Crowding alone cannot account for cosolute effect on amyloid aggregation

Amyloid fiber formation is a specific form of protein aggregation, often resulting from the misfolding of native proteins. Aimed at modeling the crowded environment of the cell, recent experiments showed a reduction in fibrillation halftimes for amyloid-forming peptides in the presence of cosolutes that are preferentially excluded from proteins and peptides. The effect of excluded cosolutes has previously been attributed to the large volume excluded by such inert cellular solutes, sometimes termed "macromolecular crowding". Here, we studied a model peptide that can fold to a stable monomeric β-hairpin conformation, but under certain solution conditions aggregates in the form of amyloid fibrils. Using Circular Dichroism spectroscopy (CD), we found that, in the presence of polyols and polyethylene glycols acting as excluded cosolutes, the monomeric β-hairpin conformation was stabilized with respect to the unfolded state. Stabilization free energy was linear with cosolute concentration, and grew with molecular volume, as would also be predicted by crowding models. After initiating the aggregation process with a pH jump, fibrillation in the presence and absence of cosolutes was followed by ThT fluorescence, transmission electron microscopy, and CD spectroscopy. Polyols (glycerol and sorbitol) increased the lag time for fibril formation and elevated the amount of aggregated peptide at equilibrium, in a cosolute size and concentration dependent manner. However, fibrillation rates remained almost unaffected by a wide range of molecular weights of soluble polyethylene glycols. Our results highlight the importance of other forces beyond the excluded volume interactions responsible for crowding that may contribute to the cosolute effects acting on amyloid formation.     

Mathematical modeling of nucleotide excision repair reveals efficiency of sequential assembly strategies

Nucleotide excision repair (NER) requires the concerted action of many different proteins that assemble at sites of damaged DNA in a sequential fashion. We have constructed a mathematical model delineating hallmarks and general characteristics for NER. We measured the assembly kinetics of the putative damage-recognition factor XPC-HR23B at sites of DNA damage in the nuclei of living cells. These and other in vivo kinetic data allowed us to scrutinize the dynamic behavior of the nucleotide excision repair process in detail. A sequential assembly mechanism appears remarkably advantageous in terms of repair efficiency. Alternative mechanisms for repairosome formation, including random assembly and preassembly, can readily become kinetically unfavorable. Based on the model, new experiments can be defined to gain further insight into this complex process and to critically test model predictions. Our work provides a kinetic framework for NER and rationalizes why many multiprotein processes within the cell nucleus show sequential assembly strategy.     

Useful applications of DOSY experiments for the study of mushroom polysaccharides

DOSY analysis has been performed on different mushroom extracts and fractions with the aim to describe a general method for the study of high molecular weight polysaccharides. These NMR experiments can be exploited to monitor the fractionation pathways performed on crude extracts in order to isolate polysaccharides. DOSY can also rapidly verify the purity of the isolated compounds, as well as evaluate their molecular size. In spite of the complexity of DOSY spectra of mixtures, this NMR technique seems to be a valid analytical tool that could be adopted as a routine method for the study of polysaccharides from different sources.     

Positive interaction between invasive plants: The influence of Pyracantha angustifolia on the recruitment of native and exotic woody species

Abstract: Positive interactions between species are known to play an important role in the dynamics of plant communities, including the enhancement of invasions by exotics. We studied the influence of the invasive shrub Pyracantha angustifolia (Rosaceae) on the recruitment of native and exotic woody species in a secondary shrubland in central Argentina mountains. We recorded woody sapling recruitment and micro‐environmental conditions under the canopies of Pyracantha and the dominant native shrub Condalia montana (Rhamnaceae), and in the absence of shrub cover, considering these situations as three treatments. We found that native and exotic species richness were higher under Pyracantha than under the other treatments. Ligustrum lucidum (Oleaceae), an exotic bird‐dispersed shade‐tolerant tree, was the most abundant species recruiting in the area, and its density was four times higher under the canopy of Pyracantha. This positive interaction may be related to Pyracantha's denser shading, to the mechanical protection of its canopy against ungulates, and/or to the simultaneous fruit ripening of both woody invaders.     

Biological traits of stream macroinvertebrates from a semi-arid catchment: patterns along complex environmental gradients

1. The relationships between biological traits of macroinvertebrates and environmental characteristics were investigated in streams with contrasting physical, chemical or landscape level attributes. We used an ordination technique, RLQ analysis, which links an environmental table (R) with traits table (Q) through an abundance table (L) to investigate the relationship between habitat characteristics and biological traits.
2. A major environmental axis explaining the distribution of species and their distinctive biological features was obtained. This axis included variables of anthropogenic pressure (agricultural and urban uses) and natural variability (climatic and geologic) that are strongly intercorrelated in the study area, with a clear spatial component.
3. The attributes of species from frequently disturbed systems (small size, multivoltinism, diapause, ovoviviparity, etc.) were associated with semi-arid areas whereas traits common in more stable and favourable environments (large body size, semi-voltinism, isolated eggs, etc.) were found in upland forested areas.
4. The natural climatic variation was proposed as a disturbance axis of a theoretical habitat templet (driven by the intense hydrological disturbances typical of semi-arid streams), while anthropogenic pressure (mainly intensive agriculture) and high salinity, a natural consequence of geology, was proposed as an adversity axis. Different life-histories associated with contrasting environmental features were superimposed in this habitat templet.
5. The ecological–evolutionary scenario in which stream macroinvertebrates have evolved and by which their communities are organized, is closely linked to disturbance, environmental harshness and human pressure.     

Synthesis and preliminary biological characterization of a new potential I-Radioligand for dopamine and serotonin receptors

The synthesis and a preliminary biological characterization of a new class of N-benzyl-aminoalcohols which have serotonin (5-HT₂) and dopamine (D₂) receptor affinity is described. In vitro competition binding studies were conducted with the new molecules and ³H-spiperone on crude membrane preparation from rat striatum and frontal cortex. One of these compounds, 3-benzylamino-1-(4-fluoro-2-iodophenyl)-propan-1-ol (6f), whose IC₅₀ values are in the micromolar range for both the D₂ and 5-HT₂ receptors, was prepared in iodine-125 labelled form (6i) by nucleophilic substitution of the bromine atom of 3-benzylamino-1-(2-bromo-4-fluorophenyl)-propan-1-ol (6d). In the in vivo studies, conducted on rats, the radiolabelled molecule 6i shows a good capacity to cross the blood–brain barrier (BBB) with a mean value of first pass cerebral extraction (E) of ca. 50% when the regional cerebral blood flow, measured with microsphere technique, is in the experimental animal's physiologic range (0.8–1 mL/min/g). A preliminary in vitro autoradiographic distribution on coronal rat brain slices of the radioiodinated molecule showed that it was preferentially localized in the striatum and in the cerebral regions rich in dopamine- and serotonin receptors, even if a high non-specific binding was observed.