| 1. |
- Aguilar, Ximena, 1978-, et al.
(författare)
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Interaction Studies of the Human and Arabidopsis thaliana Med25-ACID Proteins with the Herpes Simplex Virus VP16-and Plant-Specific Dreb2a Transcription Factors
- 2014
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 9:5, s. e98575-
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Tidskriftsartikel (refereegranskat)abstract
- Mediator is an evolutionary conserved multi-protein complex present in all eukaryotes. It functions as a transcriptional coregulator by conveying signals from activators and repressors to the RNA polymerase II transcription machinery. The Arabidopsis thaliana Med25 (aMed25) ACtivation Interaction Domain (ACID) interacts with the Dreb2a activator which is involved in plant stress response pathways, while Human Med25-ACID (hMed25) interacts with the herpes simplex virus VP16 activator. Despite low sequence similarity, hMed25-ACID also interacts with the plant-specific Dreb2a transcriptional activator protein. We have used GST pull-down-, surface plasmon resonance-, isothermal titration calorimetry and NMR chemical shift experiments to characterize interactions between Dreb2a and VP16, with the hMed25 and aMed25-ACIDs. We found that VP16 interacts with aMed25-ACID with similar affinity as with hMed25-ACID and that the binding surface on aMed25-ACID overlaps with the binding site for Dreb2a. We also show that the Dreb2a interaction region in hMed25-ACID overlaps with the earlier reported VP16 binding site. In addition, we show that hMed25-ACID/Dreb2a and aMed25-ACID/Dreb2a display similar binding affinities but different binding energetics. Our results therefore indicate that interaction between transcriptional regulators and their target proteins in Mediator are less dependent on the primary sequences in the interaction domains but that these domains fold into similar structures upon interaction.
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| 2. |
- Chaintreau, Alain, et al.
(författare)
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Site-specific C-13 content by quantitative isotopic C-13 nuclear magnetic resonance spectrometry : a pilot inter-laboratory study
- 2013
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Ingår i: Analytica Chimica Acta. - : Elsevier BV. - 0003-2670 .- 1873-4324. ; 788, s. 108-113
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Tidskriftsartikel (refereegranskat)abstract
- Isotopic C-13 NMR spectrometry, which is able to measure intra-molecular C-13 composition, is of emerging demand because of the new information provided by the C-13 site-specific content of a given molecule. A systematic evaluation of instrumental behaviour is of importance to envisage isotopic C-13 NMR as a routine tool. This paper describes the first collaborative study of intra-molecular C-13 composition by NMR. The main goals of the ring test were to establish intra-and inter-variability of the spectrometer response. Eight instruments with different configuration were retained for the exercise on the basis of a qualification test. Reproducibility at the natural abundance of isotopic C-13 NMR was then assessed on vanillin from three different origins associated with specific delta C-13(i) profiles. The standard deviation was, on average, between 0.9 and 1.2 parts per thousand for intra-variability. The highest standard deviation for inter-variability was 2.1%. This is significantly higher than the internal precision but could be considered good in respect of a first ring test on a new analytical method. The standard deviation of delta C-13(i) in vanillin was not homogeneous over the eight carbons, with no trend either for the carbon position or for the configuration of the spectrometer. However, since the repeatability for each instrument was satisfactory, correction factors for each carbon in vanillin could be calculated to harmonize the results.(C) 2013 Elsevier B.V. All rights reserved.
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| 3. |
- Drotz, Stina Harrysson, et al.
(författare)
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Effects of soil organic matter composition on unfrozen water content and heterotrophic CO2 production of frozen soils
- 2010
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Ingår i: Geochimica et Cosmochimica Acta. - : Elsevier Ltd. - 0016-7037 .- 1872-9533. ; 74:8, s. 2281-90
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Tidskriftsartikel (refereegranskat)abstract
- Several recent studies have highlighted the importance of soil organic matter (SOM) mineralization at high latitudes during winter for ecosystem carbon (C) balances, and the ability of the soil to retain unfrozen water at sub-zero temperatures has been shown to be a major determinant of C mineralization rates. Further, SOM is believed to strongly influence the liquid water contents in frozen surface layers of boreal forest soils and tundra, but the mechanisms and specific factors involved are currently unknown. Here we evaluate the effects of the chemical composition of SUM on the amount of unfrozen water, the pore size equivalents in which unfrozen water can exist, and the microbial heterotrophic activity at sub-zero temperatures in boreal forest soils. To do this, we have characterized the chemical composition of SUM in forest soil samples (surface O-horizons) using solid state CP-MAS (cross polarization magic angle spinning) NMR spectroscopy. The acquired information was then used to elucidate the extent to which different fractions of SUM can explain the observed variations in unfrozen water content, pore size equivalents, and biogenic CO2 production rates in the examined soil samples under frozen conditions (-4 degrees C). The data evaluation was done by the use of principal component analysis (PCA) and projections to latent structures by means of partial least square (PLS). We conclude that aromatic, O-aromatic, methoxy/N-alkyl and alkyl C are the major SOM components affecting frozen boreal forest soil's ability to retain unfrozen water and sustain heterotrophic activity (95% confidence level). Our results reveal that solid carbohydrates have a significant negative impact (95% confidence level) on CO2 production in frozen boreal spruce forest soils, in contrast to the positive effects of carbohydrate polymers during unfrozen conditions. We conclude that the hierarchy of environmental factors controlling SOM mineralization changes as soils freeze. The effect of SUM composition on pore size distribution and unfrozen water content has a superior influence on SUM mineralization and hence on heterotrophic CO2 production of frozen soils. (C) 2010 Elsevier Ltd. All rights reserved.
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| 4. |
- Ehlers, Ina, 1984-, et al.
(författare)
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Elucidating Turnover Pathways of Bioactive Small Molecules by Isotopomer Analysis : The Persistent Organic Pollutant DDT
- 2014
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Ingår i: PLOS ONE. - : PLoS ONE. - 1932-6203. ; 9:10, s. e110648-
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Tidskriftsartikel (refereegranskat)abstract
- The persistent organic pollutant DDT (1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane) is still indispensable in the fight againstmalaria, although DDT and related compounds pose toxicological hazards. Technical DDT contains the dichloro congenerDDD (1-chloro-4-[2,2-dichloro-1-(4-chlorophenyl)ethyl]benzene) as by-product, but DDD is also formed by reductive degradation of DDT in the environment. To differentiate between DDD formation pathways, we applied deuterium NMR spectroscopy to measure intramolecular deuterium distributions (2H isotopomer abundances) of DDT and DDD. DDD formed in the technical DDT synthesis was strongly deuterium-enriched at one intramolecular position, which we traced back to 2H/1H fractionation of a chlorination step in the technical synthesis. In contrast, DDD formed by reductive degradation was strongly depleted at the same position, which was due to the incorporation of 2H-depleted hydride equivalents during reductive degradation. Thus, intramolecular isotope distributions give mechanistic information on reaction pathways, and explain a puzzling difference in the whole-molecule 2H/1H ratio between DDT and DDD. In general, our results highlight that intramolecular isotope distributions are essential to interpret whole-molecule isotope ratios. Intramolecular isotope information allows distinguishing pathways of DDD formation, which is important to identify polluters or to assess DDT turnover in the environment. Because intramolecular isotope data directly reflect isotope fractionation of individual chemical reactions, they are broadly applicable to elucidate transformation pathways of smallbioactive molecules in chemistry, physiology and environmental science.
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| 5. |
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| 6. |
- Erhagen, Björn, et al.
(författare)
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Temperature response of litter and soil organic matter decomposition is determined by chemical composition of organic material
- 2013
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Ingår i: Global Change Biology. - : Wiley-Blackwell. - 1354-1013 .- 1365-2486. ; 19:12, s. 3858-3871
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Tidskriftsartikel (refereegranskat)abstract
- The global soil carbon pool is approximately three times larger than the contemporary atmospheric pool, therefore even minor changes to its integrity may have major implications for atmospheric CO2 concentrations. While theory predicts that the chemical composition of organic matter should constitute a master control on the temperature response of its decomposition, this relationship has not yet been fully demonstrated. We used laboratory incubations of forest soil organic matter (SOM) and fresh litter material together with NMR spectroscopy to make this connection between organic chemical composition and temperature sensitivity of decomposition. Temperature response of decomposition in both fresh litter and SOM was directly related to the chemical composition of the constituent organic matter, explaining 90% and 70% of the variance in Q10 in litter and SOM respectively. The Q10 of litter decreased with increasing proportions of aromatic and O-aromatic compounds, and increased with increased contents of alkyl- and O-alkyl carbons. In contrast, in SOM, decomposition was affected only by carbonyl compounds. To reveal why a certain group of organic chemical compounds affected the temperature sensitivity of organic matter decomposition in litter and SOM, a more detailed characterisation of the (13) C aromatic region using Heteronuclear Single Quantum Coherence (HSQC) was conducted. The results revealed considerable differences in the aromatic region between litter and SOM. This suggests that the correlation between chemical composition of organic matter and the temperature response of decomposition differed between litter and SOM. The temperature response of soil decomposition processes can thus be described by the chemical composition of its constituent organic matter, this paves the way for improved ecosystem modelling of biosphere feedbacks under a changing climate.
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| 7. |
- Greule, Markus, et al.
(författare)
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Improved rapid authentication of vanillin using δ13C and δ2H values
- 2010
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Ingår i: European Food Research and Technology. - : Springer Science and Business Media LLC. - 1438-2377 .- 1438-2385. ; 231:6, s. 933-941
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Tidskriftsartikel (refereegranskat)abstract
- Vanilla still remains one of the most important and widely used flavours in the food industry and is also extensively employed by fragrance and pharmaceutical manufacturing companies. Natural vanilla flavour, extracted from the pods of the tropic orchid vanilla, is considerably more expensive than synthetic vanillin. The disparity of prices between natural vanillin and that derived from other sources has given rise to many cases of fraudulent adulteration, and for more than 30 years, strenuous efforts have been made to authenticate sources of vanillin. Stable isotope analysis is one of the most powerful analytical tools to distinguish between natural vanillin and that originating from other sources. Recently, a rapid and precise method for analysis of both δ13C and δ2H values of plant methoxyl groups has been published. Here, we report an application of the method for the control of authenticity of vanillin. Carbon and hydrogen stable isotope values of the vanillin molecule and vanillin methoxyl groups of vanillin samples of different origins including authentic and synthetic samples were measured. The results clearly show that use of this approach provides a rapid and reliable authenticity assessment of vanillin. The technique used for these studies is robust and rapid, involves minimum sample preparation and requires only a small amount of vanillin sample, usually 1 mg for stable carbon and 4 mg for stable hydrogen analysis.
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| 8. |
- Harrysson Drotz, Stina, et al.
(författare)
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Both catabolic and anabolic heterotrophic microbial activity proceed in frozen soils
- 2010
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 107:49, s. 21046-21051
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Tidskriftsartikel (refereegranskat)abstract
- A large proportion of the global soil carbon pool is stored in soils of high-latitude ecosystems in which microbial processes and production of greenhouse gases proceed during the winter months. It has been suggested that microorganisms have limited ability to sequester substrates at temperatures around and below 0 °C and that a metabolic shift to dominance of catabolic processes occurs around these temperatures. However, there are contrary indications that anabolic processes can proceed, because microbial growth has been observed at far lower temperatures. Therefore, we investigated the utilization of the microbial substrate under unfrozen and frozen conditions in a boreal forest soil across a temperature range from -9 °C to +9 °C, by using gas chromatography-isotopic ratio mass spectrometry and (13)C magic-angle spinning NMR spectroscopy to determine microbial turnover and incorporation of (13)C-labeled glucose. Our results conclusively demonstrate that the soil microorganisms maintain both catabolic (CO(2) production) and anabolic (biomass synthesis) processes under frozen conditions and that no significant differences in carbon allocation from [(13)C]glucose into [(13)C]CO(2) and cell organic (13)C-compounds occurred between +9 °C and -4 °C. The only significant metabolic changes detected were increased fluidity of the cell membranes synthesized at frozen conditions and increased production of glycerol in the frozen samples. The finding that the processes in frozen soil are similar to those in unfrozen soil has important implications for our general understanding and conceptualization of soil carbon dynamics in high-latitude ecosystems.
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| 9. |
- Kaffarnik, Stefanie, et al.
(författare)
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Two-Dimensional P-31,H-1 NMR Spectroscopic Profiling of Phospholipids in Cheese and Fish
- 2013
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Ingår i: Journal of Agricultural and Food Chemistry. - : American Chemical Society (ACS). - 0021-8561 .- 1520-5118. ; 61:29, s. 7061-7069
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Tidskriftsartikel (refereegranskat)abstract
- Phospholipids (PLs) comprise an important lipid class in food because of their technological use as emulsifiers and their nutritional value. This study used one-dimensional P-31 NMR and two-dimensional (2D) P-31,H-1 COSY NMR spectroscopy for the determination of the PL composition of cheese and fish after liquid liquid enrichment. This extraction step enabled the identification of 10 PLs in cheese and 9 PLs in fish by 2D P-31,H-1 NMR. Variations in the P-31 shifts indicated differences in the fatty acids attached to the individual PLs. The total PL content in cheese fat and fish oil ranged from 0.3 to 0.4% and from 5 to 12%, respectively. Phosphatidylcholine was the most prominent PL in both matrices (up to 6596). Minor PLs (limit of detection = 4 nmol, i.e. 500 mu L of an 8 mu M solution) were identified in forms of phosphatidic acid, lysophosphatidic acid, and phosphatidylglycerol. Specific cross couplings and H-1 fine structures in the 2D P-31,H-1 NMR spectra proved to be valuable for the assignment and verification of known and uncommon PLs in the samples.
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| 10. |
- Nielsen, Søren B, et al.
(författare)
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The interaction of equine lysozyme : oleic acid complexes with lipid membranes suggests a cargo off-loading mechanism
- 2010
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Ingår i: Journal of Molecular Biology. - : Elsevier. - 0022-2836 .- 1089-8638. ; 398:2, s. 351-361
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Tidskriftsartikel (refereegranskat)abstract
- The normal function of equine lysozyme (EL) is the hydrolysis of peptidoglycan residues of bacterial cell walls. EL is closely related to alpha-lactalbumins with respect to sequence and structure and further possesses the calcium binding site of alpha-lactalbumins. Recently, EL multimeric complexes with oleic acids (ELOA) were shown to possess tinctorial and morphological properties, similar to amyloidal aggregates, and to be cytotoxic. ELOA's interactions with phospholipid membranes appears to be central to its biological action, similar to human alpha-lactalbumin made lethal to tumor cells (HAMLET). Here, we describe the interaction of ELOA with phospholipid membranes. Confocal scanning laser microscopy shows that ELOA, but not native EL, accumulates on the surface of giant unilamellar vesicles, without inducing significant membrane permeability. Quartz crystal microbalance with dissipation (QCM-D) data indicated an essentially non-disruptive binding of ELOA to supported lipid bilayers, leading to formation of highly dissipative and "soft" lipid membrane; at higher concentrations of ELOA, the lipid membrane desorbs from the surface probably as bilayer sheets of vesicles. This membrane rearrangement occurred to a similar extent when free oleic acid (OA) was added, but not when free OA was removed from ELOA by prior incubation with BSA, emphasizing the role of OA in this process. NMR data indicated an equilibrium between free and bound OA which shifts towards free OA as ELOA is progressively diluted indicating that OA is relatively loosely bound. Activity measurements together with fluorescence spectroscopy and circular dichroism suggested a conversion of ELOA toward a more native-like state on interaction with lipid membranes, although complete refolding was not observed. Altogether, these results suggest that ELOA may act as an OA carrier and facilitate OA transfer to the membrane. ELOA's properties illustrate that protein folding variants may possess specific functional properties distinct from the native protein. Abbreviations QCM-D, Quartz crystal microbalance with dissipation; CD, Circular dichroism; EL, equine lysozyme; ELOA, EL complex with oleic acid; OA, oleic acid, CSLM, Confocal scanning laser microscopy, Df, dissipation-frequency.
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