Understanding the specific molecular interactions between proteins and $\beta$1,3-1,4-mixed-linked d-glucans is fundamental to harvest the full biological and biotechnological potential of these carbohydrates and of proteins that specifically recognize them. The family 11 carbohydrate-binding module from Clostridium thermocellum (CtCBM11) is known for its binding preference for $\beta$1,3-1,4-mixed-linked over $\beta$1,4-linked glucans. Despite the growing industrial interest of this protein for the biotransformation of lignocellulosic biomass, the molecular determinants of its ligand specificity are not well defined. In this report, a combined approach of methodologies was used to unravel, at a molecular level, the ligand recognition of CtCBM11. The analysis of the interaction by carbohydrate microarrays and NMR and the crystal structures of CtCBM11 bound to $\beta$1,3-1,4-linked glucose oligosaccharides showed that both the chain length and the position of the $\beta$1,3-linkage are important for recognition, and identified the tetrasaccharide Glc$\beta$1,4Glc$\beta$1,4Glc$\beta$1,3Glc sequence as a minimum epitope required for binding. The structural data, along with site-directed mutagenesis and ITC studies, demonstrated the specificity of CtCBM11 for the twisted conformation of $\beta$1,3-1,4-mixed-linked glucans. This is mediated by a conformation-selection mechanism of the ligand in the binding cleft through CH-$π$ stacking and a hydrogen bonding network, which is dependent not only on ligand chain length, but also on the presence of a $\beta$1,3-linkage at the reducing end and at specific positions along the $\beta$1,4-linked glucan chain. The understanding of the detailed mechanism by which CtCBM11 can distinguish between linear and mixed-linked $\beta$-glucans strengthens its exploitation for the design of new biomolecules with improved capabilities and applications in health and agriculture. DATABASE: Structural data are available in the Protein Data Bank under the accession codes 6R3M and 6R31.

One of the trends in downstream processing comprises the use of ?anything-but-chromatography? methods to overcome the current downfalls of standard packed-bed chromatography. Precipitation and magnetic separation are two techniques already proven to accomplish protein purification from complex media, yet never used in synergy. With the aim to capture antibodies directly from crude extracts, a new approach combining precipitation and magnetic separation was developed and named as affinity magnetic precipitation. A precipitation screening, based on the Hofmeister series, and a commercial precipitation kit were tested with affinity magnetic particles to assess the best condition for antibody capture from human serum plasma and clarified cell supernatant. The best conditions were obtained when using PEG3350 as precipitant at 4°C for 1h, reaching 80% purity and 50% recovery of polyclonal antibodies from plasma, and 99% purity with 97% recovery yield of anti-TNFα mAb from cell supernatants. These results show that the synergetic use of precipitation and magnetic separation can represent an alternative for the efficient capture of antibodies. This article is protected by copyright. All rights reserved

The aim of this study is to evaluate the elemental composition of six food supplements of plant origin, commonly sold in the Portuguese market, by energy dispersive X-ray fluorescence. The presence of arsenic in all the Maca, Ashwagandha, Camu-Camu and Hemp protein samples (except the generic form) is a reason of concern due to the long-term effects of As mainly in its inorganic form. Thus, great caution must be taken on some food supplements, particularly the cases of Moringa from Egypt and Yellow/Xpresso Maca, whose inorganic As concentrations are in line with the upper bound concentration for the 95th dietary exposure according to the European Food Safety Authority which is 0.64 μg/kg bw/day. In what regards Hemp protein, if the supplier’s daily intake recommendation (30 g) is followed, values as high as 1.75 μg/kg bw/day of inorganic As will be consumed, which are dangerously above the upper bound. In this case this specific supplement lot should be removed from the market. Also the consumption of Hemp protein leads to a daily intake of Mn above the Daily Reference Intake (DRI) and Adequate Intakes (AIs) for adults. The contamination of Goji berries by Pb is a reason for concern—organic berries contained 11.3 μg/g while berries derived from conventional agriculture 11.6 μg/g, leading to daily intake doses of 315.3 μg and 324.8 μg, respectively, if the recommended daily intake of 28 g is followed. Our findings point out to an inadequacy of the recommended intakes by the supplier vis a vis the concentrations observed, greatly increasing the risk for public health.

Understanding the specific molecular interactions between proteins and β1,3-1,4-mixed-linked d-glucans is fundamental to harvest the full biological and biotechnological potential of these carbohydrates and of proteins that specifically recognize them. The family 11 carbohydrate-binding module from Clostridium thermocellum (CtCBM11) is known for its binding preference for β1,3-1,4-mixed-linked over β1,4-linked glucans. Despite the growing industrial interest of this protein for the biotransformation of lignocellulosic biomass, the molecular determinants of its ligand specificity are not well defined. In this report, a combined approach of methodologies was used to unravel, at a molecular level, the ligand recognition of CtCBM11. The analysis of the interaction by carbohydrate microarrays and NMR and the crystal structures of CtCBM11 bound to β1,3-1,4-linked glucose oligosaccharides showed that both the chain length and the position of the β1,3-linkage are important for recognition, and identified the tetrasaccharide Glcβ1,4Glcβ1,4Glcβ1,3Glc sequence as a minimum epitope required for binding. The structural data, along with site-directed mutagenesis and ITC studies, demonstrated the specificity of CtCBM11 for the twisted conformation of β1,3-1,4-mixed-linked glucans. This is mediated by a conformation–selection mechanism of the ligand in the binding cleft through CH-π stacking and a hydrogen bonding network, which is dependent not only on ligand chain length, but also on the presence of a β1,3-linkage at the reducing end and at specific positions along the β1,4-linked glucan chain. The understanding of the detailed mechanism by which CtCBM11 can distinguish between linear and mixed-linked β-glucans strengthens its exploitation for the design of new biomolecules with improved capabilities and applications in health and agriculture. Database Structural data are available in the Protein Data Bank under the accession codes 6R3M and 6R31.

Deterioration of adhesively bonded CFRP/steel systems in salt fog environment, i.e., deicing salts and ocean environments, has to be taken into account in the design of steel strengthened structures. In the present work, monotonic and quasi-static cyclic loading were applied to CFRP-to-steel double strap joints for two kinds of CFRP laminates after being aged for a period of 5000 h to evaluate the bond behavior. The bonded joints exposed to salt fog had a different failure mode than that observed in the control specimens (0 h of exposure). The severe reduction of the maximum bond stress resulted from damage initiation that occurred in the corrosion region of the steel substrate, associated with final partial rupture on the corroded steel substrate around the edge of the bonded area: it was also correlated with reduced load carrying capacity. Results of pseudo-cyclic tests showed that the relationship between a local damage parameter (D) and normalized local dissipated energy (Wd/Gf) and the normalized slip increment (ΔS/ΔSult) exhibited almost the same trend in the un-aged and aged bonded joints. The normalized slip increment can be seen as a direct indicator for the local and global damage for the un-aged and aged bonded joints. However, monotonic and quasi-static cyclic tests results revealed that the stress concentration due to local corrosion of steel substrate could lead to brittle rupture or accelerated cumulative damage once the aged bonded interface had become weaker. The bonded joints have exhibited also a smaller relative deformation capacity between CFRP and steel.

Os edifícios históricos como os palácios, castelos, igrejas e templos, são exemplos do património construído e constituem uma parte importante do nosso património cultural. Estas construções históricas incluem normalmente paredes de alvenaria maciça e pedra natural. A salvaguarda do património construído em pedra é de importância inquestionável, pelo que devem ser desenvolvidas e estudadas técnicas de intervenção adequadas em materiais e estruturas. Neste artigo, é apresentada uma metodologia de ensaios não destrutivos para avaliar a conservação de edifícios históricos de pedra e, com base nesta metodologia, é mostrado um caso de estudo na Igreja de São Leonardo, monumento português construído na Atouguia da Baleia no século XIII.

Proof plays a central role in developing, establishing, and communicating mathematical knowledge. Nevertheless it is not such a central element in school mathematics. This article discusses some issues involving mathematical proof in school, intending to characterize the understanding of mathematical proof in school, its function and the meaning and relevance attributed to the notion of simple proof. The main conclusions suggest that the idea of addressing mathematical proof at all levels of school is a recent idea that is not yet fully implemented in schools. It requires an adaptation of the understanding of proof to the age of the students, reducing the level of formality, and allowing the students to experience the different functions of proof and not only the function of verification. Among the different functions of proof, the function of explanation deserves special attention due to the illumination and empowerment that it can bring to the students and their learning. The way this function of proof relates to the notion of simple proof (and the related aesthetic issues) seems relevant enough to make it, in the future, a focus of attention for the teachers who address mathematical proof in the classroom.

The left patience sorting (lPS) monoid, also known in the literature as the Bell monoid, and the right patient sorting (rPS) monoid are introduced by defining certain congruences on words. Such congruences are constructed using insertion algorithms based on the concept of decreasing subsequences.
Presentations for these monoids are given.

Each finite-rank rPS monoid is shown to have polynomial growth and to satisfy a non-trivial identity (dependent on its rank), while the infinite rank rPS monoid does not satisfy a non-trivial identity. The lPS monoids of finite rank have exponential growth and thus do not satisfy non-trivial identities. The
complexity of the insertion algorithms is discussed.

rPS monoids of finite rank are shown to be automatic and to have recursive complete presentations. When the rank is $1$ or $2$, they are also biautomatic. lPS monoids of finite rank are shown to have finite complete presentations and to be biautomatic.

Assessing wood fracture behavior is essential in the design of structural timber elements and connections. This is particularly the case for connections with the possibility of brittle splitting failure. The numerical cohesive zone models that are used to simulate the fracture behavior of wood make it necessary to assume a cohesive law of the material that relates cohesive tractions and crack opening displacements ahead of the crack tip. This work addresses the determination of the fracture cohesive laws of Eucalyptus globulus, a hardwood species with great potential in timber engineering. This study centres on Mode I fracture loading for RL and TL crack propagation systems using Double Cantilever Beam tests. The Compliance-Based Beam Method is applied as the data reduction scheme in order to obtain the strain energy release rate from the load-displacement curves. The cohesive laws are determined by differentiating the relationship between strain energy release rate and crack tip opening displacement. The latter is measured by the digital image correlation technique. High strain energy release rates were obtained for this species, with no big differences between crack propagation systems. The difference between the crack systems is somewhat more pronounced in terms of maximum stress that determines the respective cohesive laws.