Computer calculation-based quantitative structure-activity relationships for the oxidation of phenol derivatives horseradish peroxidase compound II

 The second-order rate constants for the oxidation of a series of phenol derivatives by horseradish peroxidase compound II were compared to computer-calculated chemical parameters characteristic for this reaction step. The phenol derivatives studied were phenol, 4-chlorophenol, 3-hydroxyphenol, 3-methylphenol, 4-methylphenol, 4-hydroxybenzoate, 4-methoxyphenol and 4-hydroxybenzaldehyde. Assuming a reaction of the phenolic substrates in their non-dissociated, uncharged forms, clear correlations (r = 0.977 and r = 0.905) were obtained between the natural logarithm of the second-order rate constants (ln k _app and ln k ₂ respectively) for their oxidation by compound II and their calculated ionisation potential, i.e. minus the energy of their highest occupied molecular orbital [E(HOMO)]. In addition to this first approach in which the quantitative structure-activity relationship (QSAR) was based on a calculated frontier orbital parameter of the substrate, in a second and third approach the relative heat of formation (ΔΔHF) calculated for the process of one-electron abstraction and H^• abstraction from the phenol derivatives was used as a parameter. Plots of the natural logarithms of the second-order rate constants (k _app and k ₂) for the reaction and the calculated ΔΔHF values for the process of one-electron abstraction also provide clear QSARs with correlation coefficients of –0.968 and –0.926 respectively. Plots of the natural logarithms of the second-order rate constants (k _app and k ₂) for the reaction and the calculated ΔΔHF values for the process of H^• abstraction provide QSARs with correlation coefficients of –0.989 and –0.922 respectively. Since both mechanisms considered, i.e. initial electron abstraction versus initial H^• abstraction, provided clear QSARs, the results could not be used to discriminate between these two possible mechanisms for phenol oxidation by horseradish peroxidase compound II. The computer calculation-based QSARs thus obtained for the oxidation of the various phenol derivatives by compound II from horseradish peroxidase indicate the validity of the approaches investigated, i.e. both the frontier orbital approach and the approach in which the process is described by calculated relative heats of formation. The results also indicate that outcomes from computer calculations on relatively unrelated phenol derivatives can be reliably compared to one another. Furthermore, as the actual oxidation of peroxidase substrates by compound II is known to be the rate-limiting step in the overall catalysis by horseradish peroxidase, the QSARs of the present study may have implications for the differences in the overall rate of substrate oxidation of the phenol derivatives by horseradish peroxidase. Received: 29 March 1996 / Accepted: 17 July 1996     

Nω-Hydroxyamino-α-amino acids as a new class of very strong inhibitors of arginases

 The effects of various compounds bearing an N-OH group such as N-hydroxy-guanidines, amidoximes, and hydroxylamines, on bovine and rat liver arginases was studied. Some of these compounds with an l-α-amino acid function at an appropriate distance from the N-OH group acted as strong competitive liver arginase inhibitors, displaying Ki values between 4 and 150 μM. Two compounds, N ^ε-hydroxy-l-lysine and N ^ω-hydroxy-d,l-indospicine, which exhibited Ki values of 4 and 20 μM (at pH 7.4), were the most potent inhibitors of arginase described to date. The distance between the α-amino acid and N-OH functions appeared to be crucial for potent inhibition of arginase, as N ^δ-hydroxy-l-ornithine, which has one -CH₂ group less than N ^ε-hydroxy-l-lysine, exhibited a 37-fold higher Ki value than N ^ε-hydroxy-l-lysine. Based on these results, a model for the interaction of N ^ω-hydroxyamino-l-α-amino acids with the arginase active site is proposed. This model involves the binding of the N-OH group of the inhibitors to the arginase Mn(II) center and suggests that N ^ε-hydroxy-l-lysine is a good transition state analog of arginase.     

DISRUPTION OF CELL—CELL ADHESION IN THE PRESENCE OF SODIUM BUTYRATE ACTIVATES EXPRESSION OF THE 92kDa TYPE IV COLLAGENASE IN MDCK CELLS

We have investigated the effects of altered cell shape on the regulation of the 92kDa type IV collagenase. In MDCK cells, anti-E-cadherin antibodies alter cell shape by disrupting normal cell—cell contacts, while sodium butyrate causes a marked flattening and spreading of cells. The disruption of cell—cell contacts led to a faint expression of the 92kDa collagenase. This effect was enhanced by sodium butyrate, which by itself did not induce collagenase expression. In contrast, stromelysin expression was not regulated in these conditions. Although mRNA expression was enhanced, the secreted collagenase activity was not altered in these conditions in either cell line. Examination of cytoskeletal and extracellular matrix proteins and cell—cell and cell—matrix adhesion proteins by immunofluorescence and Western blot revealed a disruption of the actin network, tight junctions, and fibronectin deposition by anti E-cadherin antibodies, and alterations in actin, cytokeratin 8, cytokeratin 14, laminin and β1 integrin induced by sodium butyrate. Thus, the induction of collagenase expression in epithelial cells by disrupted cell—cell adhesion and sodium butyrate is associated with changes in cell shape and structure.     

Vegetation heterogeneity and diversity in flat and mountain landscapes of Patagonia (Argentina)

Abstract. We studied floristic and diversity patterns and their environmental controls in two landscapes of contrasting topography in the Patagonian steppe. The analyses were focused on the effects of water availability gradients and landscape configuration on plant species distribution and coexistence. Floristic variation was investigated using Correspondence Analysis. The relationship between floristic and environmental variation was analyzed using Canonical Correspondence Analysis and correlation tests. We explored diversity patterns by relating spatial distance to floristic dissimilarities. The floristic gradient was determined by shrub and grass species and was related to precipitation in the flat area, and to precipitation, elevation and potential radiation in the mountain area. Site species richness increased with water availability in both areas. Mean site species richness and species turnover in space was higher in the mountain than in the flat area. Landscape species richness and floristic gradients were more concentrated in the mountain than in the flat area. In contrast to shrubs and grasses, forb species distributions were uncoordinated and probably independent of any environmental gradient. Our results suggest (1) that landscape configuration affects species composition and diversity through its direct effect on abiotic environmental heterogeneity, and (2) that the environmental controls of the community composition vary depending on the plant functional type considered.     

Global vegetation models: incorporating transient changes to structure and composition

Abstract. We describe an approach for developing a Dynamic Global Vegetation Model (DGVM) that accounts for transient changes in vegetation distribution over a decadal time scale. The DGVM structure is based on a linkage between an equilibrium global vegetation model and smaller scale ecosystem dynamics modules that simulate the rate of vegetation change. Vegetation change is classified into four basic types, based largely on the projected change in above-ground biomass of the vegetation. These four types of change are: (1) dieback of forest, shrubland or grassland; (2) successional replacement within forest, shrubland or grassland; (3) invasion of forest, shrubland or grassland; (4) change in tree/grass ratio. We then propose an approach in which the appropriate ecosystem dynamics module for each type of change is applied and the grid cells of the global model updated accordingly. An approach for accounting for fire, as an example of a disturbance which may strongly influence the rate and spatial pattern of forest dieback, is incorporated. We also discuss data needs for the development, calibration and validation of the model.     

Production and characterization of ligninolytic enzymes ofBjerkandera adusta grown on wood meal/wheat bran culture and production of these enzymes using a rotary-solid fermenter

Manganese peroxidase (MnP) and lignin peroxidase (LiP) were produced by growing a white-rot fungusBjerkandera adusta statically, on a wood meal/wheat bran culture in flasks. MnP and LiP reached their maximum activity after 6 and 19 days of inoculation, respectively. Both MnP and LiP are thought to be important enzymes in lignin biodegradation byB. adusta. Ion exchange chromatography showed thatB. adusta produced a single LiP and a single MnP enzyme in wood meal/wheat bran culture. These enzymes were separated and characterized. The molecular weight of MnP was 46,500 with a pl of 3.9. The molecular weight of LiP was estimated to be 47,000 with a pl of 3.5. Spectral analysis demonstrated that both enzymes are heme proteins. Production of these enzymes was also achieved using a rotarysolid culture fermenter. MnP, LiP and veratryl alcohol oxidase were produced byB. adusta in the fermenter.     

In situ estimation of net CO2 assimilation, photosynthetic electron flow and photorespiration in Turkey oak (Q. cerris L.) leaves: diurnal cycles under different levels of water supply

Diurnal time courses of net CO₂ assimilation rates, stomatal conductance and light-driven electron fluxes were measured in situ on attached leaves of 30-year-old Turkey oak trees (Quercus cerris L.) under natural summer conditions in central Italy. Combined measurements of gas exchange and chlorophyll a fluorescence under low O₂ concentrations allowed the demonstration of a linear relationship between the photochemical efficiency of PSII (fluorescence measurements) and the apparent quantum yield of gross photosynthesis (gas exchange). This relationship was used under normal O₂ to compute total light-driven electron fluxes, and to partition them into fractions used for RuBP carboxylation or RuBP oxygenation. This procedure also yielded an indirect estimate of the rate of photorespiration in vivo. The time courses of light-driven electron flow, net CO₂ assimilation and photorespiration paralleled that of photosynthetic photon flux density, with important afternoon deviations as soon as a severe drought stress occurred, whereas photochemical efficiency and maximal fluorescence underwent large but reversible diurnal decreases. The latter observation indicated the occurrence of a large non-photochemical energy dissipation at PSII. We estimated that less than 60% of the total photosynthetic electron flow was used for carbon assimilation at midday, while about 40% was devoted to photorespiration. The rate of carbon loss by photorespiration (R₁) reached mean levels of 56% of net assimilation rates. The potential application of this technique to analysis of the relative contributions of thermal de-excitation at PSII and photorespiratory carbon recycling in the protection of photosynthesis against stress effects is discussed.     

Models for relating pH measurements in water and calcium chloride for a wide range of pH,soil types and depths

Soil pH is commonly measured in water (pHw) or 0.01 M CaCl₂ (pH_Ca). The need to convert between these methods has led to the publication of linear, quadratic and cubic polynomial relationships for limited suites of soils. Concerns over the applicability of such relationships when mapping a wide range of soils and pH led to the establishment of a database of pH_W and pH_Ca values on each of 7894 samples from soil survey and field experimental sites in Queensland. The relationship between pH_W and pH_Ca across all soils was investigated and preliminary results examining the effect of soil depth and soil type on the relationship are presented.For all soils and depths, a linear regression accounted for 93.2% of the variation but did not predict pH_Ca well at very high or low pH_W values. The inclusion of second and third powers of pH_W accounted for significantly more of the variation (R²=0.94) in pH_Ca and the resultant curve matched the data better at high and low pH.Analysis of surface, sub-surface and subsoil groupings did not reveal any appreciable differences in the relationship between pH_W and pH_Ca attributable to depth. In contrast, differences in the relationship were evident between soil types. Generally, the mildly leached soils had linear relationships, while the weathered soils were distinctly curvilinear at low pH.     

Effects of dark- and light-induced proton gradients in thylakoids on the Q and B thermoluminescence bands

Thermoluminescence experiments have been carried out to study the effect of a transmembrane proton gradient on the recombination properties of the S₂ and S₃ states of the oxygen evolving complex with Q_A ^- and Q_B ^-, the reduced electron acceptors of Photosystem II. We first determined the properties of the S₂Q_A ^- (Q band), S₂Q_B ^- and S₃Q_B ^- (B bands) recombinations in the pH range 5.5 to 9.0, using uncoupled thylakoids. The, a proton gradient was created in the dark, using the ATP-hydrolase function of ATPases, in coupled unfrozen thylakoids. A shift towards low temperature of both Q and B bands was observed to increase with the magnitude of the proton gradient measured by the fluorescence quenching of 9-aminoacridine. This downshift was larger for S₃Q_B ^- than for S₂Q_B ^- and it was suppressed by nigericin, but not by valinomycin. Similar results were obtained when a proton gradient was formed by photosystem I photochemistry. When Photosystem II electron transfer was induced by a flash sequence, the reduction of the plastoquinone pool also contributed to the downshift in the absence of an electron acceptor. In leaves submitted to a flash sequence above 0°C, a downshift was also observed, which was supressed by nigericin infiltration. Thus, thermoluminescence provides direct evidence on the enhancing effect of lumen acidification on the S₃S₂ and S₂S₁ reverse-transitions. Both reduction of the plastoquinone pool and lumen acidification induce a shift of the Q and B bands to lower temperature, with a predominance of lumen acidification in non-freezing, moderate light conditions.Abbreviations 9-AA 9-aminoacridine - E_A activation energy - F₀ constant fluorescence level - F_M maximum fluorescence, when all PS-II centers are closed - F_V variable fluorescence (F_M–F₀) - PS I, PS II Photosystem I, photosystem II - PQ plastoquinone - TL thermoluminescence     

Effects of dehydration on the electron transport of Chlorella. An in vivo fluorescence study

Chlorella was used to study the effects of dehydration on photosynthetic activities. The use of unicellular green algae assured that the extent of dehydration was uniform throughout the whole cell population during the course of desiccation. Changes in the activities of the cells were monitored by measurements of fluorescence induction kinetics. It was found that inhibition of most of the photosynthetic activities started at a similar level of cellular water content. They included CO₂ fixation, photochemical activity of Photosystem II and electron transport through Photosystem I. The blockage of electron flow through Photosystem I was complete and the whole transition occurred within a relative short time of dehydration. On the other hand, the suppression of Photosystem II activity was incomplete and the transition took a longer time of dehydration. Upon rehydration, the inhibition of Photosystem II activity was fully reversible when samples were in the middle of the transition, but was not thereafter. The electron transport through Photosystem I was also reversible during the transition, but was only partially afterward.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - F_m maximum fluorescence yield - F₀ non-variable fluorescence level emitted when all PS II centers are open - F_v variable part of fluorescence - PS photosystem - Q_A primary quinone acceptor of Photosystem II