Function of C1A barley peptidases and their inhibitors (cystatins) in barley seed germination : Funcion de las peptidasas C1A de cebada y sus inhibidores (cistatinas) en la germinacion de la semilla

Plant proteolysis is a metabolic process where specific enzymes called peptidases degrade proteins. In plants, this complex process involves broad metabolic networks and different sub-cellular compartments. Several types of peptidases take part in the proteolytic process, mainly cysteine-, serine-, aspartyl- and metallo- peptidases. Among the cysteine-peptidases, the papain-like or C1A peptidases (family C1, clan CA) are extensively present in land plants and are classified into catepsins L-, B-, H- and Flike. The catalytic mechanism of these C1A peptidases is highly conserved and involves the three amino acids Cys, His and Asn in the catalytic triad, and a Gln residue which seems essential for maintaining an active enzyme conformation. These proteins are synthesized as inactive precursors, which comprise an N-terminal signal peptide, a propeptide, and the mature protein. In barley, we have identified 33 cysteine-peptidases from the papain-like family, classifying them into 8 different groups. Five of them corresponded to cathepsins L-like (5 subgroups), 1 cathepsin B-like group, 1 cathepsin F-like group and 1 cathepsin H-like group. Besides, C1A peptidases are the specific targets of the plant proteinaceous inhibitors known as phytocystatins (PhyCys). The cystatin inhibitory mechanism is produced by a tight and reversible interaction with their target enzymes. In barley, the cystatin gene family is comprised by 13 members. In this work we have tried to elucidate the role of the C1A cysteine-peptidases and their specific inhibitors (cystatins) in the germination process of the barley grain. Therefore, we selected a representative member of each group/subgroup of C1A peptidases (1 cathepsin B-like, 1 cathepsin F-like, 1 cathepsin H-like and 5 cathepsins L-like). The molecular characterization of the cysteine-peptidases was done and the peptidase-inhibitor interaction was analyzed in vitro and in vivo. A study in the structural basis for specificity of pro-peptide/enzyme interaction in barley C1A cysteine-peptidases has been also carried out by inhibitory assays and the modeling of the three-dimensional structures. The barley grain maturation produces the accumulation of storage proteins (prolamins) in the endosperm which are mobilized during germination to supply the required nutrients until the photosynthesis is fully established. In this work, we have demonstrated the participation of the cysteine-peptidases and their inhibitors in the degradation of the different storage protein fractions (hordeins, albumins and globulins) present in the barley grain. Besides, transgenic barley plants overexpressing or silencing cysteine-peptidases or cystatins were obtained by Agrobacterium-mediated transformation of barley immature embryos to analyze their physiological function in vivo. Preliminary assays were carried out with the T1 grains of several transgenic lines. Comparing the knock-out and the overexpressing lines with the WT, alterations in the germination process were detected and were correlated with their grain hordein content. These data will be validated with the homozygous grains that are being produced through the double haploid technique by microspore culture. Resumen La proteólisis es un proceso metabólico por el cual se lleva a cabo la degradación de las proteínas de un organismo a través de enzimas específicas llamadas proteasas. En plantas, este complejo proceso comprende un entramado de rutas metabólicas que implican, además, diferentes compartimentos subcelulares. En la proteólisis participan numerosas proteasas, principalmente cisteín-, serín-, aspartil-, y metalo-proteasas. Dentro de las cisteín-proteasas, las proteasas tipo papaína o C1A (familia C1, clan CA) están extensamente representadas en plantas terrestres, y se clasifican en catepsinas tipo L, B, H y F. El mecanismo catalítico de estas proteasas está altamente conservado y la triada catalítica formada por los aminoácidos Cys, His y Asn, y a un aminoácido Gln, que parece esencial para el mantenimiento de la conformación activa de la proteína. Las proteasas C1A se sintetizan como precursores inactivos y comprenden un péptido señal en el extremo N-terminal, un pro-péptido y la proteína madura. En cebada hemos identificado 33 cisteín-proteasas de tipo papaína y las hemos clasificado filogenéticamente en 8 grupos diferentes. Cinco de ellos pertenecen a las catepsinas tipo L (5 subgrupos), un grupo a las catepsinas tipo-B, otro a las catepsinas tipo-F y un último a las catepsinas tipo-H. Las proteasas C1A son además las dianas específicas de los inhibidores protéicos de plantas denominados fitocistatinas. El mecanismo de inhibición de las cistatinas está basado en una fuerte interacción reversible. En cebada, se conoce la familia génica completa de las cistatinas, que está formada por 13 miembros. En el presente trabajo se ha investigado el papel de las cisteín-proteasas de cebada y sus inhibidores específicos en el proceso de la germinación de la semilla. Para ello, se seleccionó una proteasa representante de cada grupo/subgrupo (1 catepsina tipo- B, 1 tipo-F, 1 tipo-H, y 5 tipo-L, una por cada subgrupo). Se ha llevado a cabo su caracterización molecular y se ha analizado la interacción enzima-inhibidor tanto in vivo como in vitro. También se han realizado estudios sobre las bases estructurales que demuestran la especificidad en la interacción enzima/propéptido en las proteasas C1A de cebada, mediante ensayos de inhibición y la predicción de modelos estructurales de la interacción. Finalmente, y dado que durante la maduración de la semilla se almacenan proteínas de reserva (prolaminas) en el endospermo que son movilizadas durante la germinación para suministrar los nutrientes necesarios hasta que la nueva planta pueda realizar la fotosíntesis, en este trabajo se ha demostrado la participación de las cisteínproteasas y sus inhibidores en la degradación de las diferentes tipos de proteínas de reserva (hordeinas, albúmins y globulinas) presentes en el grano de cebada. Además, se han obtenido plantas transgénicas de cebada que sobre-expresan o silencian cistatinas y cisteín-proteasas con el fin de analizar la función fisiológica in vivo. Se han realizado análisis preliminares en las semillas T1 de varias líneas tránsgenicas de cebada y al comparar las líneas knock-out y las líneas de sobre-expresión con las silvestres, se han detectado alteraciones en la germinación que están además correlacionadas con el contenido de hordeinas de las semillas. Estos datos serán validados en las semillas homocigotas que se están generando mediante la técnica de dobles haploides a partir del cultivo de microesporas.

Transport properties of CuGaSe(2)-based thin-film solar cells as a function of absorber composition

The transport properties of thin-film solar cells based on wide-gap CuGaSe(2) absorbers have been investigated as a function of the bulk [Ga]/[Cu] ratio ranging from 1.01 to 1.33. We find that (i) the recombination processes in devices prepared from absorbers with a composition close to stoichiometry ([Ga]/[Cu] = 1.01) are strongly tunnelling assisted resulting in low recombination activation energies (E(a)) of approx. 0.95 eV in the dark and 1.36 eV under illumination. (ii) With an increasing [Ga]/[Cu] ratio, the transport mechanism changes to be dominated by thermally activated Shockley-Read-Hall recombination with similar E(a) values of approx. 1.52-1.57 eV for bulk [Ga]/[Cu] ratios of 1.12-1.33. The dominant recombination processes take place at the interface between CdS buffer and CuGaSe(2) absorber independently from the absorber composition. The increase of E(a) with the [Ga]/[Cu] ratio correlates with the open circuit voltage and explains the better performance of corresponding solar cells.

Realistic performance prediction in nanostructured solar cells as a function of nanostructure dimensionality and density

The behavior of quantum dot, quantum wire, and quantum well InAs/GaAs solar cells is studied with a very simplified model based on experimental results in order to assess their performance as a function of the low bandgap material volume fraction fLOW. The efficiency of structured devices is found to exceed the efficiency of a non-structured GaAs cell, in particular under concentration, when fLOW is high; this condition is easier to achieve with quantum wells. If three different quasi Fermi levels appear with quantum dots the efficiency can be much higher.

Non-destructive measurements of the optical properties of apples by means of time-resolved reflectance

Time-resolved reflectance is proposed and effectively used for the nondestructive measurement of the optical properties in apples. The technique is based on the detection of the temporal dispersion of a short laser pulse injected into the probed medium. The time-distribution of re-emitted photons interpreted with a solution of the Diffusion equation yields the mean values of the absorption and reduced scattering coefficients of the medium. The proposed technique proved valuable for the measurement of the absorption and scattering spectra of different varieties of apples. No major variations were observed in the experimental data when the fruit was peeled, proving that the measured optical properties are referred to the pulp. The depth of probed volume was determined to be about 2 cm. Finally, the technique proved capable to follow the change in chlorophyll absorption during storage.

Non-destructive measurements of the optical properties of apples by means of time-resolved reflectance

Time-resolved reflectance is proposed and effectively used for the nondestructive measurement of the optical properties in apples. The technique is based on the detection of the temporal dispersion of a short laser pulse injected into the probed medium. The time-distribution of re-emitted photons interpreted with a solution of the Diffusion equation yields the mean values of the absorption and reduced scattering coefficients of the medium. The proposed technique proved valuable for the measurement of the absorption and scattering spectra of different varieties of apples. No major variations were observed in the experimental data when the fruit was peeled, proving that the measured optical properties are referred to the pulp. The depth of probed volume was determined to be about 2 cm. Finally, the technique proved capable to follow the change in chlorophyll absorption during storage.

The function of the secondary lining in the New Austrian Tunnelling Method

The decision to build the Secondary Lining in Route Tunnels which are perforated using the New Austrian Tunnelling Method does not generally depend on reasons of structural strength. The paper discusses the implications of the issue as well as the pros and cons of the current alternative courses of action.

Piezoelectric and electroacoustic properties of Ti-doped AlN thin films as a function of Ti content

In this work we present the assessment of the structural and piezoelectric properties of Al(0.5-x)TixN0.5 compounds (titanium content menor que6% atomic), which are expected to possess improved properties than conventional AlN films, such as larger piezoelectric activity, thermal stability of frequency and temperature resistance. Al:Ti:N films were deposited from a twin concentric target of Al and Ti by reactive AC sputtering, which provided films with a radial gradient of the Ti concentration. The properties of the films were investigated as a function of their composition, which was measured by electron dispersive energy dispersive X-ray spectroscopy and Rutherford backscattering spectrometry. The microstructure and morphology of the films were assessed by X-ray diffraction and infrared reflectance. Their electroacoustic properties and dielectric constant were derived from the frequency response of BAW test resonators. Al:Ti:N films properties appear to be strongly dependent on the Ti content, which modifies the AlN wurtzite crystal structure leading to greater dielectric constant, lower sound velocities, lower electromechanical factor and moderately improved temperature coefficient of the resonant frequency.

Electron temperature versus laser intensity times wavelength squared: a comparison of theory and experiments

The peak temperature in the corona of plasma ejected by a laser-irradiated slab is discussed in terms of a one-electron-temperature model. Both heat-flux saturation and pulse rise-time effects are considered;the intensity in the rising half of the pulse is approximated by a linear function of time, I(t) = Iot/r. The temperature is found to be proportional to (IQX2)273 and a function of I0X4/r. Above a certain value of I0X4/T, the plasma presents two characteristic temperatures (at saturation and at the critical surface) which can be identified with experimentally observed cold- and hot-electron temperatures. The results are compared with extensive experimental data available for both nd and CO2 lasers, I0(W'cnf2) X2 (/um) starting around 1012. The agreement is good if substantial flux inhibition is assumed (flux-limit factor f = 0.03), and fails for I0X2 above 1O1S. Results for both ablation pressure and mass ablation rate are also given.

New version of a laboratory impact device for firmness sensing of fruits.

Results of previous studies conducted by different researchers have shown that impact techniques can be used to evaluate firmness (Delwiche et al., 1989; Delwiche et al.;1996; Jaren et al., 1992; Ruiz Altisent et al., 1996). To impact the fruit with a small spherical impactor of known mass and radius of curvature and measure the acceleration of the impactor is a technique described by Chen et al. (1985) and used by several researchers for sensing fruit firmness (Jaren et al., 1992; Correa et al.; 1992). The advantages of this method vs. a force sensor that measures the force as a function of time is that the measured impact-acceleration response is independent of the fruit mass and is less sensitive to the variation in the radius of curvature of the fruit (Chen et al., 1996). Ruiz Altisent et al. (1993) developed and used a 50 g impactor with a 19 mm diameter spherical tip, dropping from different height for fruits (apples, pears, avocados, melons, peaches ...). Another impact device for firmness sensing of fruits was developed by Chen and Ruiz Altisent (1996). They designed and fabricated an experimental low-mass impact sensor for high-speed sensing of fruit firmness. The impactor consisted of a semi-spherical impacting tip attached to the end (near the centre of percussion) of a pivoting arm. Impact is done by swinging the impactor to collide with the fruit. It has been implemented for on-line use. In both devices a small accelerometer is mounted behind the impacting tip. Lateral impactor and vertical impactor have been used in laboratory and the results from non-destructive impact tests have contributed to standardise methods to measure fruit firmness: Barreiro (1992) compared impact parameters and results of Magness-Taylor penetration tests for apples, pears, apricots [and peaches; Agulheiro (1994) studied the behaviour of the impact parameters during seven weeks of cold storage of two melon varieties; Ortiz (1998) used low energy impact and NIR procedures to segregate non crispy, non firm and soft peaches. Steinmetz (1996) compared various non-destructive firmness sensors, based on sound, impact and micro-deformation.

Electronic structure of copper nitrides as a function of nitrogen content

he nitrogen content dependence of the electronic properties for copper nitride thin films with an atomic percentage of nitrogen ranging from 26 ± 2 to 33 ± 2 have been studied by means of optical (spectroscopic ellipsometry), thermoelectric (Seebeck), and electrical resistivity measurements. The optical spectra are consistent with direct optical transitions corresponding to the stoichiometric semiconductor Cu3N plus a free-carrier contribution, essentially independent of temperature, which can be tuned in accordance with the N-excess. Deviation of the N content from stoichiometry drives to significant decreases from − 5 to − 50 μV/K in the Seebeck coefficient and to large enhancements, from 10− 3 up to 10 Ω cm, in the electrical resistivity. Band structure and density of states calculations have been carried out on the basis of the density functional theory to account for the experimental results.

Mechanical behavior of tungsten-vanadium-lanthana alloys as function of temperature

The mechanical behavior of three tungsten (W) alloys with vanadium (V) and lanthana (La2O3) additions (W–4%V, W–1%La2O3, W–4%V–1%La2O3) processed by hot isostatic pressing (HIP) have been compared with pure-W to analyze the influence of the dopants. Mechanical characterization was performed by three point bending (TPB) tests in an oxidizing air atmosphere and temperature range between 77 (immersion tests in liquid nitrogen) and 1273 K, through which the fracture toughness, flexural strength, and yield strength as function of temperature were obtained. Results show that the V and La2O3 additions improve the mechanical properties and oxidation behavior, respectively. Furthermore, a synergistic effect of both dopants results in an extraordinary increase of the flexure strength, fracture toughness and resistance to oxidation compared to pure-W, especially at higher temperatures. In addition, a new experimental method was developed to obtain a very small notch tip radius (around 5–7 μm) and much more similar to a crack through the use of a new machined notch. The fracture toughness results were lower than those obtained with traditional machining of the notch, which can be explained with electron microscopy, observations of deformation in the rear part of the notch tip. Finally, scanning electron microscopy (SEM) examination of the microstructure and fracture surfaces was used to determine and analyze the relationship between the macroscopic mechanical properties and the micromechanisms of failure involved, depending on the temperature and the dispersion of the alloy.

Cálculo de la recarga natural en grandes áreas, en función de la litología y la precipitación = Calculation of natural recharge in large areas, as a function of lithology and rainfall

En una región amplia como España se demuestra —mediante inferencias estadísticas sobre una muestra completa de 875 manantiales en los que se conoce su caudal medio y la litología de su área de alimentación y que han sido agrupados en regiones de distinta pluviometría— que la recarga media anual es una fracción fija de la precipitación media para cada litología. Se han establecido así unas tasas de recarga respecto de la precipitación para seis grupos litológicos de diferente permeabilidad: arenas, gravas y formaciones aluviales en general, 8.3%; conglomerados, 5.6%; areniscas, 7.3%; calizas y dolomías, 34.3%; margas, margocalizas, limos y arcillas, 3.3%; otras rocas, 1.3%. Teniendo en cuenta la representatividad de España, la cual tiene una gran variabilidad de litología, pluviometría, topografía, etcétera, estas tasas de recarga respecto de la precipita-ción son probablemente valores cuasi universales que pueden ser utilizados para estimar la recarga media o los recursos hídricos subterráneos medios de regiones amplias en cualquier parte del mundo, salvo en regiones especiales, como las que tienen permafrost, por ejemplo. En todo caso, estas tasas de recarga podrían ser retocadas para cada región según sus particulares características. Los datos de precipitación y litología son muy corrientes, por lo que el método puede ser ampliamente utilizado para completar balances hidráulicos.In a region as large as Spain, annual mean recharge is shown to be a fixed proportion of the mean rainfall for each lithology. This determination is based on statistical inferences from a complete sample of 875 springs for which mean flow and catchment areas are known and which have been grouped into distinct rainfall regions. Recharge rates have thus been established with respect to rainfall for six lithological groups with different permeability: sands, gravels and generally alluvial formations, 8.3%; conglomerates, 5.6%; sandstones, 7.3%; limestone and dolomite 34.3%; marls, marly limestones, silts and clays, 3.3%; and hard rocks, 1.3%. Considering the representativeness of Spain, which is large in size and has a highly varied lithology, topography and rainfall, these recharge rates for rainfall are probably quasi-universal values that can be used to estimate average recharge or average groundwater resources of large regions in any part of the world (except in special cases such as areas with permafrost, for example). For any case, these recharge rates can be adapted to each region according to its particular characteristics. Rainfall and lithology data are very common, and so the method can be widely used to calculate hydraulic balances.

Mechanical behavior of alkali-cement as function of the temperature.

This investigation reports on a comparative study of the mechanical behavior at different temperatures of three different alkali-activated fly ash pastes chemically activated using sodium silicate. A control Portland cement (OPC) was used as a reference. In an attempt to simulate the conditions prevailing in the event of accidental fire, post-thermal mechanical tests were performed to determine the residual strength. It has therefore been established that FA based cements can be fabricated for construction purposes and these materials have great potential for fire resistance applications.

Test Analysis Tool (TATo). A new tool for standardization of time series in maritime and port environment

During the last decades, Puertos del Estado and research groups have developed a huge effort on numerical an physical monitoring. This effort has led to the necessity to implement a tool to standardize, store and process all gathered data. The Test Analysis Tool (TATo) is described in the paper.

Decoupling of soil nutrient cycles as a function of aridity in global drylands

The biogeochemical cycles of carbon (C), nitrogen (N) and phosphorus (P) are interlinked by primary production, respiration and decomposition in terrestrial ecosystems. It has been suggested that the C, N and P cycles could become uncoupled under rapid climate change because of the different degrees of control exerted on the supply of these elements by biological and geochemical processes. Climatic controls on biogeochemical cycles are particularly relevant in arid, semi-arid and dry sub-humid ecosystems (drylands) because their biological activity is mainly driven by water availability. The increase in aridity predicted for the twenty-first century in many drylands worldwide may therefore threaten the balance between these cycles, differentially affecting the availability of essential nutrients. Here we evaluate how aridity affects the balance between C, N and P in soils collected from 224 dryland sites from all continents except Antarctica. We find a negative effect of aridity on the concentration of soil organic C and total N, but a positive effect on the concentration of inorganic P. Aridity is negatively related to plant cover, which may favour the dominance of physical processes such as rock weathering, a major source of P to ecosystems, over biological processes that provide more C and N, such as litter decomposition. Our findings suggest that any predicted increase in aridity with climate change will probably reduce the concentrations of N and C in global drylands, but increase that of P. These changes would uncouple the C, N and P cycles in drylands and could negatively affect the provision of key services provided by these ecosystems.