Green mass aggregations of Gyrodinium cf. aureolum Hulburt in the Ria of Pontevedra (north-west Spain)

In this paper we report a bloom of Gyrodinium cf. aureolum occurringin an estuary of the north-west coast of Spain. It was sampledin late August 1989, in shallow waters 2 m deep in the Ria ofPontevedra. Its temperature was 19.5°C and salinity 35     

INTRACELLULAR AND EXTRACELLULAR AMINO ACID POOLS OF THE MARINE DIATOM PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE) GROWN ON UNENRICHED SEAWATER IN HIGH-CELL-DENSITY DIALYSIS CULTURE1

The consumption of inorganic macronutrients (NO₃^?+ NO₂^?, NH₄⁺, and PO₄^?3) and the composition of intra- and extracellular dissolved free amino acid pools (IDFAA and EDFAA, respectively) were determined in continuous-reservoir batch dialysis cultures of the marine diatom Phaeodactylum tricornutum Bohlin maintained on unenriched natural seawater as a growth medium. Nutrient diffusion (Nd), which equals the nutrient uptake of the culture, increased with the cell density and the age of the culture. A concentration of 6.77 × 10⁷ cells · mL^?1 was obtained in stationary phase, which coincided with the NO₃^?+ NO₂^? diffusion limit (Nd_max) of the dialysis apparatus. The Nd_max for NH₄⁺ occurred much earlier, at the end of exponential growth, whereas Nd_max for PO₄^?3 was not attained during the growth cycle of the culture, even in early stationary phase. A significant depletion (77%) of the IDFAA pool during exponential phase was followed by a reestablishment–to approximately 60% of the initial level–of internal pools during linear and stationary growth phases. This recovery occurred during the illuminated portion of the photoperiod (12:12 h LD) and involved principally the amino acids GLN, GLU, β-GLU, and ASN. The recovery of GLN and ASN levels was particularly significant, because the intracellular concentrations of these amino acids were higher at the end of the growth cycle than before. The EDFAA pool was generally dominated by the amino acids SER and GLY+THR; however, during active growth, ORN and LYS often constituted an important fraction. The EDFAA concentration increased until linear growth phase was reached, during which a higher concentration of total free amino acids was attained in darkness than under illumination. The EDFAA component diminished afterward, and in stationary phase this fraction returned to concentrations equivalent to those observed at the beginning of the growth cycle. The variations in EDFAA concentrations were expressed by a pronounced decrease in the cellular excretion of amino acids with increasing cell density. These cellular responses of Phaeodactylum tricornutum in dense culture, specifically the regulation of amino acid excretion and intracellular pool size, may affect the N-conversion coefficient (Y_N). Consequently, by prolonging the linear phase of growth and reducing the concentration of autoinhibitory metabolites by diffusion, a markedly enhanced final cell density can be achieved in cultures grown on natural unenriched seawater.     

The deactivation pathways of the excited-states of the mycosporine-like amino acids shinorine and porphyra-334 in aqueous solution.

In vitro studies on the structurally related mycosporine-like amino acids (MAAs) porphyra-334 and shinorine in aqueous solutions were carried out aiming at their full photochemical and photophysical characterization and expanding the evidence on the assigned UV-photoprotective role of the molecules in vivo. The experiments on shinorine confirmed a high photostability and a poor fluorescence quantum yield, in concordance with previous results on porphyra-334. The estimation of triplet production quantum yields for both MAAs was achieved by laser-flash photolysis measurements. In particular, photosensitization experiments on porphyra-334 support the participation of the triplet state in the photodecomposition mechanism yielding a more precise value of [capital Phi](T). As well, photoacoustic calorimetry experiments allowed the first direct quantification of the nonradiative relaxation pathways of the excited MAAs in solution, corroborating that the vast majority (ca. 97%) of the absorbed energy is promptly delivered to the surroundings as heat, consistently with the low photodecomposition and emission yields observed.     

The ability of rufous hummingbirds Selasphorus rufus to dilute and concentrate urine

Most terrestrial animals face the challenge of having to conserve water in a desiccating environment. Not surprisingly, the ability to produce concentrated urine has been relatively well studied in birds. Nectar-feeding birds are unusual among terrestrial animals in that they often ingest and excrete prodigious water volumes to obtain adequate energy. Thus, they confront the unusual challenge of having to conserve electrolytes. The diluting abilities of birds and the renal mechanisms that may correlate with them have been relatively neglected. To elucidate diluting and concentrating abilities in nectar-feeding birds, we fed rufous hummingbirds Selasphorus rufus an electrolyte-free nectar and a nectar containing a range of NaCl concentrations. Hummingbirds had a spectacular (and possibly unique) diluting ability: when fed on electrolyte-free food they produced excreta containing less than 0.5 mM l⁻¹ each of sodium and potassium. Hummingbirds also had a poor concentrating ability, retaining sodium and chloride when their food (0.632 M l⁻¹ sucrose) contained more than 35 mM l⁻¹ of NaCl. The kidneys of hummingbirds do not appear to be suited for concentrating urine, and possibly contain structural features that give them a unique diluting ability compared with those of birds that do not feed on nectar.     

Evaluation of coupling conditions for the incorporation of Nα-Fmoc-Tyr(PO3H2)-OH in solid-phase peptide synthesis

Summary In order to minimise the formation of the pyrophosphate derivative of the target peptide when side-chain-unprotected phopshotyrosine is used in solid-phase peptide synthesis, this building block can be incorporated using benzotriazolyloxy-tris-(dimethylamino)phosphonium hexafluorophosphate/1-hydroxybenzotriazole/N-methylmorpholine (1:1:2.3) in the presence of a chaotropic salt (0.4 M LiCl in N-methyl-2-pyrrolidinone).Abbreviations BOP benzotriazolyloxy-tris-(dimethylamino)phosphonium hexafluorophosphate - DIEA diisopropylethylamine - Fmoc 9-fluorenylmethoxycarbony - HATU N-[(dimethylamino)1H-1,2,3-triazolo[4,5-b]pyridin-1-ylmethylene]-N-methylmethan-aminium hexafluorophosphate N-oxide - HOBt 1-hydroxybenzotriazole - HPLC high-performance liquid chromatography - MALDI-TOF matrix-assisted laser-desorption ionization time-of-flight mass spectrometry - NMM N-methylmorpholine - NMP N-methyl-2-pyrrolidinone - Pmc 2,2,5,7,8-pentamethyl-chroman-6-sulfonyl - ® solid support - TFA trifluoroacetic acid - TPTU 2-(2-pyridon-1-yl)-1,1,3,3-tetramethyluroniumfluoroborate. Abbreviations used for amino acids follow the recommendations of the IUPAC-IUB Commission of Biochemical Nomenclature [Eur. J. Biochem., 138 (1984) 9]     

Phase labeling of C−H and C−C spin-system topologies: Application in PFG-HACANH and PFG-HACA(CO)NH triple-resonance experiments for determining backbone resonance assignments in proteins

Summary Triple-resonance experiments can be designed to provide useful information on spin-system topologies. In this paper we demonstrate optimized proton and carbon versions of PFG-CT-HACANH and PFG-CT-HACA(CO)NH straight-through triple-resonance experiments that allow rapid and almost complete assignments of backbone H, ¹³C, ¹⁵N and H^N resonances in small proteins. This work provides a practical guide to using these experiments for determining resonance assignments in proteins, and for identifying both intraresidue and sequential connections involving glycine residues. Two types of delay tunings within these pulse sequences provide phase discrimination of backbone Gly C and H resonances: (i) C–H phase discrimination by tuning of the refocusing period _{a_f}; (ii) C–C phase discrimination by tuning of the ¹³C constant-time evolution period 2T_c. For small proteins, C–C phase tuning provides better S/N ratios in PFG-CT-HACANH experiments while C–H phase tuning provides better S/N ratios in PFG-CT-HACA(CO)NH. These same principles can also be applied to triple-resonance experiments utilizing ¹³C-¹³C COSY and TOCSY transfer from peripheral side-chain atoms with detection of backbone amide protons for classification of side-chain spin-system topologies. Such data are valuable in algorithms for automated analysis of resonance assignments in proteins.     

Redox-Bohr effect in the tetrahaem cytochrome c3 from Desulfovibrio vulgaris: a model for energy transduction mechanisms

 Using potentiometric titrations, two protons were found to participate in the redox-Bohr effect observed for cytochrome c ₃ from Desulfovibrio vulgaris (Hildenborough). Within the framework of the thermodynamic model previously presented, this finding supports the occurrence of a concerted proton-assisted 2e^– step, ideally suited for the coupling role of cytochrome c ₃ to hydrogenase. Furthermore, at physiological pH, it is shown that when sulfate-reducing bacteria use H₂ as energy source, cytochrome c ₃ can be used as a charge separation device, achieving energy transduction by energising protons which can be left in the acidic periplasmic side and transferring deenergised electrons to sulfate respiration. This mechanism for energy transduction, using a full thermodynamic data set, is compared to that put forward to explain the proton-pumping function of cytochrome c oxidase.     

The vegetable rennet of Cynara cardunculus L. contains two proteinases with chymosin and pepsin-like specificities

The flowers of cardoon (genus Cynara) are traditionally used in Portugal for cheese making. In this work the vegetable rennet of the species Cynara cardunculus L. was characterized in terms of enzymic composition and proteolytic specificity of its proteinases (cardosin A and cardosin B). Cardosin A was found to cleave insulin B chain at the bonds Leu15-Tyr16, Leu17-Val18 and Phe25-Tyr26. In addition to the bonds mentioned cardosin B cleaves also Glu13-Ala14, Ala14-Leu15 and Phe24-Phe25 indicating that it has a broader specificity. The kinetic parameters for the hydrolysis of the synthetic peptide Leu-Ser-Phe(NO₂)-Nle-Ala-Leu-oMe were also determined and compared to those of chymosin and pepsin. The results obtained indicate that in terms of specificity and kinetic parameters cardosin A is similar to chymosin whereas cardosin B is similar to pepsin. It appears therefore that the enzyme composition of cardoon rennet closely resembles that of calf rennet.     

Changes in the origin and quality of soil organic matter after pasture introduction in Rondônia (Brazil)

We examined the effects of the conversion of tropical forest to pasture on soil organic matter (SOM) origin and quality along a chronosequence of sites, including a primary forest and six pastures. Bulk soil samples received a physical size-fractionation treatment to assess the contribution of each compartment to total SOM pool. Besides a general increase in total C and N stocks along the chronosequence, we observed a reduction of the relative contribution of the coarser fractions to total soil C content, and an increased concentration in the finer fractions. The origin of the C in each size fraction was established from measurements of¹³C abundance. After 80 years about 93% of the C in the least humified fraction of the top 10 cm of soil was of pasture origin, while in the most humified it was 82%. Chemical analyses indicated that the fine silt and coarse clay fractions contained the most refractory carbon.