A crystallographic study of crystalline casts and pseudomorphs from the 3.5†Ga Dresser Formation, Pilbara Craton (Australia).

Crystallography has a long history of providing knowledge and methods for applications in other disciplines. The identification of minerals using X-ray diffraction is one of the most important contributions of crystallography to earth sciences. However, when the crystal itself has been dissolved, replaced or deeply modified during the geological history of the rocks, diffraction information is not available. Instead, the morphology of the crystal cast provides the only crystallographic information on the original mineral phase and the environment of crystal growth. This article reports an investigation of crystal pseudomorphs and crystal casts found in a carbonate-chert facies from the 3.48 Ga-old Dresser Formation (Pilbara Craton, Australia), considered to host some of the oldest remnants of life. A combination of X-ray microtomography, energy-dispersive X-ray spectroscopy and crystallographic methods has been used to reveal the original phases of these Archean pseudomorphs. It is found with a high degree of confidence that the original crystals forming in Archean times were hollow aragonite, the high-temperature polymorphs of calcium carbonate, rather than other possible alternatives such as gypsum (CaSO4·2H20) and nahcolite (NaHCO3). The methodology used is described in detail.

A morphogram for silica-witherite biomorphs and its application to microfossil identification in the early earth rock record.

Archean hydrothermal environments formed a likely site for the origin and early evolution of life. These are also the settings, however, were complex abiologic structures can form. Low-temperature serpentinization of ultramafic crust can generate alkaline, silica-saturated fluids in which carbonate-silica crystalline aggregates with life-like morphologies can self-assemble. These "biomorphs" could have adsorbed hydrocarbons from Fischer-Tropsch type synthesis processes, leading to metamorphosed structures that resemble carbonaceous microfossils. Although this abiogenic process has been extensively cited in the literature and has generated important controversy, so far only one specific biomorph type with a filamentous shape has been discussed for the interpretation of Archean microfossils. It is therefore critical to precisely determine the full distribution in morphology and size of these biomorphs, and to study the range of plausible geochemical conditions under which these microstructures can form. Here, a set of witherite-silica biomorph synthesis experiments in silica-saturated solutions is presented, for a range of pH values (from 9 to 11.5) and barium ion concentrations (from 0.6 to 40 mmol/L BaCl2 ). Under these varying conditions, a wide range of life-like structures is found, from fractal dendrites to complex shapes with continuous curvature. The size, spatial concentration, and morphology of the biomorphs are strongly controlled by environmental parameters, among which pH is the most important. This potentially limits the diversity of environments in which the growth of biomorphs could have occurred on Early Earth. Given the variety of the observed biomorph morphologies, our results show that the morphology of an individual microstructure is a poor criterion for biogenicity. However, biomorphs may be distinguished from actual populations of cellular microfossils by their wide, unimodal size distribution. Biomorphs grown by diffusion in silica gel can be differentiated by their continuous gradient in size, spatial density, and morphology along the direction of diffusion.

Local pH oscillations witness autocatalytic self-organization of biomorphic nanostructures.

Bottom-up self-assembly of simple molecular compounds is a prime pathway to complex materials with interesting structures and functions. Coupled reaction systems are known to spontaneously produce highly ordered patterns, so far observed in soft matter. Here we show that similar phenomena can occur during silica-carbonate crystallization, the emerging order being preserved. The resulting materials, called silica biomorphs, exhibit non-crystallographic curved morphologies and hierarchical textures, much reminiscent of structural principles found in natural biominerals. We have used a fluorescent chemosensor to probe local conditions during the growth of such self-organized nanostructures. We demonstrate that the pH oscillates in the local microenvironment near the growth front due to chemical coupling, which becomes manifest in the final mineralized architectures as intrinsic banding patterns with the same periodicity. A better understanding of dynamic autocatalytic crystallization processes in such simple model systems is key to the rational development of advanced materials and to unravel the mechanisms of biomineralization.

Diffusion and precipitation processes in iron-based silica gardens.

Silica gardens are tubular structures that form along the interface of multivalent metal salts and alkaline solutions of sodium silicate, driven by a complex interplay of osmotic and buoyant forces together with chemical reaction. They display peculiar plant-like morphologies and thus can be considered as one of the few examples for the spontaneous biomimetic self-ordering of purely inorganic materials. Recently, we could show that silica gardens moreover are highly dynamic systems that remain far from equilibrium for considerable periods of time long after macroscopic growth is completed. Due to initial compartmentalisation, drastic concentration gradients were found to exist across the tube walls, which give rise to noticeable electrochemical potential differences and decay only slowly in a series of coupled diffusion and precipitation processes. In the present work, we extend these studies and investigate the effect of the nature of the used metal cations on the dynamic behaviour of the system. To that end, we have grown single macroscopic silica garden tubes by controlled addition of sodium silicate sol to pellets of iron(ii) and iron(iii) chloride. In the following, the concentrations of ionic species were measured as a function of time on both sides of the formed membranes, while electrochemical potentials and pH were monitored online by immersing the corresponding sensors into the two separated solution reservoirs. At the end of the experiments, the solid tube material was furthermore characterised with respect to composition and microstructure by a combination of ex situ techniques. The collected data are compared to the previously reported case of cobalt-based silica gardens and used to shed light on ion diffusion through the inorganic membranes as well as progressive mineralisation at both surfaces of the tube walls. Our results reveal important differences in the dynamics of the three studied systems, which can be explained based on the acidity of the metal cations and the porosity of the membranes, leading to substantially dissimilar time-dependent solution chemistry as well as distinct final mineral structures. The insight gained in this work may help to better understand the diffusion properties and precipitation patterns in tubular iron (hydr)oxide/silicate structures observed in geological environments and during steel corrosion.

Impact of climate and land use change on water availability and reservoir management: scenarios in the Upper Aragón River, Spanish Pyrenees.

Streamflows in a Mediterranean mountain basin in the central Spanish Pyrenees were projected under various climate and land use change scenarios. Streamflow series projected for 2021-2050 were used to simulate the management of the Yesa reservoir, which is critical to the downstream supply of irrigation and domestic water. Streamflows were simulated using the Regional Hydro-Ecologic Simulation System (RHESSys). The results show that increased forest cover in the basin could decrease annual streamflow by 16%, mainly in early spring, summer and autumn. Regional climate models (RCMs) project a trend of warming and drying in the basin for the period 2021-2050, which will cause a 13.8% decrease in annual streamflow, mainly in late spring and summer. The combined effects of forest regeneration and climate change are expected to reduce annual streamflows by 29.6%, with marked decreases affecting all months with the exception of January and February, when the decline will be moderate. Under these streamflow reduction scenarios it is expected that it will be difficult for the Yesa reservoir to meet the current water demand, based on its current storage capacity (476 hm(3)). If the current project to enlarge the reservoir to a capacity of 1059 hm(3) is completed, the potential to apply multi-annual streamflow management, which will increase the feasibility of maintaining the current water supply. However, under future climate and land cover scenarios, reservoir storage will rarely exceed half of the expected capacity, and the river flows downstream of the reservoir is projected to be dramatically reduced.

Enantiospecific synthesis of heterocycles linked to purines: different apoptosis modulation of enantiomers in breast cancer cells.

The issue of chiral drug is now a major theme in the design, discovery and development of new drugs. It has been shown for many pharmaceuticals that only one enantiomer contains the desired activity, and the synthesis of such drug molecules in their optically pure form is becoming increasingly important. Mitsunobu reaction was carried out between (R)- and (S)-3,4-dihydro-2H-1,5-benzoxathiepin-3-ol and purines under microwave irradiation. A contraction into a six-membered ring takes place with concomitant inversion at the stereocentre with excellent enatiomeric excesses giving rise to the homochiral 9-(2,3-dihydro-1,4-benzoxathiin-3-ylmethyl)-9H-purines. The anti-tumour activity of all enantiomers is reported against the caspase-3-deficient MCF-7 and the wild type SKBR-3 human breast cancer cells. The most active homochiral compound displays an IC50 of 1.85 µM and induces inhibition of the translation initiation factor eIF2α. All homochiral compounds included in this study show different apoptotic effects between both enantiomers with levels up to 99%. We have analyzed caspase-mediated apoptotic pathways on enantiomers and racemates. We have found a homochiral derivative that activates the canonical intrinsic caspase-8/caspase-3 apoptotic pathway on the MCF-7 cells, and a racemic compound that induces caspase-2 activation. Moreover, we demonstrate the involvement of caspase activation during cell death induced by these compounds in SKBR-3 cells.

In situ X-ray data collection from highly sensitive crystals of Pseudomonas putida PtxS in complex with DNA.

Pseudomonas putida PtxS is a member of the LacI protein family of transcriptional regulators involved in glucose metabolism. All genes involved in this pathway are clustered into two operons, kgu and gad. PtxS controls the expression of the kgu and gad operons as well as its own transcription. The PtxS operator is a perfect palindrome, 5'-TGAAACCGGTTTCA-3', which is present in all three promoters. Crystallization of native PtxS failed, and PtxS-DNA crystals were finally produced by the counter-diffusion technique. A portion of the capillary used for crystal growth was attached to the end of a SPINE standard cap and directly flash-cooled in liquid nitrogen for diffraction tests. A full data set was collected with a beam size of 10×10 µm. The crystal belonged to the trigonal space group P3, with unit-cell parameters a=b=213.71, c=71.57 Å. Only unhandled crystals grown in capillaries of 0.1 mm inner diameter diffracted X-rays to 1.92 Šresolution.

The role and implications of bassanite as a stable precursor phase to gypsum precipitation.

Calcium sulfate minerals such as gypsum play important roles in natural and industrial processes, but their precipitation mechanisms remain largely unexplored. We used time-resolved sample quenching and high-resolution microscopy to demonstrate that gypsum forms via a three-stage process: (i) homogeneous precipitation of nanocrystalline hemihydrate bassanite below its predicted solubility, (ii) self-assembly of bassanite into elongated aggregates co-oriented along their c axis, and (iii) transformation into dihydrate gypsum. These findings indicate that a stable nanocrystalline precursor phase can form below its bulk solubility and that in the CaSO(4) system, the self-assembly of nanoparticles plays a crucial role. Understanding why bassanite forms prior to gypsum can lead to more efficient anti-scaling strategies for water desalination and may help to explain the persistence of CaSO(4) phases in regions of low water activity on Mars.

Crystallization and crystallographic analysis of the ligand-binding domain of the Pseudomonas putida chemoreceptor McpS in complex with malate and succinate.

Methyl-accepting chemotaxis proteins (MCPs) are transmembrane proteins that sense changes in environmental signals, generating a chemotactic response and regulating other cellular processes. MCPs are composed of two main domains: a ligand-binding domain (LBD) and a cytosolic signalling domain (CSD). Here, the crystallization of the LBD of the chemoreceptor McpS (McpS-LBD) is reported. McpS-LBD is responsible for sensing most of the TCA-cycle intermediates in the soil bacterium Pseudomonas putida KT2440. McpS-LBD was expressed, purified and crystallized in complex with two of its natural ligands (malate and succinate). Crystals were obtained by both the counter-diffusion and the hanging-drop vapour-diffusion techniques after pre-incubation of McpS-LBD with the ligands. The crystals were isomorphous and belonged to space group C2, with two molecules per asymmetric unit. Diffraction data were collected at the ESRF synchrotron X-ray source to resolutions of 1.8 and 1.9 Å for the malate and succinate complexes, respectively.

Pattern formation in stromatolites: insights from mathematical modelling.

To this day, computer models for stromatolite formation have made substantial use of the Kardar-Parisi-Zhang (KPZ) equation. Oddly enough, these studies yielded mutually exclusive conclusions about the biotic or abiotic origin of such structures. We show in this paper that, at our current state of knowledge, a purely biotic origin for stromatolites can neither be proved nor disproved by means of a KPZ-based model. What can be shown, however, is that whatever their (biotic or abiotic) origin might be, some morphologies found in actual stromatolite structures (e.g. overhangs) cannot be formed as a consequence of a process modelled exclusively in terms of the KPZ equation and acting over sufficiently large times. This suggests the need to search for alternative mathematical approaches to model these structures, some of which are discussed in this paper.

Ultraslow growth rates of giant gypsum crystals.

Mineralogical processes taking place close to equilibrium, or with very slow kinetics, are difficult to quantify precisely. The determination of ultraslow dissolution/precipitation rates would reveal characteristic timing associated with these processes that are important at geological scale. We have designed an advanced high-resolution white-beam phase-shift interferometry microscope to measure growth rates of crystals at very low supersaturation values. To test this technique, we have selected the giant gypsum crystals of Naica ore mines in Chihuahua, Mexico, a challenging subject in mineral formation. They are thought to form by a self-feeding mechanism driven by solution-mediated anhydrite-gypsum phase transition, and therefore they must be the result of an extremely slow crystallization process close to equilibrium. To calculate the formation time of these crystals we have measured the growth rates of the {010} face of gypsum growing from current Naica waters at different temperatures. The slowest measurable growth rate was found at 55 °C, 1.4 ± 0.2 × 10(-5) nm/s, the slowest directly measured normal growth rate for any crystal growth process. At higher temperatures, growth rates increase exponentially because of decreasing gypsum solubility and higher kinetic coefficient. At 50 °C neither growth nor dissolution was observed indicating that growth of giant crystals of gypsum occurred at Naica between 58 °C (gypsum/anhydrite transition temperature) and the current temperature of Naica waters, confirming formation temperatures determined from fluid inclusion studies. Our results demonstrate the usefulness of applying advanced optical techniques in laboratory experiments to gain a better understanding of crystal growth processes occurring at a geological timescale.

Structure of the mexicain-E-64 complex and comparison with other cysteine proteases of the papain family.

Mexicain is a 23.8 kDa cysteine protease from the tropical plant Jacaratia mexicana. It is isolated as the most abundant product after cation-exchange chromatography of the mix of proteases extracted from the latex of the fruit. The purified enzyme inhibited with E-64 [N-(3-carboxyoxirane-2-carbonyl)-leucyl-amino(4-guanido)butane] was crystallized by sitting-drop vapour diffusion and the structure was solved by molecular replacement at 2.1 A resolution and refined to an R factor of 17.7% (R(free) = 23.8%). The enzyme belongs to the alpha+beta class of proteins and the structure shows the typical papain-like fold composed of two domains, the alpha-helix-rich (L) domain and the beta-barrel-like (R) domain, separated by a groove containing the active site formed by residues Cys25 and His159, one from each domain. The four monomers in the asymmetric unit show one E-64 molecule covalently bound to Cys25 in the active site and differences have been found in the placement of E-64 in each monomer.

Changes in eggshell mechanical properties, crystallographic texture and in matrix proteins induced by moult in hens.

The effect of moult on eggshell mechanical properties, on composition and concentrations of organic matrix components and on eggshell microstructure was investigated. The observed changes were studied to understand the role of organic matrix and eggshell microstructure in eggshell strength. Moult was induced by zinc oxide (20 g zinc/kg diet) in 53 ISA Brown laying hens at 78 weeks of age. No difference was observed for egg or eggshell weights after moult. In contrast, moult improved the shell breaking strength (28.09 vs 33.71 N). After moult, there was a decrease in the average size of calcite crystals composing the eggshell and in their heterogeneity, whereas crystal orientation remained basically the same. After moulting, the total protein concentration in eggshell increased slightly. The comparisons of SDS-PAGE profiles of the organic matrix constituents extracted before and after moulting showed changes in staining intensity of certain bands. After moult, bands associated with main proteins specific to eggshell formation (OC-116 and OC-17) showed higher staining intensity, while the intensity of the egg white proteins (ovotransferrin, ovalbumin and lysozyme) decreased. ELISA confirmed the decrease in ovotransferrin after moult. Its concentration was inversely correlated with breaking strength before moult. These observations suggest that changes in eggshell crystal size could be due to changes in organic matrix composition. These changes may provide a mechanism for the improvement in shell solidity after moulting.

Purification, crystallization and preliminary X-ray analysis of mexicain.

Mexicain is a 23.7 kDa papain-like cysteine protease from the tropical plant Jacaratia mexicana. Extracted as a mix of proteases from the latex of the fruit, mexicain is isolated after cation-exchange chromatography as the most abundant product. The purified product inhibited with E-64 was crystallized by sitting-drop vapour diffusion in the presence of ethanolamine. Cryoprotected crystals diffracted X-rays from a home source to 1.98 A and belong to the monoclinic space group P2(1), with unit-cell parameters a = 57.36, b = 90.45, c = 80.39 A, beta = 92.64 degrees . The asymmetric unit contains four molecules of mexicain, with a corresponding crystal volume per protein weight (V(M)) of 2.24 A(3) Da(-1) and a solvent content of 45% by volume. A molecular-replacement model has been determined and refinement is in progress.

Structural study of the type II 3-dehydroquinate dehydratase from Actinobacillus pleuropneumoniae.

The structure of the type II dehydroquinate dehydratase (DHQase) from Actinobacillus pleuropneumoniae, the third enzyme of the shikimate pathway, has been determined. Crystals diffracting to 1.7 A were obtained in space and on earth using the counter-diffusion technique. The structure was solved using molecular replacement and refined to high resolution. The overall structure of the dodecameric enzyme is described and compared with structures of DHQases from other bacteria. DHQases contain a flexible loop that presumably closes over the active site upon substrate binding. The enzyme can exist in an open or closed conformation. The present structure displays the open conformation, with a sulfate anion bound in the active site. The availability of this structure opens a route to structure-based antibiotics targetting this pathogenic bacterium.

Self-assembled silica-carbonate structures and detection of ancient microfossils.

We have synthesized inorganic micron-sized filaments, whose microstucture consists of silica-coated nanometer-sized carbonate crystals, arranged with strong orientational order. They exhibit noncrystallographic, curved, helical morphologies, reminiscent of biological forms. The filaments are similar to supposed cyanobacterial microfossils from the Precambrian Warrawoona chert formation in Western Australia, reputed to be the oldest terrestrial microfossils. Simple organic hydrocarbons, whose sources may also be abiotic and indeed inorganic, readily condense onto these filaments and subsequently polymerize under gentle heating to yield kerogenous products. Our results demonstrate that abiotic and morphologically complex microstructures that are identical to currently accepted biogenic materials can be synthesized inorganically.

Theory and simulation of buoyancy-driven convection around growing protein crystals in microgravity.

We present an order-of-magnitude analysis of the Navier-Stokes equations in a time-dependent, incompressible and Boussinesq formulation. The hypothesis employed of two different length scales allows one to determine the different flow regimes on the basis of the geometrical and thermodynamical parameters alone, without solving the Navier-Stokes equations. The order-of-magnitude analysis is then applied to the field of protein crystallization, and to the flow field around a crystal, where the driving forces are solutal buoyancy-driven convection, from density dependence on species concentration, and sedimentation caused by the different densities of the crystal and the protein solution. The main result of this paper is to provide predictions of the conditions in which a crystal is growing in a convective regime, rather than in the ideal diffusive state, even under the typical microgravity conditions of space platforms.

Influence of the microstructure on the shell strength of eggs laid by hens of different ages.

1. The eggshell is a bioceramic material constructed of columnar calcite crystals preferentially oriented with their c-axis perpendicular to the shell surface. 2. The influence of microstructure (crystal size, shape and crystallographic orientation of crystal grains) on the mechanical properties of eggshells (shell strength) was investigated using eggs from hens of different ages. 3. There was a strong correlation between crystallographic texture and the strength of the eggshell in the case of eggs laid by young hens. The strength of eggshells increased as the preferential orientation of the crystals constituting the eggshell decreased. 4. By comparing two age populations, the effect of hen age on eggshell properties was evaluated. In general, eggshells from aged hens had a lower breaking strength (less than half that of those laid by young hens) and showed a greater variability in their structural properties such as thickness, grain morphology and crystallographic texture. 5. Texture analysis revealed that shells from eggs laid by aged hens have two preferred crystal orientations, after (001) and (104), compared with mainly one, after (001), in eggs laid by young hens. 6. These observed changes in eggshell properties could be due to changes in the organic matrix of the eggshell associated with ageing of the hens.