Spermidine improves fear memory persistence.

Persistence is the most characteristic attribute of long-term memory (LTM). For memory persistence, a second late event of consolidation, that occurs around 12h after the acquisition, is necessary. Although the N-methyl-d-aspartate (NMDA) receptor has been involved in the persistence of memory, whether endogenous modulators of the NMDA receptor actually modulate memory persistence is unknown. In the current study we investigated whether spermidine and arcaine, respectively agonist and antagonist of polyamine binding site at NMDA receptor, alter the persistence of the memory of contextual fear conditioning task in rats. While 12h post-training administration of spermidine (10 and 30mg/kg, i.p.) facilitated, arcaine (10mg/kg, i.p.) impaired the memory of fear assessed 2 and 7 days after training. Arcaine (0.1mg/kg) prevented the facilitatory effect of spermidine (10mg/kg, i.p.), and spermidine (1mg/kg), prevented the memory impairment induced by arcaine (10mg/kg, i.p.) when tested 2 and 7 days after training. These results suggest that endogenous polyamines improve the persistence of fear memory.

The Penicillium echinulatum secretome on sugar cane bagasse.

Plant feedstocks are at the leading front of the biofuel industry based on the potential to promote economical, social and environmental development worldwide through sustainable scenarios related to energy production. Penicillium echinulatum is a promising strain for the bioethanol industry based on its capacity to produce large amounts of cellulases at low cost. The secretome profile of P. echinulatum after grown on integral sugarcane bagasse, microcrystalline cellulose and three types of pretreated sugarcane bagasse was evaluated using shotgun proteomics. The comprehensive chemical characterization of the biomass used as the source of fungal nutrition, as well as biochemical activity assays using a collection of natural polysaccharides, were also performed. Our study revealed that the enzymatic repertoire of P. echinulatum is geared mainly toward producing enzymes from the cellulose complex (endogluganases, cellobiohydrolases and ß-glucosidases). Glycoside hydrolase (GH) family members, important to biomass-to-biofuels conversion strategies, were identified, including endoglucanases GH5, 7, 6, 12, 17 and 61, ß-glycosidase GH3, xylanases GH10 and GH11, as well as debranching hemicellulases from GH43, GH62 and CE2 and pectinanes from GH28. Collectively, the approach conducted in this study gave new insights on the better comprehension of the composition and degradation capability of an industrial cellulolytic strain, from which a number of applied technologies, such as biofuel production, can be generated.

Protective effects of anthocyanins on the ectonucleotidase activity in the impairment of memory induced by scopolamine in adult rats.

AIMS: We investigated whether the treatment with anthocyanins prevents the scopolamine-induced memory deficits and whether ectonucleotidase activities and purine levels are altered in the cerebral cortex (CC) and hippocampus (HC) in this model of mnemonic deficit in rats. MAIN METHODS: The animals were divided into 4 experimental groups: control (vehicle), anthocyanins (Antho), scopolamine (SCO), and scopolamine plus anthocyanins (SCO+Antho). After seven days of treatment, they were tested in the inhibitory avoidance task and open field test and submitted to euthanasia. The CC and the HC were collected for biochemical assays. The effect of treatment with Antho (200 mgkg(-1), i.p.) was investigated in rats trained to a stable level of performance and post-treated with SCO (1 mgkg(-1), i.p. 30 min after training). KEY FINDINGS: The treatment with SCO decreased the step-down latency in inhibitory avoidance task. Antho prevented the scopolamine-induced memory impairment and also the increase of NTPDase activity in the CC and HC. Furthermore, the treatment with anthocyanins prevents the decrease in 5'-nucleotidase activity and the increase in adenosine deaminase activity induced by SCO in HC. In addition, the treatment with Antho prevented the decrease in ATP levels induced by SCO in the CC and HC. SIGNIFICANCE: Our results show that scopolamine may affect purinergic enzymatic cascade or cause alterations in energy metabolism inducing loss of memory. In contrast Antho could reverse these changes, suggesting a neuroprotective effect of Antho on ectonucleotidase activities and neuronal energetic metabolism.

Gene expression modulation by heat stress in Acidithiobacillus ferrooxidans LR.

During bioleaching, Acidithiobacillus ferrooxidans is subjected to different types of stress, including heat stress, which affect bacterial growth. In this work, real time quantitative PCR was used to analyze the expression of heat shock genes, as well as genes that encode proteins related to several functional categories in A. ferrooxidans. Cells were submitted to long-term growth and heat shock, both at 40°C. The results showed that heat shock affected the expression levels of most genes investigated, whilst long-term growth at 40°C resulted in minor changes in gene expression, except for certain genes related to iron transport, which were strongly down-regulated, suggesting that the iron processing capability of A. ferrooxidans was affected by long-term growth at 40°C. A bioinformatic analysis of the genes' promoter regions indicated a putative transcriptional regulation by the σ(32) factor in 12 of the 31 genes investigated, suggesting the involvement of other regulatory mechanisms in the response of A. ferrooxidans to heat stress.

The small heat shock proteins from Acidithiobacillus ferrooxidans: gene expression, phylogenetic analysis, and structural modeling.

BACKGROUND: Acidithiobacillus ferrooxidans is an acidophilic, chemolithoautotrophic bacterium that has been successfully used in metal bioleaching. In this study, an analysis of the A. ferrooxidans ATCC 23270 genome revealed the presence of three sHSP genes, Afe_1009, Afe_1437 and Afe_2172, that encode proteins from the HSP20 family, a class of intracellular multimers that is especially important in extremophile microorganisms. RESULTS: The expression of the sHSP genes was investigated in A. ferrooxidans cells submitted to a heat shock at 40°C for 15, 30 and 60 minutes. After 60 minutes, the gene on locus Afe_1437 was about 20-fold more highly expressed than the gene on locus Afe_2172. Bioinformatic and phylogenetic analyses showed that the sHSPs from A. ferrooxidans are possible non-paralogous proteins, and are regulated by the σ32 factor, a common transcription factor of heat shock proteins. Structural studies using homology molecular modeling indicated that the proteins encoded by Afe_1009 and Afe_1437 have a conserved α-crystallin domain and share similar structural features with the sHSP from Methanococcus jannaschii, suggesting that their biological assembly involves 24 molecules and resembles a hollow spherical shell. CONCLUSION: We conclude that the sHSPs encoded by the Afe_1437 and Afe_1009 genes are more likely to act as molecular chaperones in the A. ferrooxidans heat shock response. In addition, the three sHSPs from A. ferrooxidans are not recent paralogs, and the Afe_1437 and Afe_1009 genes could be inherited horizontally by A. ferrooxidans.

Functional characterization and target discovery of glycoside hydrolases from the digestome of the lower termite Coptotermes gestroi.

BACKGROUND: Lignocellulosic materials have been moved towards the forefront of the biofuel industry as a sustainable resource. However, saccharification and the production of bioproducts derived from plant cell wall biomass are complex and lengthy processes. The understanding of termite gut biology and feeding strategies may improve the current state of biomass conversion technology and bioproduct production. RESULTS: The study herein shows comprehensive functional characterization of crude body extracts from Coptotermes gestroi along with global proteomic analysis of the termite's digestome, targeting the identification of glycoside hydrolases and accessory proteins responsible for plant biomass conversion. The crude protein extract from C. gestroi was enzymatically efficient over a broad pH range on a series of natural polysaccharides, formed by glucose-, xylose-, mannan- and/or arabinose-containing polymers, linked by various types of glycosidic bonds, as well as ramification types. Our proteomic approach successfully identified a large number of relevant polypeptides in the C. gestroi digestome. A total of 55 different proteins were identified and classified into 29 CAZy families. Based on the total number of peptides identified, the majority of components found in the C. gestroi digestome were cellulose-degrading enzymes. Xylanolytic enzymes, mannan- hydrolytic enzymes, pectinases and starch-degrading and debranching enzymes were also identified. Our strategy enabled validation of liquid chromatography with tandem mass spectrometry recognized proteins, by enzymatic functional assays and by following the degradation products of specific 8-amino-1,3,6-pyrenetrisulfonic acid labeled oligosaccharides through capillary zone electrophoresis. CONCLUSIONS: Here we describe the first global study on the enzymatic repertoire involved in plant polysaccharide degradation by the lower termite C. gestroi. The biochemical characterization of whole body termite extracts evidenced their ability to cleave all types of glycosidic bonds present in plant polysaccharides. The comprehensive proteomic analysis, revealed a complete collection of hydrolytic enzymes including cellulases (GH1, GH3, GH5, GH7, GH9 and CBM 6), hemicellulases (GH2, GH10, GH11, GH16, GH43 and CBM 27) and pectinases (GH28 and GH29).

Heat and phosphate starvation effects on the proteome, morphology and chemical composition of the biomining bacteria Acidithiobacillus ferrooxidans.

Acidithiobacillus ferrooxidans is a Gram negative, acidophilic, chemolithoautotrophic bacterium that plays an important role in metal bioleaching. During bioleaching, the cells are subjected to changes in the growth temperature and nutrients starvation. The aim of this study was to gather information about the response of the A.ferrooxidans Brazilian strain LR to K2HPO4 starvation and heat stress through investigation of cellular morphology, chemical composition and differential proteome. The scanning electron microscopic results showed that under the tested stress conditions, A. ferrooxidans cells became elongated while the Fourier transform infrared spectroscopy (FT-IR) analysis showed alterations in the wavenumbers between 850 and 1,275 cm(-1), which are related to carbohydrates, phospholipids and phosphoproteins. These findings indicate that the bacterial cell surface is affected by the tested stress conditions. A proteomic analysis, using 2-DE and tandem mass spectrometry, enabled the identification of 44 differentially expressed protein spots, being 30 due to heat stress (40°C) and 14 due to K2HPO4 starvation. The identified proteins belonged to 11 different functional categories, including protein fate, energy metabolism and cellular processes. The upregulated proteins were mainly from protein fate and energy metabolism categories. The obtained results provide evidences that A. ferrooxidans LR responds to heat stress and K2HPO4 starvation by inducing alterations in cellular morphology and chemical composition of the cell surface. Also, the identification of several proteins involved in protein fate suggests that the bacteria cellular homesostasis was affected. In addition, the identification of proteins from different functional categories indicates that the A. ferrooxidans response to higher than optimal temperatures and phosphate starvation involves global changes in its physiology.

Substrate cleavage pattern, biophysical characterization and low-resolution structure of a novel hyperthermostable arabinanase from Thermotoga petrophila.

Arabinan is a plant structural polysaccharide degraded by two enzymes; alpha-l-arabinofuranosidase and endo-1,5-alpha-l-arabinanase. These enzymes are highly diversified in nature, however, little is known about their biochemical and biophysical properties. We have characterized a novel arabinanase (AbnA) isolated from Thermotoga petrophila with unique thermostable properties such as the insignificant decrease of residual activity after incubation up to 90 degrees C. We determined the AbnA mode of operation through capillary zone electrophoresis, which accumulates arabinotriose and arabinobiose as end products after hydrolysis of arabinan-containing polysaccharides. Spectroscopic analyses by Far-UV circular dichroism and intrinsic tryptophan fluorescence emission demonstrated that AbnA is folded and formed mainly by beta-sheet structural elements. In silico molecular modeling showed that the AbnA structure encompasses a five-bladed beta-propeller catalytic core juxtaposed by distorted up-and-down beta-barrel domain. The low-resolution structure determined by small angle X-ray scattering indicated that AbnA is monomeric in solution and its molecular shape is in full agreement with the model.