2-mercaptoethanol and cysteine. Various nucleotide coenzyme tried inhibited the enzyme strongly.

Enzymatic hydrolysis of steam-pretreated lignocellulosic materials with Trichoderma atroviride enzymes produced in-house

PubMed Central

Kovacs, Krisztina; Macrelli, Stefano; Szakacs, George; Zacchi, Guido

2009-01-01

Background Improvement of the process of cellulase production and development of more efficient lignocellulose-degrading enzymes are necessary in order to reduce the cost of enzymes required in the biomass-to-bioethanol process. Results Lignocellulolytic enzyme complexes were produced by the mutant Trichoderma atroviride TUB F-1663 on three different steam-pretreated lignocellulosic substrates, namely spruce, wheat straw and sugarcane bagasse. Filter paper activities of the enzymes produced on the three materials were very similar, while β-glucosidase and hemicellulase activities were more dependent on the nature of the substrate. Hydrolysis of the enzyme preparations investigated produced similar glucose yields. However, the enzymes produced in-house proved to degrade the xylan and the xylose oligomers less efficiently than a commercial mixture of cellulase and β-glucosidase. Furthermore, accumulation of xylose oligomers was observed when the TUB F-1663 supernatants were applied to xylan-containing substrates, probably due to the low β-xylosidase activity of the enzymes. The efficiency of the enzymes produced in-house was enhanced by supplementation with extra commercial β-glucosidase and β-xylosidase. When the hydrolytic capacities of various mixtures of a commercial cellulase and a T. atroviride supernatant produced in the lab were investigated at the same enzyme loading, the glucose yield appeared to be correlated with the β-glucosidase activity, while the xylose yield seemed to be correlated with the β-xylosidase level in the mixtures. Conclusion Enzyme supernatants produced by the mutant T. atroviride TUB F-1663 on various pretreated lignocellulosic substrates have good filter paper activity values combined with high levels of β-glucosidase activities, leading to cellulose conversion in the enzymatic hydrolysis that is as efficient as with a commercial cellulase mixture. On the other hand, in order to achieve good xylan conversion, the supernatants produced by the mutant have to be supplemented with additional β-xylosidase activity. PMID:19580644

Assembling high activity phosphotriesterase composites using hybrid nanoparticle peptide-DNA scaffolded architectures

NASA Astrophysics Data System (ADS)

Breger, Joyce C.; Buckhout-White, Susan; Walper, Scott A.; Oh, Eunkeu; Susumu, Kimihiro; Ancona, Mario G.; Medintz, Igor L.

2017-06-01

Nanoparticle (NP) display potentially offers a new way to both stabilize and, in many cases, enhance enzyme activity over that seen for native protein in solution. However, the large, globular and sometimes multimeric nature of many enzymes limits their ability to attach directly to the surface of NPs, especially when the latter are colloidally stabilized with bulky PEGylated ligands. Engineering extended protein linkers into the enzymes to achieve direct attachment through the PEG surface often detrimentally alters the enzymes catalytic ability. Here, we demonstrate an alternate, hybrid biomaterials-based approach to achieving directed enzyme assembly on PEGylated NPs. We self-assemble a unique architecture consisting of a central semiconductor quantum dot (QD) scaffold displaying controlled ratios of extended peptide-DNA linkers which penetrate through the PEG surface to directly couple enzymes to the QD surface. As a test case, we utilize phosphotriesterase (PTE), an enzyme of bio-defense interest due to its ability to hydrolyze organophosphate nerve agents. Moreover, this unique approach still allows PTE to maintain enhanced activity while also suggesting the ability of DNA to enhance enzyme activity in and of itself.

Mineralogical impact on long-term patterns of soil nitrogen and phosphorus enzyme activities

NASA Astrophysics Data System (ADS)

Mikutta, Robert; Turner, Stephanie; Meyer-Stüve, Sandra; Guggenberger, Georg; Dohrmann, Reiner; Schippers, Axel

2014-05-01

Soil chronosequences provide a unique opportunity to study microbial activity over time in mineralogical diverse soils of different ages. The main objective of this study was to test the effect of mineralogical properties, nutrient and organic matter availability over whole soil pro-files on the abundance and activity of the microbial communities. We focused on microbio-logical processes involved in nitrogen and phosphorus cycling at the 120,000-year Franz Josef soil chronosequence. Microbial abundances (microbial biomass and total cell counts) and enzyme activities (protease, urease, aminopeptidase, and phosphatase) were determined and related to nutrient contents and mineralogical soil properties. Both, microbial abundances and enzyme activities decreased with soil depth at all sites. In the organic layers, microbial biomass and the activities of N-hydrolyzing enzymes showed their maximum at the intermediate-aged sites, corresponding to a high aboveground biomass. In contrast, the phosphatase activity increased with site age. The activities of N-hydrolyzing enzymes were positively correlated with total carbon and nitrogen contents, whereas the phosphatase activity was negatively correlated with the phosphorus content. In the mineral soil, the enzyme activities were generally low, thus reflecting the presence of strongly sorbing minerals. Sub-strate-normalized enzyme activities correlated negatively to clay content as well as poorly crystalline Al and Fe oxyhydroxides, supporting the view that the evolution of reactive sec-ondary mineral phases alters the activity of the microbial communities by constraining sub-strate availability. Our data suggest a strong mineralogical influence on nutrient cycling par-ticularly in subsoil environments.

A Modular Approach for Interlocking Enzymes in Whatman Paper.

PubMed

Riccardi, Caterina; Kumar, Challa; Kasi, Rajeswari; McCormick, Shelby

2018-06-13

We report a potentially universal approach for enzyme attachment to cellulose that significantly enhances enzyme stability while retaining high activity, and involves no chemical functionalization of cellulose. In our design, bovine serum albumin (BSA) was interlocked in cellulose to form a protein-friendly surface (named BSA-Paper), while also providing COOH and NH2 groups for subsequent attachment of enzymes. The desired enzyme is then mixed with additional BSA and interlocked on BSA-Paper. The 2nd layer dilutes and crosslinks the enzyme for improved stability. Laccase was tested as a model enzyme for interlocking on BSA-Paper, and was found to retain over 100% activity and was 240 times more stable at 25 °C (half life = 180 d) than laccase. This new approach was also tested with a few other enzymes with encouraging results, thus providing a potentially universal method for stabilization of enzymes on cellulose with retention of high activities. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Lipophilic flavonoids from Orthosiphon spicatus prevent oxidative inactivation of 15-lipoxygenase.

PubMed

Lyckander, I M; Malterud, K E

1996-04-01

15-Lipoxygenase from soybeans is inactivated by bubbling air through enzyme solutions. This inactivation is prevented by flavonoids from the East Asian medicinal plant Orthosiphon spicatus, previously found to be inhibitors of the enzyme. 5,7,4'-Trimethylapigenin, eupatorin and 5,7,3',4'-tetramethylluteolin show the strongest enzyme-stabilizing effects, decreasing loss of activity by 50% at concentrations of 2.0 +/- 0.04, 2.4 +/- 0.3 and 4.3 +/- 1.1 microM, respectively. There is no significant correlation between enzyme-inhibiting and enzyme-stabilizing effect. The Orthosiphon flavonoids show radical-scavenging activity towards the diphenylpicrylhydrazyl radical. This is correlated to their enzyme-stabilizing effect, but not to their inhibitory activity towards 15-lipoxygenase. When the enzyme is inactivated by air bubbling, a loss of sulfhydryl groups is observed. Sinensetin, a poor stabilizer of the enzyme, shows less efficiency in protecting sulfhydryl groups than tetramethylscutellarein, which stabilizes the enzyme more efficiently. Thus, oxidation of sulfhydryl groups may contribute to the observed air-induced inactivation of 15-lipoxygenase.

Technological advances and applications of hydrolytic enzymes for valorization of lignocellulosic biomass.

PubMed

Manisha; Yadav, Sudesh Kumar

2017-12-01

Hydrolytic enzymes are indispensable tools in the production of various foodstuffs, drugs, and consumables owing to their applications in almost every industrial process nowadays. One of the foremost areas of interest involving the use of hydrolytic enzymes is in the transformation of lignocellulosic biomass into value added products. However, limitations of the processes due to inadequate enzyme activity and stability with a narrow range of pH and temperature optima often limit their effective usage. The innovative technologies, involving manipulation of enzyme activity and stability through mutagenesis, genetic engineering and metagenomics lead to a major leap in all the fields using hydrolytic enzymes. This article provides recent advancement towards the isolation and use of microbes for lignocellulosic biomass utilisation, microbes producing the hydrolytic enzymes, the modern age technologies used to manipulate and enhance the hydrolytic enzyme activity and the applications of such enzymes in value added products development from lignocellulosic biomass. Copyright © 2017 Elsevier Ltd. All rights reserved.

An improved 96-well turbidity assay for T4 lysozyme activity

DTIC Science & Technology

2015-05-13

enzyme present in the reaction, resulting in a measure of activity in Unitsmg1. 10. To improve the reliability of the activity values, perform the...quantification of lysozyme activity with significantly lower enzyme concentrations, and the signal intensity can be enhanced by using greater amounts of... enzyme at the expense of a shorter linear reaction time. Several parameters of the assay are critical for obtaining reproducible activity

Antioxidative capacity and enzyme activity in Haematococcus pluvialis cells exposed to superoxide free radicals

NASA Astrophysics Data System (ADS)

Liu, Jianguo; Zhang, Xiaoli; Sun, Yanhong; Lin, Wei

2010-01-01

The antioxidative capacity of astaxanthin and enzyme activity of reactive oxygen eliminating enzymes such as superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) and ascorbate peroxidase (APX) were studied in three cell types of Haematococcus pluvialis exposed to high concentrations of a superoxide anion radical (O{2/-}). The results show that defensive enzymes and astaxanthin-related mechanisms were both active in H. pluvialis during exposure to reactive oxygen species (ROS) such as O{2/-}. Astaxanthin reacted with ROS much faster than did the protective enzymes, and had the strongest antioxidative capacity to protect against lipid peroxidation. The defensive mechanisms varied significantly between the three cell types and were related to the level of astaxanthin that had accumulated in those cells. Astaxanthin-enriched red cells had the strongest antioxidative capacity, followed by brown cells, and astaxanthin-deficient green cells. Although there was no significant increase in expression of protective enzymes, the malondialdehyde (MDA) content in red cells was sustained at a low level because of the antioxidative effect of astaxanthin, which quenched O{2/-} before the protective enzymes could act. In green cells, astaxanthin is very low or absent; therefore, scavenging of ROS is inevitably reliant on antioxidative enzymes. Accordingly, in green cells, these enzymes play the leading role in scavenging ROS, and the expression of these enzymes is rapidly increased to reduce excessive ROS. However, because ROS were constantly increased in this study, the enhance enzyme activity in the green cells was not able to repair the ROS damage, leading to elevated MDA content. Of the four defensive enzymes measured in astaxanthin-deficient green cells, SOD eliminates O{2/-}, POD eliminates H2O2, which is a by-product of SOD activity, and APX and CAT are then initiated to scavenge excessive ROS.

Homology to peptide pattern for annotation of carbohydrate-active enzymes and prediction of function.

PubMed

Busk, P K; Pilgaard, B; Lezyk, M J; Meyer, A S; Lange, L

2017-04-12

Carbohydrate-active enzymes are found in all organisms and participate in key biological processes. These enzymes are classified in 274 families in the CAZy database but the sequence diversity within each family makes it a major task to identify new family members and to provide basis for prediction of enzyme function. A fast and reliable method for de novo annotation of genes encoding carbohydrate-active enzymes is to identify conserved peptides in the curated enzyme families followed by matching of the conserved peptides to the sequence of interest as demonstrated for the glycosyl hydrolase and the lytic polysaccharide monooxygenase families. This approach not only assigns the enzymes to families but also provides functional prediction of the enzymes with high accuracy. We identified conserved peptides for all enzyme families in the CAZy database with Peptide Pattern Recognition. The conserved peptides were matched to protein sequence for de novo annotation and functional prediction of carbohydrate-active enzymes with the Hotpep method. Annotation of protein sequences from 12 bacterial and 16 fungal genomes to families with Hotpep had an accuracy of 0.84 (measured as F1-score) compared to semiautomatic annotation by the CAZy database whereas the dbCAN HMM-based method had an accuracy of 0.77 with optimized parameters. Furthermore, Hotpep provided a functional prediction with 86% accuracy for the annotated genes. Hotpep is available as a stand-alone application for MS Windows. Hotpep is a state-of-the-art method for automatic annotation and functional prediction of carbohydrate-active enzymes.

Cellulolytic enzymes, nucleic acids encoding them and methods for making and using them

DOEpatents

Gray, Kevin A [San Diego, CA; Zhao, Lishan [Emeryville, CA; Cayouette, Michelle H [San Diego, CA

2012-01-24

The invention provides polypeptides having any cellulolytic activity, e.g., a cellulase activity, a endoglucanase, a cellobiohydrolase, a beta-glucosidase, a xylanase, a mannanse, a .beta.-xylosidase, an arabinofuranosidase, and/or an oligomerase activity, polynucleotides encoding these polypeptides, and methods of making and using these polynucleotides and polypeptides. In one aspect, the invention is directed to polypeptides having any cellulolytic activity, e.g., a cellulase activity, e.g., endoglucanase, cellobiohydrolase, beta-glucosidase, xylanase, mannanse, .beta.-xylosidase, arabinofuranosidase, and/or oligomerase activity, including thermostable and thermotolerant activity, and polynucleotides encoding these enzymes, and making and using these polynucleotides and polypeptides. In one aspect, the invention provides polypeptides having an oligomerase activity, e.g., enzymes that convert recalcitrant soluble oligomers to fermentable sugars in the saccharification of biomass. The polypeptides of the invention can be used in a variety of pharmaceutical, agricultural, food and feed processing and industrial contexts. The invention also provides compositions or products of manufacture comprising mixtures of enzymes comprising at least one enzyme of this invention.

Effects of culture conditions on monosaccharide composition of Ganoderma lucidum exopolysaccharide and on activities of related enzymes.

PubMed

Peng, Lin; Qiao, Shuangkui; Xu, Zhenghong; Guan, Feng; Ding, Zhongyang; Gu, Zhenghua; Zhang, Liang; Shi, Guiyang

2015-11-20

We investigated the relationship between monosaccharide composition of Ganoderma lucidum exopolysaccharide (EPS) and activities of EPS synthesis enzymes under various culture temperatures and initial pH values. The mole percentages of three major EPS monosaccharides, glucose, galactose and mannose, varied depending on culture conditions and the resulting EPS displayed differing anti-tumor activities. In nine tested enzymes, higher enzyme activities were correlated with higher temperature and lower initial pH. Altered mole percentages of galactose and mannose under various culture conditions were associated with activities of α-phosphoglucomutase (PGM) and phosphoglucose isomerase (PGI), respectively, and that of mannose was also associated with phosphomannose isomerase (PMI) activity only under various pH. Our findings suggest that mole percentages of G. lucidum EPS monosaccharides can be manipulated by changes of culture conditions that affect enzyme activities, and that novel fermentation strategies based on this approach may enhance production and biological activity of EPS. Copyright © 2015 Elsevier Ltd. All rights reserved.

Operational Solution to the Nonlinear Klein-Gordon Equation

NASA Astrophysics Data System (ADS)

Bengochea, G.; Verde-Star, L.; Ortigueira, M.

2018-05-01

We obtain solutions of the nonlinear Klein-Gordon equation using a novel operational method combined with the Adomian polynomial expansion of nonlinear functions. Our operational method does not use any integral transforms nor integration processes. We illustrate the application of our method by solving several examples and present numerical results that show the accuracy of the truncated series approximations to the solutions. Supported by Grant SEP-CONACYT 220603, the first author was supported by SEP-PRODEP through the project UAM-PTC-630, the third author was supported by Portuguese National Funds through the FCT Foundation for Science and Technology under the project PEst-UID/EEA/00066/2013

Using soil enzymes to explain observed differences in the response of soil decomposition to nitrogen fertilization

NASA Astrophysics Data System (ADS)

Stone, M.; Weiss, M.; Goodale, C. L.

2010-12-01

Soil microbes produce extracellular enzymes that degrade a variety of carbon-rich polymers contained within soil organic matter (SOM). These enzymes are key regulators of the terrestrial carbon cycle. However, basic information about the kinetics of extracellular enzymes and key environmental variables that regulate their catalytic ability is lacking. This study aims to clarify the mechanisms by which microbial carbon-degrading enzymes drive different responses to nitrogen (N) fertilization in soil decomposition at two sites with long-term N fertilization experiments, the Bear Brook (BB) forest in Maine and Fernow Forest (FF) in West Virginia. We examined a suite of cellulolytic and lignolytic enzymes that break down common SOM constituents. We hypothesized that enzymes derived from the site with a higher mean annual temperature (FF) would be more heat-tolerant, and retain their catalytic efficiency (Km) as temperature rises, relative to enzymes from the colder environment (BB). We further hypothesized that cellulolytic enzyme activity would be unaffected by N, while oxidative enzyme activity would be suppressed in N-fertilized soils. To test these hypotheses and examine the interactive effects of temperature and N, we measured enzyme activity in unfertilized and N-fertilized soils under a range of laboratory temperature manipulations. Preliminary results show a significant decrease in cellulolytic enzyme efficiency with temperature at the colder site (BB), as well as a significant increase in efficiency due to N-fertilization for two cellulolytic enzymes. Oxidative enzyme activity shows a marginally significant reduction due to N-fertilization at BB. These results suggest that soil warming may produce a negative feedback on carbon turnover in certain climates, while N-fertilization may alter the relative decomposition rates of different soil organic matter constituents. FF activity will be analyzed in a similar manner and the two sites will be compared in order to fully assess our hypotheses.

The allosteric activator ATP induces a substrate-dependent alteration of the quaternary structure of a mutant aspartate transcarbamoylase impaired in active site closure.

PubMed Central

Baker, D. P.; Fetler, L.; Vachette, P.; Kantrowitz, E. R.

1996-01-01

Aspartate transcarbamoylase from Escherichia coli shows homotropic cooperativity for aspartate as well as heterotropic regulation by nucleotides. Structurally, it consists of two trimeric catalytic subunits and three dimeric regulatory subunits, each chain being comprised of two domains. Glu-50 and Ser-171 are involved in stabilizing the closed conformation of the catalytic chain. Replacement of Glu-50 or Ser-171 by Ala in the holoenzyme has been shown previously to result in marked decreases in the maximal observed specific activity, homotropic cooperativity, and affinity for aspartate (Dembowski NJ, Newton CJ, Kantrowitz ER, 1990, Biochemistry 29:3716-3723; Newton CJ, Kantrowitz ER, 1990, Biochemistry 29:1444-1451). We have constructed a double mutant enzyme combining both mutations. The resulting Glu-50/ser-171-->Ala enzyme is 9-fold less active than the Ser-171-->Ala enzyme, 69-fold less active than the Glu-50-->Ala enzyme, and shows 1.3-fold and 1.6-fold increases in the [S]0.5Asp as compared to the Ser-171-->Ala and Glu-50-->Ala enzymes, respectively. However, the double mutant enzyme exhibits some enhancement of homotropic cooperativity with respect to aspartate, relative to the single mutant enzymes. At subsaturating concentrations of aspartate, the Glu-50/Ser-171 -->Ala enzyme is activated less by ATP than either the Glu-50-->Ala or Ser-171-->Ala enzyme, whereas CTP inhibition is intermediate between that of the two single mutants. As opposed to the wild-type enzyme, the Glu-50/Ser-171 -->Ala enzyme is activated by ATP and inhibited by CTP at saturating concentrations of aspartate. Structural analysis of the Ser-171-->Ala and Glu-50/Ser-171-->Ala enzymes by solution X-ray scattering indicates that both mutants exist in the same T quaternary structure as the wild-type enzyme in the absence of ligands, and in the same R quaternary structure in the presence of saturating N-(phosphonoacetyl)-L-aspartate. However, saturating concentrations of carbamoyl phosphate and succinate are unable to convert a significant fraction of either mutant enzyme population to the R quaternary structure, as has been observed previously for the Glu-50-->Ala enzyme. The curves for both the Ser-171-->Ala and Glu-50/Ser-171-->Ala enzymes obtained in the presence of substoichiometric amounts of PALA are linear combinations of the two extreme T and R states. The structural consequences of nucleotide binding to these two enzymes were also investigated. Most surprisingly, the direction and amplitude of the effect of ATP upon the double mutant enzyme were shown to vary depending upon the substrate analogue used. PMID:8931146

Static and dynamic half-life and lifetime molecular turnover of enzymes.

PubMed

Miyawaki, Osato; Kanazawa, Tsukasa; Maruyama, Chika; Dozen, Michiko

2017-01-01

The static half-life of an enzyme is the half-life of a free enzyme not working without substrate and the dynamic half-life is that of an active enzyme working with plenty amount of substrate. These two half-lives were measured and compared for glucoamylase (GA) and β-galactosidase (BG). The dynamic half-life was much longer than the static half-life by one to three orders of magnitude for both enzymes. For BG, the half-life of the enzyme physically entrapped in a membrane reactor was also measured. In this case also, the half-life of BG in the membrane reactor was much longer than the free enzyme without substrate. These results suggest the large difference in stabilities between the free enzyme and the enzyme-substrate complex. This may be related to the natural enzyme metabolism. According to the difference in half-life, the lifetime molecular turnover (LMT), which is the number of product molecules produced by a single molecule of enzyme until it loses its activity completely, was much higher by one to four orders of magnitude for the active enzyme than the free enzyme. The concept of LMT, proposed here, will be important in bioreactor operations with or without immobilization. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Immobilization of polygalacturonase from Aspergillus niger onto activated polyethylene and its application in apple juice clarification.

PubMed

Saxena, Shivalika; Shukla, Surendra; Thakur, Akhilesh; Gupta, Reena

2008-03-01

The present work is focused on efficient immobilization of polygalacturonase on polyethylene matrix, followed by its application in apple juice clarification. Immobilization of polygalacturonase on activated polyethylene and its use in apple juice clarification was not reported so far. Aspergillus niger Van Tieghem (MTCC 3323) produced polygalacturonase when grown in modified Riviere's medium containing pectin as single carbon source by fed-batch culture. The enzyme was precipitated with ethanol and purified by gel filtration chromatography (Sephacryl S-100) and immobilized onto glutaraldehyde-activated polyethylene. The method is very simple and time saving for enzyme immobilization. Various characteristics of immobilized enzyme such as optimum reaction temperature and pH, temperature and pH stability, binding kinetics, efficiency of binding, reusability and metal ion effect on immobilized enzymes were evaluated in comparison to the free enzyme. Both the free and immobilized enzyme showed maximum activity at a temperature of 45 degrees C and pH 4.8. Maximum binding efficiency was 38%. The immobilized enzyme was reusable for 3 cycles with 50% loss of activity after the third cycle. Twenty-four U of immobilized enzyme at 45 degrees C and 1 h incubation time increased the transmittance of the apple juice by about 55% at 650 nm. The immobilized enzyme can be of industrial advantage in terms of sturdiness, availability, inertness, low price, reusability and temperature stability.

Biochemical characterization of a thermostable endonuclease V from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5.

PubMed

Wang, Yuxiao; Zhang, Likui; Zhu, Xinyuan; Li, Yuting; Shi, Haoqiang; Oger, Philippe; Yang, Zhihui

2018-05-22

Endonuclease V (Endo V) is an important enzyme for repairing deoxyinosine in DNA. While bacterial and eukaryotic endo Vs have been well studied, knowledge of archaeal endo Vs is limited. Here, we first presented biochemical characterization of a thermostable endonuclease V from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 (Tba endo V). The recombinant enzyme possessed optimal endonuclease activity for cleaving deoxyinosine-containing DNA at 70-90 °C. Furthermore, Tba endo V can withstand 100 °C for 120 min without significant loss of its activity, suggesting the enzyme is thermostable. Tba endo V exhibited varying cleavage efficiencies at various pH levels from 6.0 to 11.0, among which an optimal pH for the enzyme was 8.0-9.0. In addition, a divalent metal ion was required for the enzyme to cleave DNA. Mn 2+ and Mg 2+ were optimal ions for the enzyme's activity whereas Ca 2+ , Zn 2+ and Co 2+ inhibited the enzyme activity. Moreover, the enzyme activity was suppressed by high NaCl concentration. Tba endo V bound to all DNA substrates; however, the enzyme exhibited a higher affinity for binding to deoxyinosine-containing DNA than normal DNA. Our work provides valuable information for revealing the role of Tba endo V in the base excision repair pathway for deoxyinosine repair in Thermococcus. Copyright © 2018. Published by Elsevier B.V.

Characterization of the receptor-destroying enzyme activity from infectious salmon anaemia virus.

PubMed

Kristiansen, Marianne; Frøystad, Marianne K; Rishovd, Anne Lise; Gjøen, Tor

2002-11-01

Infectious salmon anaemia virus (ISAV) infects cells via the endocytic pathway and, like many other enveloped viruses, ISAV contains a receptor-destroying enzyme. We have analysed this acetylesterase activity with respect to substrate specificity, enzyme kinetics, inhibitors, temperature and pH stability. The ISAV acetylesterase was inhibited by di-isopropyl fluorophosphate (DFP) in a dose-dependent fashion but not by other known hydrolase inhibitors, suggesting that a serine residue is part of the active site. The pH optimum of the enzyme was in the range 7.5-8.0 and the enzymatic activity was lessened at temperatures above 40 degrees C. The effect of DFP on agglutination/elution of erythrocytes by ISAV demonstrated that the acetylesterase activity is the bona fide receptor-destroying enzyme. A haemadsorption assay was used to analyse whether the esterase was active on the surface of infected cells or not.

Characterization of Purified Staphylococcal Lipase1

PubMed Central

Vadehra, D. V.; Harmon, L. G.

1967-01-01

Purified staphylococcal lipase had an optimal pH of 8.3 for activity at 37 C, and an optimal temperature of 45 C at pH 8.0. During storage, the enzyme lost less than 10% of the activity over a period of 21 days at 4 and -23 C. The enzyme retained 93% of the activity when heated for 30 min at 50 C and was 95% destroyed in 30 min at 70 C. The purified lipase was capable of hydrolyzing a variety of natural fats and oils. However, the enzyme was three times more active on nonhydrogenated soybean oil than on hydrogenated soybean oil with an iodine value of <3.0. The enzyme was also capable of hydrolyzing fatty acids on the α, β, and α′ positions of a synthetic mixed triglyceride. In general, the presence of oxidizing agents increased the activity and the presence of reducing agents decreased the activity of the lipase enzyme. PMID:6035042

Release of enzymes from cells: transport and distribution within the extracellular space.

PubMed

Mattenheimer, H; Friedel, R

1977-01-01

The distribution in the extracellular space of enzymes released from organ cells was investigated using three models: (1) comparison of enzyme activities in blood plasma and lymph of the ductus thoracicus (dog) and plasma and intestinal lymph (rat); (2) i.v. injection of heterologous, homologous and autologous enzymes in order to increase acutely the activities and to measure the rate constants for the distribution and elimination of the enzymes (rat); or (3) plasmapheresis in order to create an enzyme activity gradient from the interstitial space and to determine the rate constants for the reestablishment of the equilibrium between the extra and intravascular compartments (rat). The results suggest that the enzymes are mainly released into the interstitial fluid and transported via the lymph into the intravascular compartment. From there the enzymes diffuse back into the interstitial compartment and are eliminated by a yet unknown mechanism. Transport of enzymes across the capillary membranes in both directions depends on (1) the permeability of the capillary membranes, which varies from region to region and (2) the molecular seizes of the enzymes.

Elevation of Glucose 6-Phosphate Dehydrogenase Activity Induced by Amplified Insulin Response in Low Glutathione Levels in Rat Liver

PubMed Central

Taniguchi, Misako; Mori, Nobuko; Iramina, Chizuru

2016-01-01

Weanling male Wistar rats were fed on a 10% soybean protein isolate (SPI) diet for 3 weeks with or without supplementing 0.3% sulfur-containing amino acids (SAA; methionine or cystine) to examine relationship between glutathione (GSH) levels and activities of NADPH-producing enzymes, glucose 6-phosphate dehydrogenase (G6PD) and malic enzyme (ME), in the liver. Of rats on the 10% SPI diet, GSH levels were lower and the enzyme activities were higher than of those fed on an SAA-supplemented diet. Despite the lower GSH level, γ-glutamylcysteine synthetase (γ-GCS) activity was higher in the 10% SPI group than other groups. Examination of mRNAs of G6PD and ME suggested that the GSH-suppressing effect on enzyme induction occurred prior to and/or at transcriptional levels. Gel electrophoresis of G6PD indicated that low GSH status caused a decrease in reduced form and an increase in oxidized form of the enzyme, suggesting an accelerated turnover rate of the enzyme. In primary cultured hepatocytes, insulin response to induce G6PD activity was augmented in low GSH levels manipulated in the presence of buthionine sulfoximine. These findings indicated that elevation of the G6PD activity in low GSH levels was caused by amplified insulin response for expression of the enzyme and accelerated turnover rate of the enzyme molecule. PMID:27597985

Statistical Optimization of Fibrinolytic Enzyme Production Using Agroresidues by Bacillus cereus IND1 and Its Thrombolytic Activity In Vitro

PubMed Central

Prakash Vincent, Samuel Gnana

2014-01-01

A potent fibrinolytic enzyme-producing Bacillus cereus IND1 was isolated from the Indian food, rice. Solid-state fermentation was carried out using agroresidues for the production of fibrinolytic enzyme. Among the substrates, wheat bran supported more enzyme production and has been used for the optimized enzyme production by statistical approach. Two-level full-factorial design demonstrated that moisture, supplementation of beef extract, and sodium dihydrogen phosphate have significantly influenced enzyme production (P < 0.05). A central composite design resulted in the production of 3699 U/mL of enzyme in the presence of 0.3% (w/w) beef extract and 0.05% (w/w) sodium dihydrogen phosphate, at 100% (v/w) moisture after 72 h of fermentation. The enzyme production increased fourfold compared to the original medium. This enzyme was purified to homogeneity by ammonium sulfate precipitation, diethylaminoethyl-cellulose ion-exchange chromatography, Sephadex G-75 gel filtration chromatography, and casein-agarose affinity chromatography and had an apparent molecular mass of 29.5 kDa. The optimum pH and temperature for the activity of fibrinolytic enzyme were found to be 8.0 and 60°C, respectively. This enzyme was highly stable at wide pH range (7.0–9.0) and showed 27% ± 6% enzyme activity after initial denaturation at 60°C for 1 h. In vitro assays revealed that the enzyme could activate plasminogen and significantly degraded the fibrin net of blood clot, which suggests its potential as an effective thrombolytic agent. PMID:25003130

Analysis of the activation of acetylcholinesterase by carbon nanoparticles using a monolithic immobilized enzyme microreactor: role of the water molecules in the active site gorge.

PubMed

Ibrahim, Firas; Andre, Claire; Iutzeler, Anne; Guillaume, Yves Claude

2013-10-01

A biochromatographic system was used to study the direct effect of carbon nanoparticles (CNPs) on the acetylcholinesterase (AChE) activity. The AChE enzyme was covalently immobilized on a monolithic CIM-disk via its NH2 residues. Our results showed an increase in the AChE activity in presence of CNPs. The catalytic constant (k(cat)) was increased while the Michaelis constant (K(m)) was slightly decreased. This indicated an increase in the enzyme efficiency with increase of the substrate affinity to the active site. The thermodynamic data of the activation mechanism of the enzyme, i.e. ΔH* and ΔS*, showed no change in the substrate interaction mechanism with the anionic binding site. The increase of the enthalpy (ΔH*) and the entropy (ΔS*) with decrease in the free energy of activation (Ea) was related to structural conformation change in the active site gorge. This affected the stability of water molecules in the active site gorge and facilitated water displacement by substrate for entering to the active site of the enzyme.

Functional screening of pharmacological chaperones via restoration of enzyme activity upon denaturation.

PubMed

Shanmuganathan, Meera; Britz-McKibbin, Philip

2012-10-02

Pharmacological chaperones (PCs) are small molecules that stabilize and promote protein folding. Enzyme inhibition is widely used for PC selection; however, it does not accurately reflect chaperone activity. We introduce a functional assay for characterization of PCs based on their capacity to restore enzyme activity that is abolished upon chemical denaturation. Dose-dependent activity curves were performed as a function of urea to assess the chaperone potency of various ligands to β-glucocerebrosidase as a model system. Restoration of enzyme activity upon denaturation allows direct screening of PCs for treatment of genetic disorders associated with protein deficiency, such as Gaucher disease.

PhAP protease from Pseudoalteromonas haloplanktis TAC125: Gene cloning, recombinant production in E. coli and enzyme characterization

NASA Astrophysics Data System (ADS)

de Pascale, D.; Giuliani, M.; De Santi, C.; Bergamasco, N.; Amoresano, A.; Carpentieri, A.; Parrilli, E.; Tutino, M. L.

2010-08-01

Cold-adapted proteases have been found to be the dominant activity throughout the cold marine environment, indicating their importance in bacterial acquisition of nitrogen-rich complex organic compounds. However, few extracellular proteases from marine organisms have been characterized so far, and the mechanisms that enable their activity in situ are still largely unknown. Aside from their ecological importance and use as model enzyme for structure/function investigations, cold-active proteolytic enzymes offer great potential for biotechnological applications. Our studies on cold adapted proteases were performed on exo-enzyme produced by the Antarctic marine bacterium Pseudoalteromonas haloplanktis TAC125. By applying a proteomic approach, we identified several proteolytic activities from its culture supernatant. PhAP protease was selected for further investigations. The encoding gene was cloned and the protein was recombinantly produced in E. coli cells. The homogeneous product was biochemically characterised and it turned out that the enzyme is a Zn-dependent aminopeptidase, with an activity dependence from assay temperature typical of psychrophilic enzymes.

Antioxidant enzyme activities are affected by salt content and temperature and influence muscle lipid oxidation during dry-salted bacon processing.

PubMed

Jin, Guofeng; He, Lichao; Yu, Xiang; Zhang, Jianhao; Ma, Meihu

2013-12-01

Fresh pork bacon belly was used as material and manufactured into dry-salted bacon through salting and drying-ripening. During processing both oxidative stability and antioxidant enzyme stability were evaluated by assessing peroxide value (PV), thiobarbituric acid reactive substances (TBARS) and activities of catalase, glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD), and their correlations were also analysed. The results showed that all antioxidant enzyme activities decreased (p<0.05) until the end of process; GSH-Px was the most unstable one followed by catalase. Antioxidant enzyme activities were negatively correlated with TBARS (p<0.05), but the correlations were decreased with increasing process temperature. Salt showed inhibitory effect on all antioxidant enzyme activities and was concentration dependent. These results indicated that when process temperature and salt content were low at the same time during dry-salted bacon processing, antioxidant enzymes could effectively control lipid oxidation. Copyright © 2013 Elsevier Ltd. All rights reserved.

'Enzyme Test Bench': A biochemical application of the multi-rate modeling

NASA Astrophysics Data System (ADS)

Rachinskiy, K.; Schultze, H.; Boy, M.; Büchs, J.

2008-11-01

In the expanding field of 'white biotechnology' enzymes are frequently applied to catalyze the biochemical reaction from a resource material to a valuable product. Evolutionary designed to catalyze the metabolism in any life form, they selectively accelerate complex reactions under physiological conditions. Modern techniques, such as directed evolution, have been developed to satisfy the increasing demand on enzymes. Applying these techniques together with rational protein design, we aim at improving of enzymes' activity, selectivity and stability. To tap the full potential of these techniques, it is essential to combine them with adequate screening methods. Nowadays a great number of high throughput colorimetric and fluorescent enzyme assays are applied to measure the initial enzyme activity with high throughput. However, the prediction of enzyme long term stability within short experiments is still a challenge. A new high throughput technique for enzyme characterization with specific attention to the long term stability, called 'Enzyme Test Bench', is presented. The concept of the Enzyme Test Bench consists of short term enzyme tests conducted under partly extreme conditions to predict the enzyme long term stability under moderate conditions. The technique is based on the mathematical modeling of temperature dependent enzyme activation and deactivation. Adapting the temperature profiles in sequential experiments by optimum non-linear experimental design, the long term deactivation effects can be purposefully accelerated and detected within hours. During the experiment the enzyme activity is measured online to estimate the model parameters from the obtained data. Thus, the enzyme activity and long term stability can be calculated as a function of temperature. The results of the characterization, based on micro liter format experiments of hours, are in good agreement with the results of long term experiments in 1L format. Thus, the new technique allows for both: the enzyme screening with regard to the long term stability and the choice of the optimal process temperature. The presented article gives a successful example for the application of multi-rate modeling, experimental design and parameter estimation within biochemical engineering. At the same time, it shows the limitations of the methods at the state of the art and addresses the current problems to the applied mathematics community.

A Simple and Accurate Method for Measuring Enzyme Activity.

ERIC Educational Resources Information Center

Yip, Din-Yan

1997-01-01

Presents methods commonly used for investigating enzyme activity using catalase and presents a new method for measuring catalase activity that is more reliable and accurate. Provides results that are readily reproduced and quantified. Can also be used for investigations of enzyme properties such as the effects of temperature, pH, inhibitors,…

Microbial Interactions Associated with Biofilms Attached to Trichodesmium spp. and Detrital Particles in the Ocean

DTIC Science & Technology

2010-06-01

AHLs enhanced certain organic-matter degrading hydrolytic enzyme activities. My results suggest that AHL-QS is a factor regulating...biogeochemically relevant enzyme activities on sinking POC and within the biofilms attached to Trichodesmium colonies and thereby may impact the timing and... Enzyme activity assays ......................................................................................................................232 RESULTS

Activities of Tricarboxylic Acid Cycle Enzymes, Glyoxylate Cycle Enzymes, and Fructose Diphosphatase in Bakers' Yeast During Adaptation to Acetate Oxidation

PubMed Central

Gosling, J. P.; Duggan, P. F.

1971-01-01

Bakers' yeast oxidizes acetate at a high rate only after an adaptation period during which the capacity of the glyoxylate cycle is found to increase. There was apparently no necessity for the activity of acetyl-coenzyme A synthetase, the capacity of the tricarboxylic acid cycle, or the concentrations of the cytochromes to increase for this adaptation to occur. Elevation of fructose 1,6 diphosphatase occurred only when acetate oxidation was nearly maximal. Cycloheximide almost completely inhibited adaptation as well as increases in the activities of isocitrate lyase and aconitate hydratase, the only enzymes assayed. p-Fluorophenylalanine was partially effective and chloramphenicol did not inhibit at all. The presence of ammonium, which considerably delayed adaptation of the yeast to acetate oxidation, inhibited the increases in the activities of the glyoxylate cycle enzymes to different degrees, demonstrating noncoordinate control of these enzymes. Under the various conditions, the only enzyme activity increase consistently related to the rising oxygen uptake rate was that of isocitrate lyase which apparently limited the activity of the cycle. PMID:5557595

Reduction of nuclear encoded enzymes of mitochondrial energy metabolism in cells devoid of mitochondrial DNA

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

Mueller, Edith E., E-mail: ed.mueller@salk.at; Mayr, Johannes A., E-mail: h.mayr@salk.at; Zimmermann, Franz A., E-mail: f.zimmermann@salk.at

2012-01-20

Highlights: Black-Right-Pointing-Pointer We examined OXPHOS and citrate synthase enzyme activities in HEK293 cells devoid of mtDNA. Black-Right-Pointing-Pointer Enzymes partially encoded by mtDNA show reduced activities. Black-Right-Pointing-Pointer Also the entirely nuclear encoded complex II and citrate synthase exhibit reduced activities. Black-Right-Pointing-Pointer Loss of mtDNA induces a feedback mechanism that downregulates complex II and citrate synthase. -- Abstract: Mitochondrial DNA (mtDNA) depletion syndromes are generally associated with reduced activities of oxidative phosphorylation (OXPHOS) enzymes that contain subunits encoded by mtDNA. Conversely, entirely nuclear encoded mitochondrial enzymes in these syndromes, such as the tricarboxylic acid cycle enzyme citrate synthase (CS) and OXPHOS complexmore » II, usually exhibit normal or compensatory enhanced activities. Here we report that a human cell line devoid of mtDNA (HEK293 {rho}{sup 0} cells) has diminished activities of both complex II and CS. This finding indicates the existence of a feedback mechanism in {rho}{sup 0} cells that downregulates the expression of entirely nuclear encoded components of mitochondrial energy metabolism.« less

Development of radiometric assays for quantification of enzyme activities of the key enzymes of thyroid hormones metabolism.

PubMed

Pavelka, S

2014-01-01

We newly elaborated and adapted several radiometric enzyme assays for the determination of activities of the key enzymes engaged in the biosynthesis (thyroid peroxidase, TPO) and metabolic transformations (conjugating enzymes and iodothyronine deiodinases, IDs) of thyroid hormones (THs) in the thyroid gland and in peripheral tissues, especially in white adipose tissue (WAT). We also elaborated novel, reliable radiometric methods for extremely sensitive determination of enzyme activities of IDs of types 1, 2 and 3 in microsomal fractions of different rat and human tissues, as well as in homogenates of cultured mammalian cells. The use of optimized TLC separation of radioactive products from the unconsumed substrates and film-less autoradiography of radiochromatograms, taking advantage of storage phosphor screens, enabled us to determine IDs enzyme activities as low as 10(-18) katals. In studies of the interaction of fluoxetine (Fluox) with the metabolism of THs, we applied adapted radiometric enzyme assays for iodothyronine sulfotransferases (ST) and uridine 5'-diphospho-glucuronyltransferase (UDP-GT). Fluox is the most frequently used representative of a new group of non-tricyclic antidepressant drugs--selective serotonin re-uptake inhibitors. We used the elaborated assays for quantification the effects of Fluox and for the assessment of the degree of potential induction of rat liver ST and/or UDP-GT enzyme activities by Fluox alone or in combination with T(3). Furthermore, we studied possible changes in IDs activities in murine adipose tissue under the conditions that promoted either tissue hypertrophy (obesogenic treatment) or involution (caloric restriction), and in response to leptin, using our newly developed radiometric enzyme assays for IDs. Our results suggest that deiodinase D1 has a functional role in WAT, with D1 possibly being involved in the control of adipose tissue metabolism and/or accumulation of the tissue. Significant positive correlation between specific enzyme activity of D1 in WAT and plasma leptin levels was found. The newly developed and adapted radiometric enzyme assays proved to be very useful tools for studies of factors modulating THs metabolism, not only in model animals but also in clinical studies of human obesity.

Screening and Characterization of Polygalacturonase as Potential Enzyme for Keprok Garut Orange (Citrus nobilis var. chrysocarpa) Juice Clarification

NASA Astrophysics Data System (ADS)

Widowati, E.; Utami, R.; Kalistyatika, K.

2017-11-01

Use of thermostable enzyme from bacilli for industrial application is significant. This research aimed to isolate thermophilic pectinolytic bacteria from orange peel and vegetable waste which produced thermostable polygalacturonase, to investigate the polygalacturonase ability in clarifying keprok Garut orange juice, and to characterize polygalacturonase based on pH optimum, temperature optimum, enzyme stability, enzyme kinetics KM, and Vmax. Obtained, 14 isolates that further selected to 4 best isolates based on highest polygalacturonase activity and keprok Garut orange juice clarification ability. Four selected enzyme isolates were AR 2, AR 4, KK 4, and KK 5 had ability to increase juice transmittance, decrease juice viscosity and also reduce total soluble solid. Furthermore 4 selected isolates were partially purified by ammonium sulphate precipitation and dialysis method. Four partially purified enzymes were known that enzyme character of AR 2 optimum at pH 6; AR 4 optimum at pH 5.5; KK 4 optimum at pH 6; and KK 5 optimum at pH 4.5. Four enzymes were optimum at temperature 60°C thus stable at temperature 50-60°C, this characteristic indicate that enzymes were thermostable. AR 2 showed active activity stable at pH 4-7; AR 4 showed active activity stable at pH 6-7; KK 4 showed active activity stable at pH 4-6; however KK 5 stable at pH 4-5. Enzyme AR 2 and KK 4 was getting inactive at pH 11, thus AR 4 and KK 5 inactive at pH 12. KM value of AR 2, AR 4, KK 4, and KK 5 was 0.0959; 0.0974; 0.0966; and 0.178 mg/ml respectively. Vmax of AR 2, AR 4, KK 4, and KK 5 was 0.0203; 0.0202; 0.0185; and 0.0229 U/ml respectively. Enzyme AR 2 was the most compatible enzyme to be applied in keprok Garut orange juice clarification for it had the lowest KM value.

The 46 kDa dimeric protein from Variovorax paradoxus shows faster methotrexate degrading activity in its nanoform compare to the native enzyme.

PubMed

Bayineni, Venkata Krishna; Venkatesh, Krishna; Sahu, Chandan Kumar; Kadeppagari, Ravi-Kumar

2016-04-01

Methotrexate degrading enzymes are required to overcome the toxicity of the methotrexate while treating the cancer. The enzyme from Variovorax paradoxus converts the methotrexate in to non toxic products. Methotrexate degrading enzyme from V. paradoxus is a dimeric protein with a molecular mass of 46 kDa and it acts on casein and gelatin. This enzyme is optimally active at pH 7.5 and 40°C and nanoparticles of this enzyme were prepared by desolvation-crosslinking method. Enzyme nanoparticles could degrade methotrexate faster than the native enzyme and they show lower Km compare to the native enzyme. Enzyme nanoparticles show better thermostability and they were stable for much longer time in the serum compare to the native enzyme. Enzyme nanoparticles show better functionality than the native enzyme while clearing the methotrexate added to the serum suggesting their advantage over the native enzyme for the therapeutic and biotechnological applications. Copyright © 2016 Elsevier Inc. All rights reserved.

Purification and characterization of a tartrate-resistant acid phosphatase from human osteoclastomas.

PubMed Central

Hayman, A R; Warburton, M J; Pringle, J A; Coles, B; Chambers, T J

1989-01-01

Tartrate-resistant acid phosphatase is one of the major enzymes produced and secreted by osteoclasts. To obtain sufficient enzyme for biochemical characterization, we have purified this enzyme from human osteoclastomas by sequential chromatography on SP-Sephadex, CM-Sephadex, hydroxylapatite, Sephadex G-150 and concanavalin A-Sepharose. The purification over the original tumour extract was about 2000-fold, with a yield of 10%. The enzyme appeared to be homogeneous when assessed by SDS/polyacrylamide-gel electrophoresis. Both gel filtration and SDS/polyacrylamide-gel electrophoresis indicated an Mr of about 30,000. The reduced and alkylated enzyme consists of two subunits with Mrs of 15,000 and 17,500. The N-terminal amino acid sequence of both subunits indicates that there is a high degree of identity between the osteoclastoma enzyme and similar enzymes purified from spleen and uterus. Using 4-methylumbelliferyl phosphate as substrate, the specific activity of the purified enzyme was 387 units.mg-1, and the Km was 284 microns. The pH optimum was 5.7. Unlike similar enzymes purified from human and bovine bone, osteoclastoma acid phosphatase is not activated by reducing agents (2-mercaptoethanol or ascorbic acid). The enzyme contains 4.8 mol of Fe2+/3+, 0.3 mol of Mn2+ and 1.7 mol of Mg2+ per mol of enzyme. Although the enzyme loses 50% of its activity in the presence of EDTA, it is not inhibited by the iron chelator 1,10-phenanthroline. However, the enzyme is activated to a small extent by Mn2+ and Mg2+. Using a variety of substrates and inhibitors, we demonstrate that there are differences between the osteoclastoma acid phosphatase and the enzyme purified from other sources. Images Fig. 1. Fig. 2. Fig. 4. PMID:2775236

Ultrasound assisted intensification of enzyme activity and its properties: a mini-review.

PubMed

Nadar, Shamraja S; Rathod, Virendra K

2017-08-22

Over the last decade, ultrasound technique has emerged as the potential technology which shows large applications in food and biotechnology processes. Earlier, ultrasound has been employed as a method of enzyme inactivation but recently, it has been found that ultrasound does not inactivate all enzymes, particularly, under mild conditions. It has been shown that the use of ultrasonic treatment at appropriate frequencies and intensity levels can lead to enhanced enzyme activity due to favourable conformational changes in protein molecules without altering its structural integrity. The present review article gives an overview of influence of ultrasound irradiation parameters (intensity, duty cycle and frequency) and enzyme related factors (enzyme concentration, temperature and pH) on the catalytic activity of enzyme during ultrasound treatment. Also, it includes the effect of ultrasound on thermal kinetic parameters and Michaelis-Menten kinetic parameters (k m and V max ) of enzymes. Further, in this review, the physical and chemical effects of ultrasound on enzyme have been correlated with thermodynamic parameters (enthalpy and entropy). Various techniques used for investigating the conformation changes in enzyme after sonication have been highlighted. At the end, different techniques of immobilization for ultrasound treated enzyme have been summarized.

Correction: Mesoporous titania thin films as efficient enzyme carriers for paraoxon determination/detoxification: effects of enzyme binding and pore hierarchy on the biocatalyst activity and reusability.

PubMed

Frančič, N; Bellino, M G; Soler-Illia, G J A A; Lobnik, A

2016-07-07

Correction for 'Mesoporous titania thin films as efficient enzyme carriers for paraoxon determination/detoxification: effects of enzyme binding and pore hierarchy on the biocatalyst activity and reusability' by N. Frančičet al., Analyst, 2014, 139, 3127-3136.

The interaction of the Eco R1 restriction enzyme E.coli with nucleotides

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

Hollis, Donald F.

1979-11-01

The Eco R1 restriction enzyme can be shown to be inhibited by nucleotides which correspond to any part of its known site of phosphodiesterase activity. A series of di-, tetra-, and hexa-nucleotide fragments were synthesized and their effect on the activity of the enzyme upon superhelical Co1 E1 DNA studied. The inhibition caused by the individual mononucleotides were also studied. In general all the nucleotide fragments showed some form of interaction with the enzyme system. Tetranucleotides were stronger inhibitors than dinucleotides, which in turn were stronger inhibitors than the mononucleotides. Within each category of inhibitors, those containing the phosphodiester bondmore » which is acted upon by the enzyme were the strongest inhibitors. Only those fragments which were consistent with the enzymes site of activity showed competitive inhibition kinetics. Nucleotides which do not fit within the site of phosphodiesterase activity show non-competitive inhibition kinetics.« less

Modification of enzymes by use of high-pressure homogenization.

PubMed

Dos Santos Aguilar, Jessika Gonçalves; Cristianini, Marcelo; Sato, Helia Harumi

2018-07-01

High-pressure is an emerging and relatively new technology that can modify various molecules. High-pressure homogenization (HPH) has been used in several studies on protein modification, especially in enzymes used or found in food, from animal, plant or microbial resources. According to the literature, the enzymatic activity can be modulated under pressure causing inactivation, stabilization or activation of the enzymes, which, depending on the point of view could be very useful. Homogenization can generate changes in the structure of the enzyme modifying various chemical bonds (mainly weak bonds) causing different denaturation levels and, consequently, affecting the catalytic activity. This review aims to describe the various alterations due to HPH treatment in enzymes, to show the influence of high-pressure on proteins and to report the HPH effects on the enzymatic activity of different enzymes employed in the food industry and research. Copyright © 2018 Elsevier Ltd. All rights reserved.

Microbial Activity and Silica Degradation in Rice Straw

NASA Astrophysics Data System (ADS)

Kim, Esther Jin-kyung

Abundantly available agricultural residues like rice straw have the potential to be feedstocks for bioethanol production. Developing optimized conditions for rice straw deconstruction is a key step toward utilizing the biomass to its full potential. One challenge associated with conversion of rice straw to bioenergy is its high silica content as high silica erodes machinery. Another obstacle is the availability of enzymes that hydrolyze polymers in rice straw under industrially relevant conditions. Microbial communities that colonize compost may be a source of enzymes for bioconversion of lignocellulose to products because composting systems operate under thermophilic and high solids conditions that have been shown to be commercially relevant. Compost microbial communities enriched on rice straw could provide insight into a more targeted source of enzymes for the breakdown of rice straw polysaccharides and silica. Because rice straw is low in nitrogen it is important to understand the impact of nitrogen concentrations on the production of enzyme activity by the microbial community. This study aims to address this issue by developing a method to measure microbial silica-degrading activity and measure the effect of nitrogen amendment to rice straw on microbial activity and extracted enzyme activity during a high-solids, thermophilic incubation. An assay was developed to measure silica-degrading enzyme or silicase activity. This process included identifying methods of enzyme extraction from rice straw, identifying a model substrate for the assay, and optimizing measurement techniques. Rice straw incubations were conducted with five different levels of nitrogen added to the biomass. Microbial activity was measured by respiration and enzyme activity. A microbial community analysis was performed to understand the shift in community structure with different treatments. With increased levels of nitrogen, respiration and cellulose and hemicellulose degrading activity increased. Silicase activity did not change across nitrogen treatments despite a shift in microbial community with varied nitrogen concentration. Samples treated with different nitrogen concentrations had similar levels of diversity, however the microbial community composition differed with added nitrogen. The results demonstrated that adding nitrogen to rice straw during thermophilic decomposition nurtured a more active microbial community and promoted enzyme secretion thus improving the ability to discover enzymes for rice straw deconstruction. These results can inform future experiments for cultivating a unique, thriving compost-derived microbial community that can successfully decompose rice straw. Understanding the silicase activity of microorganisms may alleviate the challenges associated with silica in various feedstocks.

Expression of a Deschampsia antarctica Desv. Polypeptide with Lipase Activity in a Pichia pastoris Vector

PubMed Central

Rabert, Claudia; Gutiérrez-Moraga, Ana; Navarrete-Gallegos, Alejandro; Navarrete-Campos, Darío; Bravo, León A.; Gidekel, Manuel

2014-01-01

The current study isolated and characterized the Lip3F9 polypeptide sequence of Deschampsia antarctica Desv. (GeneBank Accession Number JX846628), which was found to be comprised of 291 base pairs and was, moreover, expressed in Pichia pastoris X-33 cells. The enzyme was secreted after 24 h of P. pastoris culture incubation and through induction with methanol. The expressed protein showed maximum lipase activity (35 U/L) with an optimal temperature of 37 °C. The lipase-expressed enzyme lost 50% of its specific activity at 25 °C, a behavior characteristic of a psychrotolerant enzyme. Recombinant enzyme activity was measured in the presence of ionic and non-ionic detergents, and a decrease in enzyme activity was detected for all concentrations of ionic and non-ionic detergents assessed. PMID:24514564

The inhibitory effect of metals and other ions on acid phosphatase activity from Vigna aconitifolia seeds.

PubMed

Srivastava, Pramod Kumar; Anand, Asha

2015-01-01

Sensitivity of acid phosphatase from Vigna aconitifolia seeds to metal ions, fluoride, and phosphate was examined. All the effectors had different degree of inhibitory effect on the enzyme. Among metal ions, molybdate and ferric ion were observed to be most potent inhibitors and both exhibited mixed type of inhibition. Acid phosphatase activity was inhibited by Cu2+ in a noncompetitive manner. Zn and Mn showed mild inhibition on the enzyme activity. Inhibition kinetics analysis explored molybdate as a potent inhibitor for acid phosphatase in comparison with other effectors used in this study. Fluoride was the next most strong inhibitor for the enzyme activity, and caused a mixed type of inhibition. Phosphate inhibited the enzyme competitively, which demonstrates that inhibition due to phosphate is one of the regulatory factors for enzyme activity.

Hereditary fructose intolerance mimicking a biochemical phenotype of mucolipidosis: A review of the literature of secondary causes of lysosomal enzyme activity elevation in serum.

PubMed

Ferreira, Carlos R; Devaney, Joseph M; Hofherr, Sean E; Pollard, Laura M; Cusmano-Ozog, Kristina

2017-02-01

We describe a patient with failure to thrive, hepatomegaly, liver dysfunction, and elevation of multiple plasma lysosomal enzyme activities mimicking mucolipidosis II or III, in whom a diagnosis of hereditary fructose intolerance (HFI) was ultimately obtained. She presented before introduction of solid foods, given her consumption of a fructose-containing infant formula. We present the most extensive panel of lysosomal enzyme activities reported to date in a patient with HFI, and propose that multiple enzyme elevations in plasma, especially when in conjunction with a normal plasma α-mannosidase activity, should elicit a differential diagnosis of HFI. We also performed a review of the literature on the different etiologies of elevated lysosomal enzyme activities in serum or plasma. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

A-Type Carrier Protein ErpA Is Essential for Formation of an Active Formate-Nitrate Respiratory Pathway in Escherichia coli K-12

PubMed Central

Pinske, Constanze

2012-01-01

A-type carrier (ATC) proteins of the Isc (iron-sulfur cluster) and Suf (sulfur mobilization) iron-sulfur ([Fe-S]) cluster biogenesis pathways are proposed to traffic preformed [Fe-S] clusters to apoprotein targets. In this study, we analyzed the roles of the ATC proteins ErpA, IscA, and SufA in the maturation of the nitrate-inducible, multisubunit anaerobic respiratory enzymes formate dehydrogenase N (Fdh-N) and nitrate reductase (Nar). Mutants lacking SufA had enhanced activities of both enzymes. While both Fdh-N and Nar activities were strongly reduced in an iscA mutant, both enzymes were inactive in an erpA mutant and in a mutant unable to synthesize the [Fe-S] cluster scaffold protein IscU. It could be shown for both Fdh-N and Nar that loss of enzyme activity correlated with absence of the [Fe-S] cluster-containing small subunit. Moreover, a slowly migrating form of the catalytic subunit FdnG of Fdh-N was observed, consistent with impeded twin arginine translocation (TAT)-dependent transport. The highly related Fdh-O enzyme was also inactive in the erpA mutant. Although the Nar enzyme has its catalytic subunit NarG localized in the cytoplasm, it also exhibited aberrant migration in an erpA iscA mutant, suggesting that these modular enzymes lack catalytic integrity due to impaired cofactor biosynthesis. Cross-complementation experiments demonstrated that multicopy IscA could partially compensate for lack of ErpA with respect to Fdh-N activity but not Nar activity. These findings suggest that ErpA and IscA have overlapping roles in assembly of these anaerobic respiratory enzymes but demonstrate that ErpA is essential for the production of active enzymes. PMID:22081393

Hydrolytic and ligninolytic enzyme activities in the Pb contaminated soil inoculated with litter-decomposing fungi.

PubMed

Kähkönen, Mika A; Lankinen, Pauliina; Hatakka, Annele

2008-06-01

The impact of Pb contamination was tested to five hydrolytic (beta-glucosidase, beta-xylosidase, beta-cellobiosidase, alpha-glucosidase and sulphatase) and two ligninolytic (manganese peroxidase, MnP and laccase) enzyme activities in the humus layer in the forest soil. The ability of eight selected litter-degrading fungi to grow and produce extracellular enzymes in the heavily Pb (40 g Pb of kg ww soil(-1)) contaminated and non-contaminated soil in the non-sterile conditions was also studied. The Pb content in the test soil was close to that of the shooting range at Hälvälä (37 g Pb of kg ww soil(-1)) in Southern Finland. The fungi were Agaricus bisporus, Agrocybe praecox, Gymnopus peronatus, Gymnopilus sapineus, Mycena galericulata, Gymnopilus luteofolius, Stropharia aeruginosa and Stropharia rugosoannulata. The Pb contamination (40 g Pb of kg ww soil(-1)) was deleterious to all five studied hydrolytic enzyme activities after five weeks of incubation. All five hydrolytic enzyme activities were significantly higher in the soil than in the extract of the soil indicating that a considerable part of enzymes were particle bound in the soils. Hydrolytic enzyme activities were higher in the non-contaminated soil than in the Pb contaminated soil. Fungal inocula increased the hydrolytic enzyme activities beta-cellobiosidase and beta-glucosidase in non-contaminated soils. All five hydrolytic enzyme activities were similar with fungi and without fungi in the Pb contaminated soil. This was in line that Pb contamination (40 g Pb of kg ww soil(-1)) depressed the growth of all fungi compared to those grown without Pb in the soil. Laccase and MnP activities were low in both Pb contaminated and non-contaminated soil cultures. MnP activities were higher in soil cultures containing Pb than without Pb. Our results showed that Pb in the shooting ranges decreased fungal growth and microbial functioning in the soil.

Effect of the electrical currents generated by the intestinal smooth muscle layers on pancreatic enzyme activity: an in vitro study.

PubMed

Dabek, Marta; Podgurniak, Paweł; Piedra, Jose L Valverde; Szymańczyk, Sylwia; Filip, Rafał; Wojtasz-Pajak, Anna; Werpachowska, Eliza; Podgurniak, Malgorzata; Pierzynowski, Stefan G

2007-05-01

Gut enzymes in the small intestine are exposed to extremely low electrical currents (ELEC) generated by the smooth muscle. In the present study, the in vitro tests were undertaken to evaluate the effect of these electric currents on the activity of the proteolytic pancreatic digestive enzymes. A simulator generating the typical electrical activity of pig gut was used for these studies. The electric current emitted by the simulator was transmitted to the samples, containing enzyme and its substrate, using platinum plate electrodes. All samples were incubated at 37 degrees C for 1 h. The changes in optical density, corresponding to enzyme activity, in samples stimulated for 1 h with ELEC was compared with that not exposed to ELEC. The obtained results show that the electrical current with the characteristics of the myoelectrical migrating complex (MMC) has an influence on pancreatic enzyme activity. Increased endopeptidase and reduced exopeptidase activity was noticed in samples treated with ELEC. This observation can be of important as analyzed factors which can alter enzymatic activity of the gut, can thus also affect feed/food digestibility. (c) 2007 Wiley-Liss, Inc.

Screening and characterization of thermo-active enzymes of biotechnological interest produced by thermophilic Bacillus isolated from hot springs in Tunisia.

PubMed

Thebti, Wajdi; Riahi, Yosra; Gharsalli, Rawand; Belhadj, Omrane

2016-01-01

As part of the contribution to the global efforts in research of thermostable enzymes being of industrial interest, we focus on the isolation of thermophilic bacteria from Tunisian hot springs. Among the collection of 161 strains of thermophilic Bacillus isolated from different samples of thermal water in Tunisia, 20% are capable of growing at 100°C and the rest grow at 70°C or above. Preliminary activity tests on media supplemented with enzyme-substrates confirmed that 35 strains produced amylases, 37 - proteases, 43 - cellulases, 31 - xylanases and 37 - mannanases. The study of the effect of temperature on enzyme activity led to determination of the optimal temperatures of activities that vary between 60 and 100°C. Several enzymes were active at high temperatures (80, 90 and 100°C) and kept their activity even at 110°C. Several isolated strains producing enzymes with high optimal temperatures of activity were described for the first time in this study. Both strains B62 and B120 are producers of amylase, protease, cellulase, xylanase, and mannanase. The sequencing of 16S DNA identified isolated strains as Geobacillus kaustophillus, Aeribacillus pallidus, Geobacillus galactosidasus and Geobacillus toebii.

Potentials for Soil Enzyme as Indicators of Ecological Management

NASA Technical Reports Server (NTRS)

Senwo, Z. N.; Manu, A.; Coleman, T. L.

1997-01-01

Activity measurements of selected soil enzymes (cellulase, glucosidase, amidohydrolase, phosphatase, arylsulfatase) involved in carbon, nitrogen, phosphorus, and sulfur cycling in the biosphere, hold potential as early and sensitive indicators of soil ecological stress and restoration, These measurements are advantageous because the procedures are simple, rapid, and reproducible over time. Enzyme activities are sensitive to short-term changes in soil and kind-use management. Enzyme activities have also been observed to be closely related to soil organic matter proposed as an index of soil quality.

Changes in the biological activity of chestnut soils upon the long-term application of fertilizers in a rotation with oil-bearing crops

NASA Astrophysics Data System (ADS)

Eleshev, R. E.; Bakenova, Z. B.

2012-11-01

Experimental studies showed that irrigated chestnut soils on the piedmont of the Zailiiskiy Alatau Range are characterized by the moderate activity of the hydrolytic and redox enzymes. The use of these soils in the crop rotation system increases the hydrolytic activity of the enzymes (invertase, urease, and ATP synthase) by 30% in comparison with the monoculture; at the same time, it does not have a significant impact on the changes in the biological activity of the redox enzymes (catalase and dehydrogenase). The hydrolytic activity of the soils is activated to a greater extent in the crop rotation and in the monoculture against the background application of organic fertilizers. In this case, the recommended rates of mineral fertilizers do not inhibit the activity of the hydrolytic and redox enzymes. An increase in the hydrolytic activity of the enzymes directly affects the yield of oilseed flax. Therefore, indices of the hydrolytic activity of soils can be used as a test for the diagnostics of the efficiency of fertilizers both in crop rotation and monoculture systems.

Hydroxycinnamic acid decarboxylase activity of Brettanomyces bruxellensis involved in volatile phenol production: relationship with cell viability.

PubMed

Laforgue, R; Lonvaud-Funel, A

2012-12-01

Brettanomyces bruxellensis populations have been correlated with an increase in phenolic off-flavors in wine. The volatile phenols causing the olfactory defect result from the successive decarboxylation and reduction of hydroxycinnamic acids that are normal components of red wines. The growth of B. bruxellensis is preventable by adding sulfur dioxide (SO(2)), with variable effectiveness. Moreover, it was hypothesized that SO(2) was responsible for the entry of B. bruxellensis into a viable but non-culturable (VBNC) state. The aim of this project was to investigate the effects of SO(2) on the remaining enzyme activities of B. bruxellensis populations according to their viability and cultivability, focusing on the hydroxycinnamate decarboxylase enzyme, the first enzyme needed, rather than the metabolites produced. Enzyme activity was determined both in cell-free extracts and resting cells after various SO(2) treatments in synthetic media. After slight sulfiting (around 50 mg/L total SO(2)), the yeasts had lost part of their enzyme activity but not their cultivability. At higher doses (at least 75 mg/L total SO(2)) the majority of yeasts had lost their cultivability but still retained part of their enzyme activity. These results suggested that non culturable cells retained some enzyme activity. Copyright © 2012 Elsevier Ltd. All rights reserved.

A comparison of maximal bioenergetic enzyme activities obtained with commonly used homogenization techniques.

PubMed

Grace, M; Fletcher, L; Powers, S K; Hughes, M; Coombes, J

1996-12-01

Homogenization of tissue for analysis of bioenergetic enzyme activities is a common practice in studies examining metabolic properties of skeletal muscle adaptation to disease, aging, inactivity or exercise. While numerous homogenization techniques are in use today, limited information exists concerning the efficacy of specific homogenization protocols. Therefore, the purpose of this study was to compare the efficacy of four commonly used approaches to homogenizing skeletal muscle for analysis of bioenergetic enzyme activity. The maximal enzyme activity (Vmax) of citrate synthase (CS) and lactate dehydrogenase (LDH) were measured from homogenous muscle samples (N = 48 per homogenization technique) and used as indicators to determine which protocol had the highest efficacy. The homogenization techniques were: (1) glass-on-glass pestle; (2) a combination of a mechanical blender and a teflon pestle (Potter-Elvehjem); (3) a combination of the mechanical blender and a biological detergent; and (4) the combined use of a mechanical blender and a sonicator. The glass-on-glass pestle homogenization protocol produced significantly higher (P < 0.05) enzyme activities compared to all other protocols for both enzymes. Of the four protocols examined, the data demonstrate that the glass-on-glass pestle homogenization protocol is the technique of choice for studying bioenergetic enzyme activity in skeletal muscle.

The inhibitory effect of convulsant agents on the enzyme in brain which inactivates nerveside.

PubMed

Toh, C C

1969-07-01

1. An enzyme which can be extracted from brain inactivates nerveside in the optimum pH range 5.8-7.0.2. The polybasic acids trypan blue and its analogue trypan red, bromphenol blue and its analogue bromthymol blue at concentrations of 0.22 mM and ethylenediaminetetra-acetic acid (EDTA) at a concentration of 1 mM are strong inhibitors of the enzyme.3. Penicillin which is a monobasic carboxylic acid also inhibits the enzyme but only if concentrations as high as 3.6 mM are used. The antibiotic streptomycin which is a basic substance does not inhibit the enzyme.4. Caffeine at a concentration of 7.2 mM only weakly inhibits the enzyme.5. Chymotrypsin and wheat germ acid phosphatase also inactivate nerveside at pH 5.9 and are inhibited by the acidic dyes and penicillin. EDTA inhibits wheat germ phosphatase but activates chymotrypsin.6. Inactivation of nerveside by the brain enzyme and by wheat germ phosphatase is different from the action of chymotrypsin. Nerveside solutions incubated with chymotrypsin completely lose all biological activity whereas if incubation is carried out with either the brain enzyme or wheat germ acid phosphatase a residual biological activity remains even when the concentration of these two enzymes is increased. This residual biological activity is due to a peptide as it is destroyed by chymotrypsin.7. The manner in which nerveside is inactivated by the brain enzyme is uncertain as the preparation of the latter contained phosphodiesterase and protease activities which were similarly inhibited by the acid dyes, penicillin and EDTA.8. Pentylenetetrazole, picrotoxin, strychnine and tetanus toxin do not inhibit the brain enzyme.9. The nerveside-inactivating enzyme is not identical with the Substance P-inactivating enzyme in brain as the former is inhibited by EDTA while the latter is not.

Changes of Serum Angiotensin-Converting Enzyme Activity During Treatment of Patients with Graves’ Disease*

PubMed Central

Lee, Dong Soo; Chung, June-Key; Cho, Bo Youn; Koh, Chang-Soon; Lee, Munho

1986-01-01

Serum angiotensin-converting enzyme activity was measured spectrophotometrically, and serum thyrotropin-binding-inhibitory immunoglobulin (TBII) activity was measured by radioreceptor assay in normal subjects and in patients with Graves’ disease serially before and during treatment, and these activities were compared with each other and with thyroid hormone levels in various thyroid functional status. Correlation between serum angiotensin-converting enzyme activity and serum thyroid hormone level was pursued with relation to the changes of thyroid functional status in patients with Graves’ disease during treatment. Serum angiotensin-converting enzyme activity was significantly elevated in patients with hyperthyroid Graves’ disease before the start of treatment (35 ± 13 nmol/min/ml, n=50), and not in patients with Graves’ disease, euthyroid state during treatment with antithyroid drugs or radioactive iodine (23 ± 9 nmol/min/ml, n=12), but decreased significantly in patients with Graves’ disease, hypothyroid state transiently during treatment (15 ± 4 nmol/min/ml, n=12), respectively in comparison with normal control subjects. Serum angiotensin-converting enzyme activity was positively correlated with the log value of serum T3 concentration (r=0.62, p<0.001, n=95), and with the log value of free thyroxine index (r=0.66, p<0.001, n=91) but not statistically significantly with serum TBII activity. Serum angiotensin-converting enzyme activity was followed in 11 patients with initially increased activity and the activity decreased in proportion to serum thyroid hormone level during treatment, irrespective of treatment modality. It is suggested that thyroid hormones play a role in the increase and decrease of serum angiotensin-converting enzyme activity directly or indirectly influencing the peripheral tissues (probably reticuloendothelial cells or peripheral endothelial cells) in patients with Graves’ disease. PMID:15759385

Processing of poultry feathers by alkaline keratin hydrolyzing enzyme from Serratia sp. HPC 1383.

PubMed

Khardenavis, Anshuman A; Kapley, Atya; Purohit, Hemant J

2009-04-01

The present study describes the production and characterization of a feather hydrolyzing enzyme by Serratia sp. HPC 1383 isolated from tannery sludge, which was identified by the ability to form clear zones around colonies on milk agar plates. The proteolytic activity was expressed in terms of the micromoles of tyrosine released from substrate casein per ml per min (U/mL min). Induction of the inoculum with protein was essential to stimulate higher activity of the enzyme, with 0.03% feathermeal in the inoculum resulting in increased enzyme activity (45U/mL) that further increased to 90U/mL when 3d old inoculum was used. The highest enzyme activity, 130U/mL, was observed in the presence of 0.2% yeast extract. The optimum assay temperature and pH for the enzyme were found to be 60 degrees C and 10.0, respectively. The enzyme had a half-life of 10min at 60 degrees C, which improved slightly to 18min in presence of 1mM Ca(2+). Inhibition of the enzyme by phenylmethyl sulfonyl fluoride (PMSF) indicated that the enzyme was a serine protease. The enzyme was also partially inhibited (39%) by the reducing agent beta-mercaptoethanol and by divalent metal ions such as Zn(2+) (41% inhibition). However, Ca(2+) and Fe(2+) resulted in increases in enzyme activity of 15% and 26%, respectively. The kinetic constants of the keratinase were found to be 3.84 microM (K(m)) and 108.7 microM/mLmin (V(max)). These results suggest that this extracellular keratinase may be a useful alternative and eco-friendly route for handling the abundant amount of waste feathers or for applications in other industrial processes.

Accurate Detection of Adenylation Domain Functions in Nonribosomal Peptide Synthetases by an Enzyme-linked Immunosorbent Assay System Using Active Site-directed Probes for Adenylation Domains.

PubMed

Ishikawa, Fumihiro; Miyamoto, Kengo; Konno, Sho; Kasai, Shota; Kakeya, Hideaki

2015-12-18

A significant gap exists between protein engineering and enzymes used for the biosynthesis of natural products, largely because there is a paucity of strategies that rapidly detect active-site phenotypes of the enzymes with desired activities. Herein, we describe a proof-of-concept study of an enzyme-linked immunosorbent assay (ELISA) system for the adenylation (A) domains in nonribosomal peptide synthetases (NRPSs) using a combination of active site-directed probes coupled to a 5'-O-N-(aminoacyl)sulfamoyladenosine scaffold with a biotin functionality that immobilizes probe molecules onto a streptavidin-coated solid support. The recombinant NRPSs have a C-terminal His-tag motif that is targeted by an anti-6×His mouse antibody as the primary antibody and a horseradish peroxidase-linked goat antimouse antibody as the secondary antibody. These probes can selectively capture the cognate A domains by ligand-directed targeting. In addition, the ELISA technique detected A domains in the crude cell-free homogenates from the Escherichia coli expression systems. When coupled with a chromogenic substrate, the antibody-based ELISA technique can visualize probe-protein binding interactions, which provides accurate readouts of the A-domain functions in NRPS enzymes. To assess the ELISA-based engineering of the A domains of NRPSs, we reprogramed 2,3-dihydroxybenzoic acid (DHB)-activating enzyme EntE toward salicylic acid (Sal)-activating enzymes and investigated a correlation between binding properties for probe molecules and enzyme catalysts. We generated a mutant of EntE that displayed negligible loss in the kcat/Km value with the noncognate substrate Sal and a corresponding 48-fold decrease in the kcat/Km value with the cognate substrate DHB. The resulting 26-fold switch in substrate specificity was achieved by the replacement of a Ser residue in the active site of EntE with a Cys toward the nonribosomal codes of Sal-activating enzymes. Bringing a laboratory ELISA technique and adenylating enzymes together using a combination of active site-directed probes for the A domains in NRPSs should accelerate both the functional characterization and manipulation of the A domains in NRPSs.

Induction of antioxidant enzyme activity and lipid peroxidation level in ion-beam-bombarded rice seeds

NASA Astrophysics Data System (ADS)

Semsang, Nuananong; Yu, LiangDeng

2013-07-01

Low-energy ion beam bombardment has been used to mutate a wide variety of plant species. To explore the indirect effects of low-energy ion beam on biological damage due to the free radical production in plant cells, the increase in antioxidant enzyme activities and lipid peroxidation level was investigated in ion-bombarded rice seeds. Local rice seeds were bombarded with nitrogen or argon ion beams at energies of 29-60 keV and ion fluences of 1 × 1016 ions cm-2. The activities of the antioxidant enzymes; superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), glutathione reductase (GR), glutathione S-transferase (GST) and lipid peroxidation level were assayed in the germinated rice seeds after ion bombardment. The results showed most of the enzyme activities and lipid peroxidation levels in both the argon and nitrogen bombarded samples were higher than those in the natural control. N-ion bombardment could induce higher levels of antioxidant enzyme activities in the rice samples than the Ar-ion bombardment. Additional effects due to the vacuum condition were found to affect activities of some antioxidant enzymes and lipid peroxidation level. This study demonstrates that ion beam bombardment and vacuum condition could induce the antioxidant enzyme activity and lipid peroxidation level which might be due to free radical production in the bombarded rice seeds.

Cajanus cajan Linn. (Leguminosae) prevents alcohol-induced rat liver damage and augments cytoprotective function.

PubMed

Kundu, Rakesh; Dasgupta, Suman; Biswas, Anindita; Bhattacharya, Anirban; Pal, Bikas C; Bandyopadhyay, Debashis; Bhattacharya, Shelley; Bhattacharya, Samir

2008-08-13

Cajanus cajan Linn. (Leguminosae) is a nontoxic edible herb, widely used in Indian folk medicine for the prevention of various liver disorders. In the present study we have demonstrated that methanol-aqueous fraction (MAF2) of Cajanus cajan leaf extract could prevent the chronically treated alcohol induced rat liver damage. Chronic doses of alcohol (3.7 g/ kg) orally administered to rats for 28 days and liver function marker enzymes such as GPT, GOT, ALP and anti-oxidant enzyme activities were determined. Effect of MAF2 at a dose of 50mg/kg body weight on alcohol treated rats was noted. Alcohol effected significant increase in liver marker enzyme activities and reduced the activities of anti-oxidant enzymes. Co-administration of MAF2 reversed the liver damage due to alcohol; it decreased the activities of liver marker enzymes and augmented antioxidant enzyme activities. We also demonstrate significant decrease of the phase II detoxifying enzyme, UDP-glucuronosyl transferase (UGT) activity along with a three- and two-fold decrease of UGT2B gene and protein expression respectively. MAF2 co-administration normalized UGT activity and revived the expression of UGT2B with a concomitant expression and nuclear translocation of Nrf2, a transcription factor that regulates the expression of many cytoprotective genes. Cajanus cajan extract therefore shows a promise in therapeutic use in alcohol induced liver dysfunction.

Effect of ethyleneoxide groups of anionic surfactants on lipase activity.

PubMed

Magalhães, Solange S; Alves, Luís; Sebastião, Marco; Medronho, Bruno; Almeida, Zaida L; Faria, Tiago Q; Brito, Rui M M; Moreno, Maria J; Antunes, Filipe E

2016-09-01

The use of enzymes in laundry and dish detergent products is growing. Such tendency implies dedicated studies to understand surfactant-enzyme interactions. The interactions between surfactants and enzymes and their impact on the catalytic efficiency represent a central problem and were here evaluated using circular dichroism, dynamic light scattering, and enzyme activity determinations. This work focuses on this key issue by evaluating the role of the ethyleneoxide (EO) groups of anionic surfactants on the structure and activity of a commercial lipase, and by focusing on the protein/surfactant interactions at a molecular level. The conformational changes and enzymatic activity of the protein were evaluated in the presence of sodium dodecyl sulfate (SDS also denoted as SLE 0 S) and of sodium lauryl ether sulfate with two EO units (SLE 2 S). The results strongly suggest that the presence of EO units in the surfactant polar headgroup determines the stability and the activity of the enzyme. While SDS promotes enzyme denaturation and consequent loss of activity, SLE 2 S preserves the enzyme structure and activity. The data further highlights that the electrostatic interactions among the protein groups are changed by the presence of the adsorbed anionic surfactants being such absorption mainly driven by hydrophobic interactions. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1276-1282, 2016. © 2016 American Institute of Chemical Engineers.

[Extracellular proteolytic enzymes of Azospirillum brasilensis strain Sp7 and regulation of their activity by a homologous lectin].

PubMed

Chernyshova, M P; Alen'kina, S A; Nikitina, V E; Ignatov, V V

2005-01-01

It was found that Azospirillum brasilensis strain Sp7 is able to produce extracellular proteolytic enzymes. The enzymes were active within a broad range of pH values, with two peaks of activity being located in the acid and alkaline pH areas; required calcium ions; and exhibited substrate specificity with respect to azogelatin. Zymography allowed at least four proteolytic enzymes with molecular weights of 32, 45, 52, and 174 kDa to be detected in A. brasilense Sp7 culture liquid. It was shown that the lectin from A. brasilense Sp7 can inhibit proteolytic enzymes.

Studies of a Halophilic NADH Dehydrogenase. 1: Purification and Properties of the Enzyme

NASA Technical Reports Server (NTRS)

Hochstein, Lawrence I.; Dalton, Bonnie P.

1973-01-01

An NADH dehydrogenase obtained from an extremely halophilic bacterium was purified 570-fold by a combination of gel filtration, chromatography on hydroxyapatite, and ion-exchange chromatography on QAE-Sephadex. The purified enzyme appeared to be FAD-linked and bad an apparent molecular weight of 64000. Even though enzyme activity was stimulated by NaCl, considerable activity (430 % of the maximum activity observed in the presence of 2.5 M NaCl) was observed in the absence of added NaCl. The enzyme was unstable when incubated in solutions of low ionic strength. The presence of NADH enhanced the stability of the enzyme.

Biomedical Applications of Enzymes From Marine Actinobacteria.

PubMed

Kamala, K; Sivaperumal, P

Marine microbial enzyme technologies have progressed significantly in the last few decades for different applications. Among the various microorganisms, marine actinobacterial enzymes have significant active properties, which could allow them to be biocatalysts with tremendous bioactive metabolites. Moreover, marine actinobacteria have been considered as biofactories, since their enzymes fulfill biomedical and industrial needs. In this chapter, the marine actinobacteria and their enzymes' uses in biological activities and biomedical applications are described. © 2017 Elsevier Inc. All rights reserved.

A thermodynamic and theoretical view for enzyme regulation.

PubMed

Zhao, Qinyi

2015-01-01

Precise regulation is fundamental to the proper functioning of enzymes in a cell. Current opinions about this, such as allosteric regulation and dynamic contribution to enzyme regulation, are experimental models and substantially empirical. Here we proposed a theoretical and thermodynamic model of enzyme regulation. The main idea is that enzyme regulation is processed via the regulation of abundance of active conformation in the reaction buffer. The theoretical foundation, experimental evidence, and experimental criteria to test our model are discussed and reviewed. We conclude that basic principles of enzyme regulation are laws of protein thermodynamics and it can be analyzed using the concept of distribution curve of active conformations of enzymes.

Proteomic analyses for profiling regulated proteins/enzymes by Fucus vesiculosus fucoidan in B16 melanoma cells: A combination of enzyme kinetics functional study.

PubMed

Wang, Zhi-Jiang; Zheng, Li; Yang, Jun-Mo; Kang, Yani; Park, Yong-Doo

2018-06-01

Fucoidans are complex sulfated polysaccharides that have a wide range of biological activities. Previously, we reported the various effects of Fucus vesiculosus fucoidan on tyrosinase and B16 melanoma cells. In this study, to identify fucoidan-targeted proteins in B16 melanoma cells, we performed a proteomics study and integrated enzyme kinetics. We detected 19 candidate proteins dysregulated by fucoidan treatment. Among the probed proteins, the enzyme kinetics of two candidate enzymes, namely lactate dehydrogenase (LDH) as an upregulated protein and superoxide dismutase (SOD) as a downregulated enzyme, were determined. The enzyme kinetics results showed that Fucus vesiculosus fucoidan significantly inhibited LDH catalytic function while it did not affect SOD activity even at a high dose, while only slightly decreased activity (up to 10%) at a low dose. Based on our previous and present observations, fucoidan could inhibit B16 melanoma cells growth via regulating proteins/enzymes expression levels such as LDH and SOD known as cell survival biomarkers. Interestingly, both expression level and enzyme catalytic activity of LDH were regulated by fucoidan, which could directly induce the apoptotic effect on B16 melanoma cells along with SOD downregulation. This study highlights how combining proteomics with enzyme kinetics can yield valuable insights into fucoidan targets. Copyright © 2018 Elsevier B.V. All rights reserved.

Expression and characterization of thermostable glycogen branching enzyme from Geobacillus mahadia Geo-05.

PubMed

Mohtar, Nur Syazwani; Abdul Rahman, Mohd Basyaruddin; Raja Abd Rahman, Raja Noor Zaliha; Leow, Thean Chor; Salleh, Abu Bakar; Mat Isa, Mohd Noor

2016-01-01

The glycogen branching enzyme (EC 2.4.1.18), which catalyses the formation of α -1,6-glycosidic branch points in glycogen structure, is often used to enhance the nutritional value and quality of food and beverages. In order to be applicable in industries, enzymes that are stable and active at high temperature are much desired. Using genome mining, the nucleotide sequence of the branching enzyme gene ( glgB ) was extracted from the Geobacillus mahadia Geo-05 genome sequence provided by the Malaysia Genome Institute. The size of the gene is 2013 bp, and the theoretical molecular weight of the protein is 78.43 kDa. The gene sequence was then used to predict the thermostability, function and the three dimensional structure of the enzyme. The gene was cloned and overexpressed in E. coli to verify the predicted result experimentally. The purified enzyme was used to study the effect of temperature and pH on enzyme activity and stability, and the inhibitory effect by metal ion on enzyme activity. This thermostable glycogen branching enzyme was found to be most active at 55 °C, and the half-life at 60 °C and 70 °C was 24 h and 5 h, respectively. From this research, a thermostable glycogen branching enzyme was successfully isolated from Geobacillus mahadia Geo-05 by genome mining together with molecular biology technique.

Biomimicry enhances sequential reactions of tethered glycolytic enzymes, TPI and GAPDHS.

PubMed

Mukai, Chinatsu; Gao, Lizeng; Bergkvist, Magnus; Nelson, Jacquelyn L; Hinchman, Meleana M; Travis, Alexander J

2013-01-01

Maintaining activity of enzymes tethered to solid interfaces remains a major challenge in developing hybrid organic-inorganic devices. In nature, mammalian spermatozoa have overcome this design challenge by having glycolytic enzymes with specialized targeting domains that enable them to function while tethered to a cytoskeletal element. As a step toward designing a hybrid organic-inorganic ATP-generating system, we implemented a biomimetic site-specific immobilization strategy to tether two glycolytic enzymes representing different functional enzyme families: triose phosphoisomerase (TPI; an isomerase) and glyceraldehyde 3-phosphate dehydrogenase (GAPDHS; an oxidoreductase). We then evaluated the activities of these enzymes in comparison to when they were tethered via classical carboxyl-amine crosslinking. Both enzymes show similar surface binding regardless of immobilization method. Remarkably, specific activities for both enzymes were significantly higher when tethered using the biomimetic, site-specific immobilization approach. Using this biomimetic approach, we tethered both enzymes to a single surface and demonstrated their function in series in both forward and reverse directions. Again, the activities in series were significantly higher in both directions when the enzymes were coupled using this biomimetic approach versus carboxyl-amine binding. Our results suggest that biomimetic, site-specific immobilization can provide important functional advantages over chemically specific, but non-oriented attachment, an important strategic insight given the growing interest in recapitulating entire biological pathways on hybrid organic-inorganic devices.

Biomimicry Enhances Sequential Reactions of Tethered Glycolytic Enzymes, TPI and GAPDHS

PubMed Central

Mukai, Chinatsu; Gao, Lizeng; Bergkvist, Magnus; Nelson, Jacquelyn L.; Hinchman, Meleana M.; Travis, Alexander J.

2013-01-01

Maintaining activity of enzymes tethered to solid interfaces remains a major challenge in developing hybrid organic-inorganic devices. In nature, mammalian spermatozoa have overcome this design challenge by having glycolytic enzymes with specialized targeting domains that enable them to function while tethered to a cytoskeletal element. As a step toward designing a hybrid organic-inorganic ATP-generating system, we implemented a biomimetic site-specific immobilization strategy to tether two glycolytic enzymes representing different functional enzyme families: triose phosphoisomerase (TPI; an isomerase) and glyceraldehyde 3-phosphate dehydrogenase (GAPDHS; an oxidoreductase). We then evaluated the activities of these enzymes in comparison to when they were tethered via classical carboxyl-amine crosslinking. Both enzymes show similar surface binding regardless of immobilization method. Remarkably, specific activities for both enzymes were significantly higher when tethered using the biomimetic, site-specific immobilization approach. Using this biomimetic approach, we tethered both enzymes to a single surface and demonstrated their function in series in both forward and reverse directions. Again, the activities in series were significantly higher in both directions when the enzymes were coupled using this biomimetic approach versus carboxyl-amine binding. Our results suggest that biomimetic, site-specific immobilization can provide important functional advantages over chemically specific, but non-oriented attachment, an important strategic insight given the growing interest in recapitulating entire biological pathways on hybrid organic-inorganic devices. PMID:23626684

Compounds from silicones alter enzyme activity in curing barnacle glue and model enzymes.

PubMed

Rittschof, Daniel; Orihuela, Beatriz; Harder, Tilmann; Stafslien, Shane; Chisholm, Bret; Dickinson, Gary H

2011-02-17

Attachment strength of fouling organisms on silicone coatings is low. We hypothesized that low attachment strength on silicones is, in part, due to the interaction of surface available components with natural glues. Components could alter curing of glues through bulk changes or specifically through altered enzyme activity. GC-MS analysis of silicone coatings showed surface-available siloxanes when the coatings were gently rubbed with a cotton swab for 15 seconds or given a 30 second rinse with methanol. Mixtures of compounds were found on 2 commercial and 8 model silicone coatings. The hypothesis that silicone components alter glue curing enzymes was tested with curing barnacle glue and with commercial enzymes. In our model, barnacle glue curing involves trypsin-like serine protease(s), which activate enzymes and structural proteins, and a transglutaminase which cross-links glue proteins. Transglutaminase activity was significantly altered upon exposure of curing glue from individual barnacles to silicone eluates. Activity of purified trypsin and, to a greater extent, transglutaminase was significantly altered by relevant concentrations of silicone polymer constituents. Surface-associated silicone compounds can disrupt glue curing and alter enzyme properties. Altered curing of natural glues has potential in fouling management.

Compounds from Silicones Alter Enzyme Activity in Curing Barnacle Glue and Model Enzymes

PubMed Central

Rittschof, Daniel; Orihuela, Beatriz; Harder, Tilmann; Stafslien, Shane; Chisholm, Bret; Dickinson, Gary H.

2011-01-01

Background Attachment strength of fouling organisms on silicone coatings is low. We hypothesized that low attachment strength on silicones is, in part, due to the interaction of surface available components with natural glues. Components could alter curing of glues through bulk changes or specifically through altered enzyme activity. Methodology/Principal Findings GC-MS analysis of silicone coatings showed surface-available siloxanes when the coatings were gently rubbed with a cotton swab for 15 seconds or given a 30 second rinse with methanol. Mixtures of compounds were found on 2 commercial and 8 model silicone coatings. The hypothesis that silicone components alter glue curing enzymes was tested with curing barnacle glue and with commercial enzymes. In our model, barnacle glue curing involves trypsin-like serine protease(s), which activate enzymes and structural proteins, and a transglutaminase which cross-links glue proteins. Transglutaminase activity was significantly altered upon exposure of curing glue from individual barnacles to silicone eluates. Activity of purified trypsin and, to a greater extent, transglutaminase was significantly altered by relevant concentrations of silicone polymer constituents. Conclusions/Significance Surface-associated silicone compounds can disrupt glue curing and alter enzyme properties. Altered curing of natural glues has potential in fouling management. PMID:21379573

Altering lipase activity and enantioselectivity in organic media using organo-soluble bases: Implication for rate-limiting proton transfer in acylation step.

PubMed

Chen, Chun-Chi; Chen, Teh-Liang; Tsai, Shau-Wei

2006-06-05

With the hydrolytic resolution of (R,S)-naproxen 2,2,2-trifluoroethyl esters via a partially purified papaya lipase (PCPL) in water-saturated isooctane as the model system, the enzyme activity, and enantioselectivty is altered by adding a variety of organo-soluble bases that act as either enzyme activators (i.e., TEA, MP, TOA, DPA, PY, and DMA) or enzyme inhibitors (i.e., PDP, DMAP, and PP). Triethylamine (TEA) is selected as the best enzyme activator as 2.24-fold increase of the initial rate for the (S)-ester is obtained when adding 120 mM of the base. By using an expanded Michaelis-Menten mechanism for the acylation step, the kinetic analysis indicates that the proton transfer for the breakdown of tetrahedral intermediates to acyl-enzyme intermediates is the rate-limiting step, or more sensitive than that for the formation of tetrahedral intermediates when the enzyme activators of different pKa are added. However, no correlation for the proton transfers in the acylation step is found when adding the bases acting as enzyme deactivators. Copyright 2006 Wiley Periodicals, Inc.

In Situ Observation of Chymotrypsin Catalytic Activity Change Actuated by Nonheating Low-Frequency Magnetic Field.

PubMed

Efremova, Maria V; Veselov, Maxim M; Barulin, Alexander V; Gribanovsky, Sergey L; Le-Deygen, Irina M; Uporov, Igor V; Kudryashova, Elena V; Sokolsky-Papkov, Marina; Majouga, Alexander G; Golovin, Yuri I; Kabanov, Alexander V; Klyachko, Natalia L

2018-04-24

Magnetomechanical modulation of biochemical processes is a promising instrument for bioengineering and nanomedicine. This work demonstrates two approaches to control activity of an enzyme, α-chymotrypsin immobilized on the surface of gold-coated magnetite magnetic nanoparticles (GM-MNPs) using a nonheating low-frequency magnetic field (LF MF). The measurement of the enzyme reaction rate was carried out in situ during exposure to the magnetic field. The first approach involves α-chymotrypsin-GM-MNPs conjugates, in which the enzyme undergoes mechanical deformations with the reorientation of the MNPs under LF MF (16-410 Hz frequency, 88 mT flux density). Such mechanical deformations result in conformational changes in α-chymotrypsin structure, as confirmed by infrared spectroscopy and molecular modeling, and lead to a 63% decrease of enzyme initial activity. The second approach involves an α-chymotrypsin-GM-MNPs/trypsin inhibitor-GM-MNPs complex, in which the activity of the enzyme is partially inhibited. In this case the reorientation of MNPs in the field leads to disruption of the enzyme-inhibitor complex and an almost 2-fold increase of enzyme activity. The results further demonstrate the utility of magnetomechanical actuation at the nanoscale for the remote modulation of biochemical reactions.

Identification of a botulinum C3-like enzyme in bovine brain that catalyzes ADP-ribosylation of GTP-binding proteins.

PubMed

Maehama, T; Takahashi, K; Ohoka, Y; Ohtsuka, T; Ui, M; Katada, T

1991-06-05

A novel enzyme activity was found in bovine brain cytosol that transfers the ADP-ribosyl moiety of NAD to proteins with Mr values of 22,000 and 25,000. The substrates were the same GTP-binding proteins serving as the substrate of an ADP-ribosyltransferase C3 which was produced by a type C strain of Clostridium botulinum. The brain enzyme was partially purified from the cytosol and had a molecular mass of approximately 20,000 on a gel filtration column. The brain endogenous enzyme displayed unique properties similar to those observed with botulinum C3 enzyme. The enzyme activity was markedly stimulated by a protein factor that had been initially found in the cytosol as an activator for botulinum C3-catalyzed ADP-ribosylation (Ohtsuka, T., Nagata, K., Iiri, T., Nozawa, Y., Ueno, K., Ui, M., and Katada, T. (1989) J. Biol. Chem. 264, 15000-15005). The activity of the brain enzyme was also affected by certain types of detergents or phospholipids. The substrate of the brain enzyme was specific for GTP-binding proteins serving as the substrate of botulinum C3 enzyme; the alpha-subunits of trimeric GTP-binding proteins which served as the substrate of cholera or pertussis toxin were not ADP-ribosylated by the endogenous enzyme. Thus, this is the first report showing an endogenous enzyme in mammalian cells that catalyzes ADP-ribosylation of small molecular weight GTP-binding proteins.

Directed evolution of new and improved enzyme functions using an evolutionary intermediate and multidirectional search.

PubMed

Porter, Joanne L; Boon, Priscilla L S; Murray, Tracy P; Huber, Thomas; Collyer, Charles A; Ollis, David L

2015-02-20

The ease with which enzymes can be adapted from their native roles and engineered to function specifically for industrial or commercial applications is crucial to enabling enzyme technology to advance beyond its current state. Directed evolution is a powerful tool for engineering enzymes with improved physical and catalytic properties and can be used to evolve enzymes where lack of structural information may thwart the use of rational design. In this study, we take the versatile and diverse α/β hydrolase fold framework, in the form of dienelactone hydrolase, and evolve it over three unique sequential evolutions with a total of 14 rounds of screening to generate a series of enzyme variants. The native enzyme has a low level of promiscuous activity toward p-nitrophenyl acetate but almost undetectable activity toward larger p-nitrophenyl esters. Using p-nitrophenyl acetate as an evolutionary intermediate, we have generated variants with altered specificity and catalytic activity up to 3 orders of magnitude higher than the native enzyme toward the larger nonphysiological p-nitrophenyl ester substrates. Several variants also possess increased stability resulting from the multidimensional approach to screening. Crystal structure analysis and substrate docking show how the enzyme active site changes over the course of the evolutions as either a direct or an indirect result of mutations.

Transformation of cyclodextrin glucanotransferase (CGTase) from aqueous suspension to fine solid particles via electrospraying.

PubMed

Saallah, Suryani; Naim, M Nazli; Mokhtar, Mohd Noriznan; Abu Bakar, Noor Fitrah; Gen, Masao; Lenggoro, I Wuled

2014-10-01

In this study, the potential of electrohydrodynamic atomization or electrospraying to produce nanometer-order CGTase particles from aqueous suspension was demonstrated. CGTase enzyme was prepared in acetate buffer solution (1% v/v), followed by electrospraying in stable Taylor cone-jet mode. The deposits were collected on aluminium foil (collector) at variable distances from the tip of spraying needle, ranging from 10 to 25 cm. The Coulomb fission that occurs during electrospraying process successfully transformed the enzyme to the solid state without any functional group deterioration. The functional group verification was conducted by FTIR analysis. Comparison between the deposit and the as-received enzyme in dry state indicates almost identical spectra. By increasing the distance of the collector from the needle tip, the average particle size of the solidified enzyme was reduced from 200±117 nm to 75±34 nm. The average particle sizes produced from the droplet fission were in agreement with the scaling law models. Enzyme activity analysis showed that the enzyme retained its initial activity after the electrospraying process. The enzyme particles collected at the longest distance (25 cm) demonstrated the highest enzyme activity, which indicates that the activity was controlled by the enzyme particle size. Copyright © 2014 Elsevier Inc. All rights reserved.

Human recombinant soluble guanylyl cyclase: expression, purification, and regulation

NASA Technical Reports Server (NTRS)

Lee, Y. C.; Martin, E.; Murad, F.

2000-01-01

The alpha1- and beta1-subunits of human soluble guanylate cyclase (sGC) were coexpressed in the Sf9 cells/baculovirus system. In addition to the native enzyme, constructs with hexahistidine tag at the amino and carboxyl termini of each subunit were coexpressed. This permitted the rapid and efficient purification of active recombinant enzyme on a nickel-affinity column. The enzyme has one heme per heterodimer and was readily activated with the NO donor sodium nitroprusside or 3-(5'-hydroxymethyl-2'furyl)-1-benzyl-indazole (YC-1). Sodium nitroprusside and YC-1 treatment potentiated each other in combination and demonstrated a remarkable 2,200-fold stimulation of the human recombinant sGC. The effects were inhibited with 1H-(1,2, 4)oxadiazole(4,3-a)quinoxalin-1one (ODQ). The kinetics of the recombinant enzyme with respect to GTP was examined. The products of the reaction, cGMP and pyrophosphate, inhibited the enzyme. The extent of inhibition by cGMP depended on the activation state of the enzyme, whereas inhibition by pyrophosphate was not affected by the enzyme state. Both reaction products displayed independent binding and cooperativity with respect to enzyme inhibition. The expression of large quantities of active enzyme will facilitate structural characterization of the protein.

Electrochemistry of metalloproteins: protein film electrochemistry for the study of E. coli [NiFe]-hydrogenase-1.

PubMed

Evans, Rhiannon M; Armstrong, Fraser A

2014-01-01

Protein film electrochemistry is a technique which allows the direct control of redox-active enzymes, providing particularly detailed information on their catalytic properties. The enzyme is deposited onto a working electrode tip, and through control of the applied potential the enzyme activity is monitored as electrical current, allowing for direct study of inherent activity as electrons are transferred to and from the enzyme redox center(s). No mediators are used. Because the only enzyme present in the experiment is bound at the electrode surface, gaseous and liquid phase inhibitors can be introduced and removed whilst the enzyme remains in situ. Potential control means that kinetics and thermodynamics are explored simultaneously; the kinetics of a reaction can be studied as a function of potential. Steady-state catalytic rates are observed directly as current (for a given potential) and non-steady-state rates (such as interconversions between different forms of the enzyme) are observed from the change in current with time. The more active the enzyme, the higher the current and the better the signal-to-noise. In this chapter we outline the practical aspects of PFE for studying electroactive enzymes, using the Escherichia coli [NiFe]-hydrogenase 1 (Hyd-1) as an example.