Natural abundance of 15N in tropical plants with emphasis on tree legumes

Natural abundance of ¹⁵N ( ¹⁵N) of leaves harvested from tropical plants in Brazil and Thailand was analyzed. The ¹⁵N values of non-N₂-fixing trees in Brazil were +4.5±1.9, which is lower than those of soil nitrogen (+8.0±2.2). In contrast, mimosa and kudzu had very low ¹⁵N values (–1.4+0.5). The ¹⁵N values of Panicum maximum and leguminous trees, except Leucaena leucocephala, were similar to those of non-N₂-fixing trees, suggesting that the contribution of fixed N in these plants is negligible. The ¹⁵N values of non-N₂-fixing trees in Thailand were +4.9±2.0. Leucaena leucocephala, Sesbania grandiflora, Casuarina spp. and Cycas spp. had low ¹⁵N values, close to the value of atmospheric N₂ (0), pointing to a major contribution of N₂ fixation in these plants. Cassia spp. and Tamarindus indica had high ¹⁵N values, which confirms that these species are non-nodulating legumes. The ¹⁵N values of Acacia spp. and Gliricidia sepium and other potentially nodulating tree legumes were, on average, slightly lower than those of non-N₂-fixing trees, indicating a small contribution of N₂ fixation in these legumes.     

15N natural abundance studies in Australian commercial sugarcane

The measurement of natural ¹⁵N abundance is a well-established technique for the identification and quantification of biological N₂ fixation in plants. Associative N₂ fixing bacteria have been isolated from sugarcane and reported to contribute potentially significant amounts of N to plant growth and development. It has not been established whether Australian commercial sugarcane receives significant input from biological N₂ fixation, even though high populations of N₂ fixing bacteria have been isolated from Australian commercial sugarcane fields and plants. In this study, ¹⁵N measurements were used as a primary measure to identify whether Australian commercial sugarcane was obtaining significant inputs of N via biological N₂ fixation. Quantification of N input, via biological N₂ fixation, was not possible since suitable non-N₂ fixing reference plants were not present in commercial cane fields. The survey of Australian commercially grown sugarcane crops showed the majority had positive leaf ¹⁵N values (73% >3.00, 63% of which were >5.00), which was not indicative of biological N₂ fixation being the major source of N for these crops. However, a small number of sites had low or negative leaf ¹⁵N values. These crops had received high N fertiliser applications in the weeks prior to sampling. Two possible pathways that could result in low ¹⁵N values for sugarcane leaves (other than N₂ fixation) are proposed; high external N concentrations and foliar uptake of volatilised NH₃. The leaf ¹⁵N value of sugarcane grown in aerated solution culture was shown to decrease by approximately 5 with increasing external N concentration (0.5–8.0 mM), with both NO₃ ^– and NH₄ ⁺ nitrogen forms. Foliar uptake of atmospheric NH₃ has been shown to result in depleted leaf ¹⁵N values in many plant species. Acid traps collected atmospheric N with negative ¹⁵N value (–24.45±0.90) from above a field recently surface fertilised with urea. The ¹⁵N of leaves of sugarcane plants either growing directly in the soil or isolated from soil in pots dropped by 3.00 in the same field after the fertiliser application. Both the high concentration of external N in the root zone (following the application of N-fertilisers) and/or subsequent foliar uptake of volatilised NH₃ could have caused the depleted leaf ¹⁵N values measured in the sugarcane crops at these sites.     

Interactions between nitrate uptake and N2 fixation in white clover

Summary White clover (Trifolium repens L.) plants grown in pots and supplied with the same concentration x days of¹⁵N labelled nitrate, but in contrasting patterns and doses had similar N concentrations but differed in the proportions devived from N₂ fixation and nitrate. N₂-fixation and nodule dry weight responded rapidly (2–3 days) to changes in nitrate availability. Plants exposed frequently to small doses of nitrate took up more nitrate (and hence relied less on N₂-fixation) and had greater dry weights and shoot: root ratios than those exposed to larger doses less often. In mixed ryegrass (Lolium perenne L.)/clover communities clover's ability to either successfully compete for nitrate or fix N₂ gave it consistently higher N concentrations than grass whether they were given high or low nitrate nutrient. This higher N concentration was accompanied by greater dry weights than grass in the low nitrate swards but not where high levels of nitrate were applied.     

Spatial variability of soil total C and N and their stable isotopes in an upland Scottish grassland

As preparation for a below ground food web study, the spatial variability of three soil properties (total N, total C and pH) and two stable isotopes (¹³C and ¹⁵N of whole soil) were quantified using geostatistical approaches in upland pastures under contrasting management regimes (grazed, fertilised and ungrazed, unfertilised) in Scotland. This is the first such study of upland, north maritime grasslands. The resulting patterns of variability suggest that to obtain statistically independent samples in this system, a sampling distance of 13.5 m is required. Additionally, temporal change (a decline of 1) was observed in whole soil ¹⁵N for the grazed, fertilised plot. This may have been caused by new inputs of symbiotically-fixed atmospheric N₂.     

The 15N/14N ratios of plants in South Africa and Namibia: relationship to climate and coastal/saline environments

Summary Data are presented for the ¹⁵N/¹⁴N ratios of 140 indigenous terrestrial plants from a wide variety of natural habitats in South Africa and Namibia. Over much of the area, from high-rainfall mountains to arid deserts, the ¹⁵N values of plants lie typically in the range -1 to +6; with no evident differences between C₃ plants and C₄ grasses. There is a slight correlation between ¹⁵N and aridity, but this is less marked than the correlation between the ¹⁵N values of animal bones and aridity. At coastal or saline sites, however, the mean ¹⁵N values for plants are higher than those at nearby inland or non-saline sites-e.g.: arid Namib coast (10 higher than inland Namib); wet Natal beach (5 higher than inland Natal); saline soils 500 km from coast (4 higher than non-saline soils). High values were also found at one site where there were no marked coastal or saline influences. These environmental effects on the isotopic composition of plants will extend upwards to the animals and humans they support. They therefore have important consequences for the use of nitrogen isotope data in the study of the dietary habits and trophic structures of modern and prehistoric communities.     

Variation in the natural abundance of 15N in the halophyte,Salicornia virginica,associated with groundwater subsidies of nitrogen in a southern California salt-marsh

To provide insight into the importance of the salt-marsh ecotone as a sink for inorganic nitrogen in perched groundwater, measurements were made of the natural abundance of ¹⁵N in dissolved NO₃-N and NH₄-N and in the salt-marsh halophyte, Salicornia virginica, along an environmental gradient from agricultural land into a salt-marsh. The increase in the natural abundance of ¹⁵N (expressed by convention as ¹⁵N) of NO₃-N, accompanied by the decrease in NO₃-N (and total dissolved inorganic N, DIN) concentration along the gradient, suggested that the salt-marsh ecotone is a site of transformation, most likely through denitrification, of inorganic nitrogen in groundwater. ¹⁵N enrichment in S. virginica (and the parasitic herb, Cuscuta salina), along the tidal marsh boundary, relative to high and middle marsh locations, indicated the retention of groundwater nitrogen as vegetative biomass. The correlation between ¹⁵N _Salicornia and ¹⁵N_NH4 suggested a preference for NH₄-N over NO₃-N during uptake by this plant. Groundwater inputs enhanced the standing crop, above-ground productivity, and nitrogen content of S. virginica but the ralative effects of pore water salinity and DIN concentration on these parameters were not determined. ¹⁵N enrichment of marsh plants by groundwater DIN inputs could prove useful in tracing the fate of these inputs in the marsh food web.     

Assignment of aliphatic side-chain 1HN/15N resonances in perdeuterated proteins

Summary The perdeuteration of aliphatic sites in large proteins has been shown to greatly facilitate the process of sequential backbone and side-chain ¹³C assignments and has also been utilized in obtaining long-range NOE distance restraints for structure calculations. To obtain the maximum information from a 4D ¹⁵N/¹⁵N-separated NOESY, as many main-chain and side-chain ¹H_N/¹⁵N resonances as possible must be assigned. Traditionally, only backbone amide ¹H_N/¹⁵N resonances are assigned by correlation experiments, whereas slowly exchanging side-chain amide, amino, and guanidino protons are assigned by NOEs to side-chain aliphatic protons. In a perdeuterated protein, however, there is a minimal number of such protons. We have therefore developed several gradient-enhanced and sensitivity-enhanced pulse sequences, containing water-flipback pulses, to provide through-bond correlations of the aliphatic side-chain ¹H_N/¹⁵N resonances to side-chain ¹³C resonances with high sensitivity: NH₂-filtered 2D ¹H-¹⁵N HSQC (H₂N-HSQC), 3D H₂N(CO)C_/ and 3D H₂N(COC_/)C_/ for glutamine and asparagine side-chain amide groups; 2D refocused H(N_/)C_/ and H(N_/C_/)C_/ for arginine side-chain amino groups and non-refocused versions for lysine side-chain amino groups; and 2D refocused H(N)C and nonrefocused H(N_.)C for arginine side-chain guanidino groups. These pulse sequences have been applied to perdeuterated ¹³C-/¹⁵N-labeled human carbonic anhydrase II (²H-HCA II). Because more than 95% of all side-chain ¹³C resonances in ²H-HCA II have already been assigned with the C(CC)(CO)NH experiment, the assignment of the side-chain ¹H_N/¹⁵N resonances has been straightforward using the pulse sequences mentioned above. The importance of assigning these side-chain H_N protons has been demonstrated by recent studies in which the calculation of protein global folds was simulated using only ¹H_N-¹H_N NOE restraints. In these studies, the inclusion of NOE restraints to side-chain H_N protons significantly improved the quality of the global fold that could be determined for a perdeuterated protein [R.A. Venters et al. (1995) J. Am. Chem. Soc., 117, 9592–9593].To whom correspondence should be addressed.     

Carbon isotope composition of N2-fixing and N-fertilized legumes along an elevational gradient

Dinitrogen-fixing legumes are frequently assumed to be less water-use efficient than plants utilizing soil mineral N, because of the high respiratory requirements for driving N₂ fixation. However, since respiration is assumed not to discriminate against ¹³C, any differences in water-use efficiency exclusively due to respiration should not be apparent in carbon isotope discrimination () values. Our objective was to determine if the source of N (N₂ fixation versus soil N) had any effect on of field-grown grain legumes grown at different elevations. Four legume species, Glycine max, Phaseolus lunatus, P. vulgaris, and Vigna unguiculata, were grown on five field sites spanning a 633 m elevational gradient on the island of Maui, Hawaii. The legumes were either inoculated with a mixture of three effective strains of rhizobia or fertilized weekly with urea at 100 kg N ha^-1 in an attempt to completely suppress symbiotic N₂-fixing activity. In 14 of 20 analyses of stover and 12 of 15 analyses of seed values were significantly higher (p=0.10) in the inoculated plants than the N-fertilized plants. Nitrogen concentrations were generally higher in the fertilized treatments than the inoculated treatments. The different values obtained depending on N-source may have implications in using as an indicator of water-use efficiency or yield potential of legumes.     

Calcium and tip growth inNeurospora crassa

Summary We examined the ionic regulation of tip growth inNeurospora crassa by a combination of electrophysiology and confocal microscopy. To determine if transmembrane ionic fluxes are required for tip growth, we voltage clamped the membrane from –200 to +50 mV. In this voltage range, transmembrane ionic fluxes would either reverse (e.g., K⁺) or change dramatically (e.g., Ca²⁺ influx) but had no effect on hyphal growth rates. Therefore, ionic fluxes (including Ca²⁺ influx) may not be required for tip growth. However, intracellular Ca²⁺ may still play an obligatory role in tip growth. To assess this possibility, we first increased cytosolic Ca²⁺ directly by ionophoresis. Elevated Ca²⁺ induced subapical branch initiation, often multiple tips. At hyphal tips, fluorescence ratio imaging using fluo-3 and fura-red revealed a pronounced tip-high Ca²⁺ gradient within 10 m of the tip in growing hyphae which was not observed in nongrowing hyphae. Injection of the Ca²⁺ chelator 1,2-bis(ortho-aminophenoxy)ethane-N,N,N,N-tetrapotassium acetate consistently inhibited growth concomitantly with a depletion of intracellular Ca²⁺ and dissipation of the tip-high gradient. We conclude that Ca²⁺ plays a regulatory role in tip initiation and the maintenance of tip growth. Because plasma membrane ionic fluxes do not play a role in tip growth, we suggest that the tip-high Ca²⁺ gradient is generated from intracellular Ca²⁺ stores in the ascomyceteN. crassa.Abbreviations BAPTA 1,2-bis(ortho-aminophenoxy)ethane-N,N,N,N-tetrapotassium acetate - [Ca²⁺]_i intracellular Ca²⁺ concentration - fluo-3 2,7-dichloro-6-hydroxy-3-oxo-9-xanthenyl-4-methyl-2,2-(ethylenedioxy)dianiline-N,N,N,N-tetraacetic acid     

15N natural abundances and N use by tundra plants

Plant species collected from tundra ecosystems located along a north-south transect from central Alaska to the north coast of Alaska showed large and consistent differences in ¹⁵N natural abundances. Foliar ¹⁵N values varied by about 10% among species within each of two moist tussock tundra sites. Differences in ¹⁵N contents among species or plant groups were consistent across moist tussock tundra at several other sites and across five other tundra types at a single site. Ericaceous species had the lowest ¹⁵N values, ranging between about –8 to –6. Foliar ¹⁵N contents increased progressively in birch, willows and sedges to maximum ¹⁵N values of about +2 in sedges. Soil ¹⁵N contents in tundra ecosystems at our two most intensively studied sites increased with depth and ¹⁵N values were usually higher for soils than for plants. Isotopic fractionations during soil N transformations and possibly during plant N uptake could lead to observed differences in ¹⁵N contents among plant species and between plants and soils. Patterns of variation in ¹⁵N content among species indicate that tundra plants acquire nitrogen in extremely nutrient-poor environments by competitive partitioning of the overall N pool. Differences in plant N sources, rooting depth, mycorrhizal associations, forms of N taken up, and other factors controlling plant N uptake are possible causes of variations in ¹⁵N values of tundra plant species.     

Calcium requirement in nitrogen fixation in the cyanobacterium Synechococcus RF-1

Under diurnal 16/8-h light-dark cycles, ethyleneglycol-bis-(-aminoethyl ether)-N,N,N,N-tetraacetic acid (EGTA) at 1 mM completely blocked the appearance of rhythmic N₂-fixing activity in Synechococcus RF-1. Ca²⁺ at 2 mM, when supplied either together with or several hours after the EGTA application, restored the nitrogenase activity, whereas, when Ca²⁺ was supplied several hours later, the peak of nitrogenase activity was shifted from the dark to the light period in which the activity is normally suppressed. Sr²⁺ also reversed the inhibition by EGTA, but only partially. When O₂ in the gas phase above the culture was below 1%, the inhibition of nitrogenase activity by EGTA was reduced to less than 20% of the control value without EGTA. Thus Ca²⁺ appears to be required by the cell to protect its nitrogenase from inactivation by O₂. In media without EGTA, a close correlation between nitrogenase activity and concentrations of Ca²⁺ was also observed.Abbreviation EGTA ethyleneglycol-bis-(-aminoethyl ether)-N,N,N,N-tetraacetic acid     

Estimates of nitrogen fixation by trees on an aridity gradient in Namibia

Summary Nitrogen (N₂) fixation was estimated along an aridity gradient in Namibia from the natural abundance of ¹⁵N (¹⁵N value) in 11 woody species of the Mimosacease which were compared with the ¹⁵N values in 11 woody non-Mimosaceae. Averaging all species and habitats the calculated contribution of N₂ fixation (N _f ) to leaf nitrogen (N) concentration of Mimosaceae averaged about 30%, with large variation between and within species. While in Acacia albida N _f was only 2%, it was 49% in Acacia hereroensis and Dichrostachys cinerea, and reached 71% in Acacia melifera. In the majority of species N _f was 10–30%. There was a marked variation in background ¹⁵N values along the aridity gradient, with the highest ¹⁵N values in the lowland savanna. The difference between ¹⁵N values of Mimosaceae and non-Mimosaceae, which is assumed to result mainly from N₂ fixation, was also largest in the lowland savanna. Variations in ¹⁵N of Mimosaceae did not affect N concentrations, but higher ¹⁵N-values of Mimosaeae are associated with lower carbon isotope ratios (¹³C value). N₂ fixation was associated with reduced intrinsic water use efficiency. The opposite trends were found in non-Mimosaceae, in which N-concentration increased with ¹⁵N, but ¹³C was unaffected. The large variation among species and sites is discussed.This paper is prepared in memory of J. Visser, who took part in the collection of species, but died in 1990     

Structure and function of the yeast vacuolar membrane proton ATPase

Our current work on a vacuolar membrane proton ATPase in the yeastSaccharomyces cerevisiae has revealed that it is a third type of H⁺-translocating ATPase in the organism. A three-subunit ATPase, which has been purified to near homogeneity from vacuolar membrane vesicles, shares with the native, membrane-bound enzyme common enzymological properties of substrate specificities and inhibitor sensitivities and are clearly distinct from two established types of proton ATPase, the mitochondrial F₀F₁-type ATP synthase and the plasma membrane E₁E₂-type H⁺-ATPase. The vacuolar membrane H⁺-ATPase is composed of three major subunits, subunita (M _r =67 kDa),b (57kDa), andc (20 kDa). Subunita is the catalytic site and subunitc functions as a channel for proton translocation in the enzyme complex. The function of subunitb has not yet been identified. The functional molecular masses of the H⁺-ATPase under two kinetic conditions have been determined to be 0.9–1.1×10⁵ daltons for single-cycle hydrolysis of ATP and 4.1–5.3×10⁵ daltons for multicycle hydrolysis of ATP, respectively.N,N-Dicyclohexylcarbodiimide does not inhibit the former reaction but strongly inhibits the latter reaction. The kinetics of single-cycle hydrolysis of ATP indicates the formation of an enzyme-ATP complex and subsequent hydrolysis of the bound ATP to ADP and Pi at a 7-chloro-4-nitrobenzo-2-oxa-1,3-diazolesensitive catalytic site. Cloning of structural genes for the three subunits of the H⁺-ATPase (VMA1, VMA2, andVMA3) and their nucleotide sequence determination have been accomplished, which provide greater advantages for molecular biological studies on the structure-function relationship and biogenesis of the enzyme complex. Bioenergetic aspects of the vacuole as a main, acidic compartment ensuring ionic homeostasis in the cytosol have been described.Abbreviations CCCP carbonyl cyanidem-chlorophenyl hydrazone - DCCD N,N-dicyclohexylcarbondiimide - DES diethylstilbestrol - DIDS 4,4-diisothiocyano-2,2-stilbene disulfonic acid - NBD-Cl 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole - Pi inorganic phosphate - SDS sodium dodecylsulfate - SF6847 3,5-di-tert-butyl-4-hydroxybenzylidenemalononitrile - SITS 4-acetamide-4-isothiocyanatostilbene-2,2-disulfonic acid - ZW3-14 N-tetradecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate     

Electron transport-linked proton translocation at nitrite reduction inCampylobacter sputorum subspeciesbubulus

Campylobacter sputorum subspeciesbubulus contains a membrane-bound nitrite reductase which catalyses the six-electron reduction of nitrite to ammonia. Formate andL-lactate are used as hydrogen donors. Cells ofC. sputorum grown with nitrate or nitrite contain cytochromes of theb-andc-type and a carbon monoxide-binding cytochromec. In addition, a special membrane-bound carbon monoxide-binding pigment is found. Nitrite reduction with formate orL-lactate as a hydrogen donor is strongly inhibited by 2-n-heptyl-4-hydroxyquinoline-N-oxide (HQNO). Nitrite reduction by bacterial suspensions with lactate as a hydrogen donor is strongly inhibited by carbonylcyanide-m-chlorophenyl-hydrazone (CCCP) whereas nitrite reduction with formate as a hydrogen donor is not inhibited at all. H⁺/O values and H⁺/NO ₂ ^- values were measured with ascorbate + N,N,N,N-tetramethyl-p-phenylenediamine (TMPD), formate (in the absence and presence of carbonic anhydrase) andL-lactate as a hydrogen donor. The results are summarized in a scheme for electron transport from formate or lactate to oxygen or nitrite which shows a periplasmic orientation of formate dehydrogenase and nitrite reductase and a cytoplasmic orientation of lactate dehydrogenase and oxygen reduction, and which shows proton translocation with a H⁺/2e value of 2.0. The H⁺/O and H⁺/NO ₂ ^- values predicted by this scheme are in good agreement with the experimental values.Abbreviations CCCP carbonylcyanide-m-chlorophenylhydrazone - HQNO 2-n-heptyl-4-hydroxyquinoline-N-oxide - MTPP⁺ methyltriphenylphosphonium cation - TMPD N,N,N,N-tetramethyl-p-phenylenediamine; H⁺/O (H⁺/NO ₂ ^- ), number of protons liberated in the outer bulk phase at the reduction of one atom O (one ion NO ₂ ^- ); H⁺/2e (q⁺/2e), number of protons (charges) translocated across the cytoplasmic membrane during flow of two electrons to an acceptor     

Theβ-subunit of human chorionic gonadotropin containsN-glycosidic trisialo tri- and tri′-antennary carbohydrate chains

TheN-linked carbohydrate chains of the-subunit of highly purified urinary human chorionic gonadotropin have been re-investigated. The oligosaccharides were released enzymatically by peptide-N ⁴-(N-acetyl--glucosaminyl)asparagine amidase-F, and fractionated by a combination of FPLC and HPLC. As a result of the application of improved fractionation methods, apart from the earlier reported carbohydrate chains, also small amounts of trisialo tri- and tri-antennary oligosaccharides were found. The primary structures of the latter carbohydrate chains have been determined by 500-MHz¹H-NMR spectroscopy to beAbbreviations hCG human chorionic gonadotropin - hCG- -subunit - hCG- -subunit - PNGase-F peptide-N ⁴-(N-acetyl--glucosaminyl)asparagine amidase-F (E.C. 3.5.1.52) - endo-F endo--N-acetylglucosaminidase-F (E.C. 3.2.1.96) - SDS sodium dodecyl sulphate - PAGE polyacrylamide gel electrophoresis - CBB coomassie brilliant blue R 250 - GlcNAc N-acetylglucosamine - NeuAc N-acetylneuraminic acid - Man mannose - Gal galactose - Fuc fucose     

A study on Na+-coupled oxidative phosphorylation: ATP formation supported by artificially imposed ΔpNa and ΔpK inVibrio alginolyticus cells

Addition of Na⁺ to the K⁺-loadedVibrio alginolyticus cells, creating a 250-fold Na⁺ gradient, is shown to induce a transient increase in the intracellular ATP concentration, which is abolished by the Na⁺/H⁺ antiporter, monensin. The pNa-supported ATP synthesis requires an additional driving force supplied by endogenous respiration or, alternatively, by a K⁺ gradient (high [K⁺] inside). In the former case, ATP formation is resistant to the protonophorous uncoupler. Dicyclohexylcarbodiimide and diethylstilbestrol, but not vanadate, completely inhibit Na⁺ pulse-induced ATP formation. The data agree with the assumption that Na⁺-ATP-synthase is involved in oxidative phosphorylation inV. alginolyticus. Interrelation of H⁺ and Na⁺ cycles in bacteria is discussed.Abbreviations and electrochemical gradients of H⁺ and Na⁺, respectively - transmembrane electric potential difference - pH, pNa, and pK concentration gradients of H⁺, Na⁺, and K⁺, respectively - CCCP carbonyl cyanidem-chlorophenylhydrazone - DCCD N,N-dicyclohexylcarbodiimide - DES diesthylstilbestrol - HQNO 2-heptyl-4-hydroxyquinolineN-oxide - Tricine N[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine     

Angular dependence of 1J(Ni,Calphai) and 2J(Ni,Calpha(i-1)) coupling constants measured in J-modulated HSQCs

A new method to measure ¹J(N_i,C _i) and ²J(N_i,C _{(i – 1)}) coupling constants in proteins based on a J-modulated sensitivity enhanced HSQC was introduced. Coupling constants were measured in the denatured and in the native state of ubiquitin and found to depend on the conformation of the protein backbone. Using a combined data set of experimental coupling constants from ubiquitin and staphylococcal nuclease (Delaglio et al., 1991), the angular dependence of the coupling constants on the backbone angles and was investigated. It was found that the size of ²J(N_i,C _{(i – 1)}) correlates strongly with the backbone conformation, while only a weak conformational dependence on the size of ¹J(N_i,C _i) coupling constants was observed. Coupling constants in the denatured state of ubiquitin were uniform along the sequence of the protein and not dependent on a given residue type. Furthermore it was shown that the observed coupling constants were in good agreement with predicted coupling constants using a simple model for the random coil.     

Sodium efflux from perfused giant algal cells

Internodal cells of the giant alga Chara corallina were perfused internally to replace the native cytoplasm, tonoplast and vacuole with artificial cytoplasm. Sodium efflux from perfused cells, measured by including ²²Na in the perfusion media, was increased by increasing the internal sodium concentration and by decreasing the external pH, and was inhibited by external application of the renal diuretic amiloride. The sodium efflux was markedly ATP-dependent, with a 50-fold decrease in efflux observed after perfusion with media lacking ATP. Efflux in the presence of ATP was reduced by 33% by inclusion of 10 M N,N-dicyclohexylcarbodiimide in the perfusion medium. The membrane potential of the perfused cells approximated that of intact cells from the same culture. It is suggested that sodium efflux in perfused Chara cells proceeds via a secondary antiporter with protons, regulated by ATP in a catalytic role and with the proton motive force acting as the energy source.Abbreviations DCCD N,N-dicyclohexylcarbodiimide - EGTA ethylene glycol-bis(-aminoethyl ether)-N,N,N,N-tetraacetic acid - Mes 2-(N-morpholino)ethanesulphonic acid - Mops 3(N-morpholino)propanesulphonic acid - Taps tris(hydroxymethyl)methylaminopropanesulphonic acid     

Stable oxygen isotopes in<Emphasis Type="Italic"> Porites</Emphasis> corals monitor weekly temperature variations in the northern Gulf of Aqaba,Red Sea

In order to assess the ability of Porites corals to accurately record environmental variations, high-resolution (weekly/biweekly) coral ¹⁸O records were obtained from four coral colonies from the northern Gulf of Aqaba, which grew at depths of 7, 19, 29, and 42 m along one transect. Adjacent to each colony, hourly temperatures, biweekly salinities, and monthly ¹⁸O of seawater were continuously recorded over a period of 14 months (April 1999 to June 2000). Contrary to water temperature, which shows a regular and strong seasonal variation and change with depth, seawater ¹⁸O exhibits a weak seasonality and little change with depth. Positive correlations between seawater ¹⁸O and salinity were observed. The two parameters were related to each other by the equation ¹⁸O _Seawater (, VSMOW) = 0.281 × Salinity – 9.14. The high-resolution coral ¹⁸O records from this study show a regular pattern of seasonality and are able to capture fine details of the weekly average temperature records. They resolve more than 95% of the weekly average temperature range. On the other hand, attenuation and amplification of coral seasonal amplitudes were recorded in deep, slow-growing corals, which were not related to environmental effects (temperature and/or seawater ¹⁸O) or sampling resolution. We propose that these result from a combined effect of subannual variations in extension rate and variable rates of spine thickening of skeletal structures within the tissue layer. However, no smoothing or distortion of the isotopic signals was observed due to calcification within the tissue layer in shallow-water, fast-growing corals. The calculations from coral ¹⁸O calibrations against the in situ measurements show that temperature (T) is related to coral ¹⁸O ( _c ) and seawater ¹⁸O ( _w ) by the equation T (°C) = –5.38 ( _c – _w ) –1.08. Our results demonstrate that coral ¹⁸O from the northern Gulf of Aqaba is a reliable recorder of temperature variations, and that there is a minor contribution of seawater ¹⁸O to this proxy, which could be ignored.     

Interaction between photosynthetic and respiratory electron-transfer chains in the membranes of Anabaena variabilis

The rate of CO₂- and p-benzoquione-dependent photosynthetic O₂ evolution by Anabaena variabilis cells remained unaltered and the rate of O₂ uptake observed after switching off the light (endogenous respiration) was enhanced by a factor of 6–8 when the O₂ concentration was increased from 200 to 400 M. Photosystem-I-linked O₂ uptake and respiration of the cells incubated with ascorbate and N,N,NN-tetramethyl-p-phenylenediamine was not appreciable influenced by the O₂ concentration. 2-Iodo-6-isopropyl-3-methyl-2,4,4-trinitrodiphenyl ether, blocking electron transfer at the plastoquinone level, suppressed O₂ evolution and had no influence on endogenous respiration. 2-n-Heptyl-4-hydroxyquinoline-N-oxide, an inhibitor of electron transfer between photosystems II and I, as well as the cytochrome-oxidase inhibitors N ₃ ^- , CN^- and NH₂OH, caused a 35–50% retardation of endogenous respiration and blocked photosynthetic O₂ evolution. The molar ratio of cytochromes b₆, f, c-553, aa₃ and photosystem-I reaction centers in the isolated membranes equalled approx. 2:1:2:0.7:2. It is inferred that endogenous respiration of A. variabilis cells is inhibited by the light-induced electron flow through both photosystems at the level of the plastoquinone-plastocyanin-oxidoreductase complex.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DNP-INT 2-iodo-6-isopropyl-3-methyl-2,4,4-trinitrodiphenyl ether - Hepes 4-(2-hydroxyethyl)-1-piperazine ethansulfonic acid - TMPD N,N,NN-tetramethyl-p-phenylenediamine