Ecophysiology and distribution of the endemic leafless spurgeEuphorbia aphylla and the introducedE. tirucalli (Euphorbiaceae,Euphorbia sect.Tirucalli) in the Canary Islands

The leafless spurgeEuphorbia aphylla (Euphorbiaceae), an endemic species restricted to several of the Canary Islands where it inhabits coastal and arid localities, expresses Crassulacean Acid Metabolism (CAM) when it is subjected to summer drought. A flexible CAM is consistent with the general ecology of the species. It is the only member of sect.Tirucalli native to the Canary Islands and is at the north-western edge of the section's biogeographical range. The other members of the section have a paleotropic distribution and are found throughout Africa. Many of them are regarded as obligate CAM plants, includingE. tirucalli which was used as a comparison in ecophysiological experiments examining the response ofE. aphylla to drought and temperature.     

Na/H exchange in cultured epithelial cells from fish gills

Using primary cultures of gill pavement cells from freshwater rainbow trout, a method is described for achieving confluent monolayers of the cells on glass coverslips. A continuous record of intracellular pH was obtained by loading the cells with the pH-sensitive flourescent dye 2,7-bis(2-carboxyethyl)-5(6)-carboxyfluorescein and mounting the coverslips in the flowthrough cuvette of a spectrofluorimeter. Experiments were performed in HEPES-buffered media nominally free of HCO₃. Resting intracellular pH (7.43 at extracellular pH=7.70) was insensitive to the removal of Cl^– or the application of 4-acetamido-4-isothiocyanatostilbene-2,2-disulfonic acid (0.1 mmol·l^–1), but fell by about 0.3 units when Na⁺ was removed or in the presence of amiloride (0.2 mmol·l^–1). Exposure to elevated ammonia (ammonia prepulse; 30 mmol·l^–1 as NH₄Cl for 6–9 min) produced an increase in intracellular pH (to about 8.1) followed by a slow decay, and washout of the pulse caused intracellular pH to fall to about 6.5. Intracellular non-HCO ₃ ^– buffer capacity was about 13.4 slykes. Rapid recovery of intracellular pH from intracellular acidosis induced by ammonia prepulse was inhibited more than 80% in Na⁺-free conditions or in the presence of amiloride (0.2 mmol·l^–1). Neither bafilomycin A₁ (3 mol·l^–1) nor Cl removal altered the intracellular pH recovery rate. The K _m for Na⁺ of the intracellular pH recovery mechanism was 8.3 mmol·l^–1, and the rate constant at V _max was 0.008·s^–1 (equivalent to 5.60 mmol H⁺·l^–1 cell water·min^–1), which was achieved at external Na⁺ levels from 25 to 140 mmol·l^–1. We conclude that intracellular pH in cultured gill pavement cells in HEPES-buffered, HCO ₃ ^– -free media, both at rest and during acidosis, is regulated by a Na⁺/H⁺ antiport and not by anion-dependent mechanisms or a vacuolar H⁺-ATPase.Abbreviations BCECF 2,7-bis(2-carboxyethyl)-5(6)-carboxy-fluorescein - BCECF/AM 2,7-bis(2-carboxyethyl)-5(6)-carboxy-fluorescein, acetoxymethylester - Cholin-Cl choline chloride - DMSO dimethyl sulfoxide - EDTA ethylene diamine tetra-acetic acid - FBS foetal bovine serum - H ⁺ -ATPase Proton-dependent adenosine triphosphatase - HEPES N-[2-hydroxyethyl]piperazine-N[2-ethanesulfonic acid] - pH _i intracellular pH - pH _e extracellular pH - PBS phosphate-buffered saline - SITS 4-acetamido-4-isothiocyanatostilbene-2,2-disulfonic acid     

Human neurotrophin-3: A one-step peptide mapping method and complete disulfide characterization of the recombinant protein

Human neurotrophin-3 (NT-3) is a member of the nerve growth factor (NGF) family of neurotrophic factors, and the recombinant protein is being developed as a therapeutic for neurodegenerative diseases. The final product purity and lot-to-lot variation are monitored routinely by peptide mapping. However, only the N-terminal region of NT-3 was susceptible to proteolysis under native conditions. Complete digestion required that the protein be chemically modified by reduction and S-alkylation prior to proteolysis. Complete proteolytic degradation of the protein was achieved simply by an intial denaturation of NT-3 in 6 M guanidinium chloride (pH 6) for 2 hr at 37°C, followed by a tenfold dilution with the digestion buffer (0.1 M Tris-HCl, 1 mM CaCl₂ at pH 7.0) and immediate addition of chymotrypsin at 1% by weight. Direct comparison of the peptide map with an identical aliquot that had been reduced and alkylated also allowed the establishment of the cystine linkages present in NT-3: Cys¹⁴ to Cys⁷⁹, Cys⁵⁷ to Cys¹⁰⁸, and Cys⁶⁷ to Cys¹¹⁰. This disulfide structure is homologous to the NGF family of neurotrophic factors.     

Fluorescence and Fourier-transform infrared spectroscopic studies on the role of disulfide bond in the calcium binding in the 33 kDa protein of Photosystem II

The 33 kDa protein of Photosystem II has one intrachain disulfide bond. Fluorescence spectroscopy shows that the major groups in the protein that bind to Ca²⁺ should be the carboxylic side groups of glutamic acid and/or aspartic acid. Fluorescence and Fourier-transform infrared (FTIR) spectroscopic studies indicate that the conformation of the 33 kDa protein is altered upon reduction, while the reduced protein still retains the secondary structure. FTIR spectroscopy also shows that the metal ions induce a relative decrease of unordered structure and -sheet, and a substantial increase of -helix in both the intact and the reduced 33 kDa protein. This indicates that the addition of cations results in a much more compact structure and that both the intact and the reduced 33 kDa proteins have the ability to bind calcium. The above results may suggest that the disulfide bridge is not essential for calcium binding.Abbreviations CD circular dichroism - FTIR Fourier transform infrared - La lanthanum - PS photosystem - Tb terbium     

CAM variations in the leaf-succulent Delosperma tradescantioides (Mesembryanthemaceae), native to southern Africa

Drought responses of diurnal gas exchange, malic acid accumulation and water status were examined in Delosperma tradescantioides , a succulent that grows in drought-prone microenvironments in summer rainfall and all-year rainfall regions of southern Africa. When well-watered, this species exhibited Crassulacean acid metabolism (CAM)-cycling, but its carbon fixation pattern changed during the development of drought, shifting to either low-level CAM or to CAM-idling. The rate and pattern of this change depended on environmental conditions, duration of water stress and leaf age. At the onset of drought, diurnal malate fluctuation increased, but was strongly depressed (by ca 70%) as drought continued, and when leaf water content and water potential were low (ca 35 and 50% of the initial levels, respectively). When rewatered, rates of growth and photosynthesis, gas exchange and water status recovered fully to pre-stressed values within two days. Whole-shoot carbon uptake rates suggested that leaf growth had continued unabated during a short-term (∼ one week) drought. This emphasises that CAM-idling allows the maintenance of active metabolism with negligible gas exchange when soil water is limiting. It is possible that old or senescent leaves may provide water for the expansion of developing leaves during initial periods of drought. Regardless of the water regime and environmental conditions, leaf nocturnal malate accumulation and water content were positively correlated and increased with leaf age. Thus the gradual loss of water from older mature leaves may induce CAM-idling, which reduces water loss. An important ecological consequence of this combination of CAM modes is the potential to switch rapidly between fast growth via C₃ gas exchanges when well-watered to water-conserving CAM-idling during drought.     

Relative growth rate, biomass allocation pattern and water use efficiency of three wheat cultivars during early ontogeny as dependent on water availability

We have investigated the water use efficiency of whole plants and selected leaves and allocation patterns of three wheat cultivars (Mexipak, Nesser and Katya) to explore how variation in these traits can contribute to the ability to grow in dry environments. The cultivars exhibited considerable differences in biomass allocation and water use efficiency. Cultivars with higher growth rates of roots and higher proportions of biomass in roots (Nesser and Katya) also had higher leaf growth rates, higher proportions of their biomass as leaves and higher leaf area ratios. These same cultivars had lower rates of transpiration per unit leaf area or unit root weight and higher biomass production per unit water use. They also had higher ratios of photosynthesis to transpiration, and lower ratios of intercellular to external CO₂ partial pressure. The latter resulted from large differences in stomatal conductance associated with relatively small differences in rates of photosynthesis. There was little variation between cultivars in response to drought, and differences in allocation pattern and plant water use efficiency between cultivars as found under well-watered conditions persisted under dry conditions. At the end of the non-watered treatment, relative growth rates and transpiration rates decreased to similar values for all cultivars. High ratios of photosynthesis to transpiration, and accordingly high biomass production per unit of transpiration, is regarded as a favourable trait for dry environments, since more efficient use of water postpones the decrease in plant water status.     

Chemical soil conditions in pristineNothofagus forests of New Zealand as compared to German forests

In many German forest soils low base saturation of CEC in deeper soil layers was reported and acidic deposition is seen as the major cause of these findings. To test this hypothesis we sampled 5 New Zealand forest soils from pristine beech (Nothofagus fusca, N. menziesii, N. solandri) sites under climatic and geological conditions comparable to higher elevations in Germany. The soils developed from granite and greywacke. Soil samples were analyzed for pH and the exchangeable cations were extracted with 1M NH₄Cl. The base saturation of all soil profiles was very low, even in deeper layers and was thus similar to the patterns found in many German forest soils. The pH was generally higher in the New Zealand soils as compared to Germany. The reason for the depletion of base cations in deeper soil layers of New Zealand forest soils is most likely the leaching of base cations with HCO₃ ^- resulting from the dissociation of carbonic acid in connection with high amounts of seepage. Thus, under high rainfall conditions, the low base saturation found in deeper layers of forest soils cannot exclusively be attributed to the effects of acidic depositions and land use. ei]Section editor: R F Huettl     

Exchange of NO and NO2 between wheat canopy monoliths and the atmosphere

The fluxes of NO and NO₂ between wheat canopy monoliths and the atmosphere were investigated with the dynamic chamber technique. For this purpose monoliths were dug out at different plant growth stages from a field site, transported to the institute, and placed in an environmental growth chamber. The wheat canopy monoliths were exposed over a period of four days to the average ratios of atmospheric NO₂ and NO measured at the field site, i.e. NO₂ concentration of about 18 mL L^-1 plus NO concentration lower than 0.5 nL L^-1. Under these conditions NO emission into the atmosphere and NO₂ deposition into canopy monoliths was observed. Both fluxes showed diurnal variation with maximum rates during the light and minimum rates during darkness. NO₂ fluxes correlated with soil temperature as well as with light intensity. NO fluxes correlated with soil temperature but not with light intensity. From the investigation performed the diurnal variation of the NO and NO₂ compensation points, the maximum rates of NO and NO₂ emission, and the total resistances of NO and NO₂ fluxes were calculated. Under the assumption that the measured data are representative for the whole vegetation period, annual fluxes of NO and NO₂ were estimated. Annual NO emission into the atmosphere amounted to 87 mg N m^-2 y^-1 (0.87 kg ha^-1 y^-1), annual NO₂ deposition into canopy monoliths amounted to 1273 mg N m^-2 y^-1 (12.73 kg ha^-1 y^-1). Apparently, the uptake of atmospheric nitrogen by the wheat field from NO₂ deposition is about 15 times higher than the loss of nitrogen from NO emission. It can therefore be assumed that even in rural areas wheat fields are a considerable sink for atmospheric nitrogen. The annual sink strength estimated in the present study is ca. 12 kg N ha^-1 y^-1. The possible origin of the NO emitted and the fate of atmospheric NO₂ taken up by the wheat canopy monoliths are discussed.Preliminary results of this paper were presented at the Joint Workshop COST 611/Working Party 3 and EUROTRAC in Delft, The Netherlands (Ludwig et al., 1991).     

无花果蛋白酶在阴离子交换树脂上的固定化

无花果蛋白酶通过８％戊二醛活化载体,共价结合到聚苯乙烯阴离子交换树脂ＧＭ２０１上,固定化作用在ｐＨ７．７,酶浓度０．８ｍｇ／ｇ树脂,４℃下进行６ｈ。得到的固定化酶表观Ｋ_ｍ值（酪蛋白,１．１１×１０~（－４）ｍｏｌ／Ｌ）小于溶液酶Ｋ_ｍ值（１．９６×１０~（－４）ｍｏｌ／Ｌ）;固定化酶活性在ｐＨ６～８保持稳定,溶液酶最适ｐＨ为７．２;固定化酶最适温度由溶液酶的５０～６０℃移至３７℃;固定化酶２５℃保持７ｄ,重复水解酪蛋白７次后,保留８３．３％活性。固定化酶对酪蛋白水解度达４７．５％,对大豆球蛋白达１１．６％。