Synthetic Peptides Derived From Lycosa Erythrognatha Venom: Interaction With Phospholipid Membranes and Activity Against Resistant Bacteria.

Superbugs are a public health problem, increasing the need of new drugs and strategies to combat them. Our group has previously identified LyeTxI, an antimicrobial peptide isolated from Lycosa erythrognatha spider venom. From LyeTxI, we synthesized and characterized a derived peptide named LyeTxI-b, which has shown significant in vitro and in vivo activity. In this work, we elucidate the interaction of LyeTxI-b with artificial membranes as well as its effects on resistant strains of bacteria in planktonic conditions or biofilms. Isothermal titration calorimetry revealed that LyeTxI-b interacts more rapidly and with higher intensity with artificial vesicles, showing higher affinity to anionic vesicles, when compared to synthetic LyeTxI. In calcein experiments, LyeTxI-b caused greater levels of vesicle cleavage. Both peptides showed antibacterial activity at concentrations of µmol L-1 against 12 different clinically isolated strains, in planktonic conditions, in a concentration-dependent manner. Furthermore, both peptides elicited a dose-dependent production of reactive oxygen species in methicillin-resistant Staphylococcus aureus. In S. aureus biofilm assay, LyeTxI-b was more potent than LyeTxI. However, none of these peptides reduced Escherichia coli biofilms. Our results show LyeTxI-b as a promising drug against clinically resistant strains, being a template for developing new antibiotics.

Effect of slight structural changes on the gelation properties of N-phenylstearamide supramolecular gels.

Supramolecular gels present several applications in which the gelator properties are closely dependent on their structure and solvent. Despite this, there are few studies on the effect of the gelation ability of gelators with slight molecular changes. Therefore, N-arylestearamides (in which aryl = phenyl (1), 4-tolyl (2) and 4-acetylphenyl (3)) were evaluated in different solvents. The critical gelefication concentration (CGC) values indicated that the substituents can significantly affect the concentration at which the supramolecular gels are formed, mainly in non-aromatic solvents (e.g. cyclohexane, acetonitrile and DMSO). From UV-Vis and DSC data, we verified that the gel-sol and sol-gel transitions (Tgel-sol and Tsol-gel) increase in the order of 1 < 2 < 3. Organogel strength was evaluated for 1-3 as a function of concentration and solvent type using rheology data. Gel strength is concentration-dependent and a strength order was found in acetonitrile, cyclohexane and DMSO, in which: 1 ∼ 2 > 3. Dynamic viscoelastic measurements as a function of temperature sweeps indicate a predominantly enthalpic contribution to the elasticity of the organogels formed from 1-3. Temperature-dependent 1H NMR indicates that NHO interactions may be responsible for the molecular association of molecules into 1D fibers, while 3D fibers were formed from van der Waals interactions.

LyeTxI-b, a Synthetic Peptide Derived From Lycosa erythrognatha Spider Venom, Shows Potent Antibiotic Activity in Vitro and in Vivo.

The antimicrobial peptide LyeTxI isolated from the venom of the spider Lycosa erythrognatha is a potential model to develop new antibiotics against bacteria and fungi. In this work, we studied a peptide derived from LyeTxI, named LyeTxI-b, and characterized its structural profile and its in vitro and in vivo antimicrobial activities. Compared to LyeTxI, LyeTxI-b has an acetylated N-terminal and a deletion of a His residue, as structural modifications. The secondary structure of LyeTxI-b is a well-defined helical segment, from the second amino acid to the amidated C-terminal, with no clear partition between hydrophobic and hydrophilic faces. Moreover, LyeTxI-b shows a potent antimicrobial activity against Gram-positive and Gram-negative planktonic bacteria, being 10-fold more active than the native peptide against Escherichia coli. LyeTxI-b was also active in an in vivo model of septic arthritis, reducing the number of bacteria load, the migration of immune cells, the level of IL-1ß cytokine and CXCL1 chemokine, as well as preventing cartilage damage. Our results show that LyeTxI-b is a potential therapeutic model for the development of new antibiotics against Gram-positive and Gram-negative bacteria.

NMR structures in different membrane environments of three ocellatin peptides isolated from Leptodactylus labyrinthicus.

The peptides ocellatin-LB1, -LB2 and -F1 have previously been isolated from anurans of the Leptodactylus genus and the sequences are identical from residue 1-22, which correspond to ocellatin-LB1 sequence (GVVDILKGAAKDIAGHLASKVM-NH2), whereas ocellatin-LB2 carries an extra N and ocellatin-F1 extra NKL residues at their C-termini. These peptides showed different spectra of activities and biophysical investigations indicated a direct correlation between membrane-disruptive properties and antimicrobial activities, i.e. ocellatin-F1 > ocellatin-LB1 > ocellatin-LB2. To better characterize their membrane interactions, we report here the detailed three-dimensional NMR structures of these peptides in TFE-d2:H2O (60:40) and in the presence of zwitterionic DPC-d38 and anionic SDS-d25 micellar solutions. Although the three peptides showed significant helical contents in the three mimetic environments, structural differences were noticed. When the structures of the three peptides in the presence of DPC-d38 micelles are compared to each other, a more pronounced curvature is observed for ocellatin-F1 and the bent helix, with the concave face composed mostly of hydrophobic residues, is consistent with the micellar curvature and the amphipathic nature of the molecule. Interestingly, an almost linear helical segment was observed for ocellatin-F1 in the presence of SDS-d25 micelles and the conformational differences in the two micellar environments are possibly related to the presence of the extra Lys residue near the peptide C-terminus, which increases the affinity of ocellatin-F1 to anionic membranes in comparison with ocellatin-LB1 and -LB2, as proved by isothermal titration calorimetry. To our knowledge, this work reports for the first time the three-dimensional structures of ocellatin peptides.

Synthesis, Crystal Structure, and Supramolecular Understanding of 1,3,5-Tris(1-phenyl-1H-pyrazol-5-yl)benzenes.

Understanding the supramolecular environment of crystal structures is necessary to facilitate designing molecules with desirable properties. A series of 12 novel 1,3,5-tris(1-phenyl-1H-pyrazol-5-yl)benzenes was used to assess the existence of planar stacking columns in supramolecular structures of pyrazoles. This class of molecules with different substituents may assist in understanding how small structural changes affect the supramolecular environment. The obtained compounds did not present the formation of planar stacking interactions between benzenes in solid or liquid states. This supposition was indicated by single crystal diffraction, Density Functional Theory (DFT) and quantum theory of atoms in molecules (QTAIM) calculations, and concentration-dependent liquid-state ¹H nuclear magnetic resonance (NMR). NMR showed that chemical shifts of benzene and pyrazole hydrogens confirm that planar stacking interactions are not formed in solution. The crystalline structures presented different molecular conformations. The molecular structures of 5 and 9b are in a twisted conformation, while compound 7 showed a conformation analogous to a calyx form.

First Assessment of Carbon Stock in the Belowground Biomass of Brazilian Mangroves

ABSTRACT Studies on belowground roots biomass have increasingly reported the importance of the contribution of this compartment in carbon stock maintenance in mangrove forests. To date, there are no estimates of this contribution in Brazilian mangrove forests, although the country has the second largest area of mangroves worldwide. For this study, trenches dug in fringing forests in Guaratiba State Biological Reserve (Rio de Janeiro, Brazil) were used to evaluate the contribution of the different classes of roots and the vertical stratification of carbon stock. The total carbon stock average in belowground roots biomass in these forests was 104.41 ± 20.73 tC.ha−1. From that, an average of 84.13 ± 21.34 tC.ha−1 corresponded to the carbon stock only in fine roots, which have diameters smaller than 5 mm and are responsible for over 80% of the total belowground biomass. Most of the belowground carbon stock is concentrated in the first 40 cm below the surface (about 70%). The root:shoot ratio in this study is 1.14. These estimates demonstrate that the belowground roots biomass significantly contributes, more than 50%, to the carbon stock in mangrove forests. And the mangrove root biomass can be greater than that of other Brazilian ecosystems.

First Assessment of Carbon Stock in the Belowground Biomass of Brazilian Mangroves.

Studies on belowground roots biomass have increasingly reported the importance of the contribution of this compartment in carbon stock maintenance in mangrove forests. To date, there are no estimates of this contribution in Brazilian mangrove forests, although the country has the second largest area of mangroves worldwide. For this study, trenches dug in fringing forests in Guaratiba State Biological Reserve (Rio de Janeiro, Brazil) were used to evaluate the contribution of the different classes of roots and the vertical stratification of carbon stock. The total carbon stock average in belowground roots biomass in these forests was 104.41 ± 20.73 tC.ha-1. From that, an average of 84.13 ± 21.34 tC.ha-1 corresponded to the carbon stock only in fine roots, which have diameters smaller than 5 mm and are responsible for over 80% of the total belowground biomass. Most of the belowground carbon stock is concentrated in the first 40 cm below the surface (about 70%). The root:shoot ratio in this study is 1.14. These estimates demonstrate that the belowground roots biomass significantly contributes, more than 50%, to the carbon stock in mangrove forests. And the mangrove root biomass can be greater than that of other Brazilian ecosystems.

Upregulation of p38 pathway accelerates proliferation and migration of MDA-MB-231 breast cancer cells.

Tumor cells capture the signaling pathways used by normal tissue to promote their own survival and dissemination and among them, the NF-κB and MAPK pathways (ERK, JNK and p38). MAPK activation has ambiguous effects on tumor cell fate depending on cell type, cancer stage and the engaged MAPK isoforms. A synthetic peptide named LyeTx II, derived from the venom of the Brazilian spider Lycosa erythrognatha, was capable of increasing MDA-MB-231 aggressive breast cancer cell proliferation as indicated by MTT and BrdU (5-bromo-2'-deoxyuridine) incorporation assay and cell migration. A correlation has been established between the accelerated proliferation and migration observed in the presence of LyeTx II and the upregulation of p38 MAPK phosphorylation. The use of the selective inhibitor of p38α/ß (SB203580) abrogated the peptide effect in MDA-MB-231 cells. Besides, an augment of the canonical NF-κB pathway activation considered as crucial in cancer progression was noted after cell incubation with LyeTx II. Importantly, activation of p38 and NF-κB pathways was dependent on TAK1 activity. Together, these data suggest that TAK1-p38 pathway may represent an interesting target for treatment of aggressive breast cancers.

Bacillus spp. Isolated from Puba as a Source of Biosurfactants and Antimicrobial Lipopeptides.

Several products of industrial interest are produced by Bacillus, including enzymes, antibiotics, amino acids, insecticides, biosurfactants and bacteriocins. This study aimed to investigate the potential of two bacterial isolates (P5 and C3) from puba, a regional fermentation product from cassava, to produce multiple substances with antimicrobial and surface active properties. Phylogenetic analyses showed close relation of isolates P5 and C3 with Bacillus amyloliquefaciens and Bacillus thuringiensis, respectively. Notably, Bacillus sp. P5 showed antimicrobial activity against pathogens such as Listeria monocytogenes and Bacillus cereus, in addition to antifungal activity. The presence of genes encoding pre-subtilosin (sboA), malonyl CoA transacylase (ituD), and the putative transcriptional terminator of surfactin (sfp) were detected in Bacillus sp. P5, suggesting the production of the bacteriocin subtilosin A and the lipopeptides iturin A and surfactin by this strain. For Bacillus sp. C3 the presence of sboA and spas (subtilin) genes was observed by the first time in members of B. cereus cluster. Bacillus sp. P5 showed emulsifying capability on mineral oil, soybean biodiesel and toluene, while Bacillus sp. C3 showed emulsifying capability only on mineral oil. The reduction of the surface tension in culture medium was also observed for strain P5, confirming the production of surface-active compounds by this bacterium. Monoprotonated molecular species and adducts of sodium and potassium ions of surfactin, iturin, and fengycin were detected in the P5 culture medium. Comparative MS/MS spectra of the peak m/z 1030 (C14 surfactin A or C15 surfactin B [M+Na]+) and peak m/z 1079 (C15 iturin [M+Na]+) showed the same fragmentation profile of standards, confirming the molecular identification. In conclusion, Bacillus sp. P5 showed the best potential for the production of antifungal, antibacterial, and biosurfactant substances.

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions.

BACKGROUND: The availability of antimicrobial peptides from several different natural sources has opened an avenue for the discovery of new biologically active molecules. To the best of our knowledge, only two peptides isolated from the frog Leptodactylus labyrinthicus, namely pentadactylin and ocellatin-F1, have shown antimicrobial activities. Therefore, in order to explore the antimicrobial potential of this species, we have investigated the biological activities and membrane interactions of three peptides isolated from the anuran skin secretion. METHODS: Three peptide primary structures were determined by automated Edman degradation. These sequences were prepared by solid-phase synthesis and submitted to activity assays against gram-positive and gram-negative bacteria and against two fungal strains. The hemolytic properties of the peptides were also investigated in assays with rabbit blood erythrocytes. The conformational preferences of the peptides and their membrane interactions have been investigated by circular dichroism spectroscopy and liposome dye release assays. RESULTS: The amino acid compositions of three ocellatins were determined and the sequences exhibit 100% homology for the first 22 residues (ocellatin-LB1 sequence). Ocellatin-LB2 carries an extra Asn residue and ocellatin-F1 extra Asn-Lys-Leu residues at C-terminus. Ocellatin-F1 presents a stronger antibiotic potential and a broader spectrum of activities compared to the other peptides. The membrane interactions and pore formation capacities of the peptides correlate directly with their antimicrobial activities, i.e., ocellatin-F1 > ocellatin-LB1 > ocellatin-LB2. All peptides acquire high helical contents in membrane environments. However, ocellatin-F1 shows in average stronger helical propensities. CONCLUSIONS: The obtained results indicate that the three extra amino acid residues at the ocellatin-F1 C-terminus play an important role in promoting stronger peptide-membrane interactions and antimicrobial properties. The extra Asn-23 residue present in ocellatin-LB2 sequence seems to decrease its antimicrobial potential and the strength of the peptide-membrane interactions.

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions

Background The availability of antimicrobial peptides from several different natural sources has opened an avenue for the discovery of new biologically active molecules. To the best of our knowledge, only two peptides isolated from the frog Leptodactylus labyrinthicus, namely pentadactylin and ocellatin-F1, have shown antimicrobial activities. Therefore, in order to explore the antimicrobial potential of this species, we have investigated the biological activities and membrane interactions of three peptides isolated from the anuran skin secretion. Methods Three peptide primary structures were determined by automated Edman degradation. These sequences were prepared by solid-phase synthesis and submitted to activity assays against gram-positive and gram-negative bacteria and against two fungal strains. The hemolytic properties of the peptides were also investigated in assays with rabbit blood erythrocytes. The conformational preferences of the peptides and their membrane interactions have been investigated by circular dichroism spectroscopy and liposome dye release assays. Results The amino acid compositions of three ocellatins were determined and the sequences exhibit 100% homology for the first 22 residues (ocellatin-LB1 sequence). Ocellatin-LB2 carries an extra Asn residue and ocellatin-F1 extra Asn-Lys-Leu residues at C-terminus. Ocellatin-F1 presents a stronger antibiotic potential and a broader spectrum of activities compared to the other peptides. The membrane interactions and pore formation capacities of the peptides correlate directly with their antimicrobial activities, i.e., ocellatin-F1 > ocellatin-LB1 > ocellatin-LB2. All peptides acquire high helical contents in membrane environments. However, ocellatin-F1 shows in average stronger helical propensities. Conclusions The obtained results indicate that the three extra amino acid residues at the ocellatin-F1 C-terminus play an important role in promoting stronger peptide-membrane interactions and antimicrobial properties. The extra Asn-23 residue present in ocellatin-LB2 sequence seems to decrease its antimicrobial potential and the strength of the peptide-membrane interactions.(AU)

Ocellatin peptides from the skin secretion of the South American frog Leptodactylus labyrinthicus (Leptodactylidae): characterization, antimicrobial activities and membrane interactions

Abstract Background The availability of antimicrobial peptides from several different natural sources has opened an avenue for the discovery of new biologically active molecules. To the best of our knowledge, only two peptides isolated from the frog Leptodactylus labyrinthicus, namely pentadactylin and ocellatin-F1, have shown antimicrobial activities. Therefore, in order to explore the antimicrobial potential of this species, we have investigated the biological activities and membrane interactions of three peptides isolated from the anuran skin secretion. Methods Three peptide primary structures were determined by automated Edman degradation. These sequences were prepared by solid-phase synthesis and submitted to activity assays against gram-positive and gram-negative bacteria and against two fungal strains. The hemolytic properties of the peptides were also investigated in assays with rabbit blood erythrocytes. The conformational preferences of the peptides and their membrane interactions have been investigated by circular dichroism spectroscopy and liposome dye release assays. Results The amino acid compositions of three ocellatins were determined and the sequences exhibit 100% homology for the first 22 residues (ocellatin-LB1 sequence). Ocellatin-LB2 carries an extra Asn residue and ocellatin-F1 extra Asn-Lys-Leu residues at C-terminus. Ocellatin-F1 presents a stronger antibiotic potential and a broader spectrum of activities compared to the other peptides. The membrane interactions and pore formation capacities of the peptides correlate directly with their antimicrobial activities, i.e., ocellatin-F1 > ocellatin-LB1 > ocellatin-LB2. All peptides acquire high helical contents in membrane environments. However, ocellatin-F1 shows in average stronger helical propensities. Conclusions The obtained results indicate that the three extra amino acid residues at the ocellatin-F1 C-terminus play an important role in promoting stronger peptide-membrane interactions and antimicrobial properties. The extra Asn-23 residue present in ocellatin-LB2 sequence seems to decrease its antimicrobial potential and the strength of the peptide-membrane interactions.

Identification of metal-binding to proteins in seed samples using RF-HPLC-UV, GFAAS and MALDI-TOF-MS.

An extraction procedure using Tris-HCl buffer solution was employed in order to extract water-soluble proteins from seed samples of oat, wheat and soybean. Initially, the total protein concentration was determined by the Bradford method in each solution, after the extraction procedure. The soybean sample showed a higher concentration of total protein compared to the others. The protein extracts obtained were separated by reverse-phase chromatography (RP-HPLC-UV). The protein fractions were collected and analyzed by graphite furnace atomic absorption spectrometry (GFAAS) and matrix-assisted laser desorption/ionization (MALDI-TOF-MS) for determination of Cu, Fe, Mn and Zn and identification of proteins, respectively. The combination of techniques such as RP-HPLC-UV, GFAAS and MALDI-TOF-MS allowed the identification of several proteins bound to metals present in the seed samples.

Exposure to an extremely low-frequency electromagnetic field only slightly modifies the proteome of Chromobacterium violaceumATCC 12472.

Several studies of the physiological responses of different organisms exposed to extremely low-frequency electromagnetic fields (ELF-EMF) have been described. In this work, we report the minimal effects of in situ exposure to ELF-EMF on the global protein expression of Chromobacterium violaceum using a gel-based proteomic approach. The protein expression profile was only slightly altered, with five differentially expressed proteins detected in the exposed cultures; two of these proteins (DNA-binding stress protein, Dps, and alcohol dehydrogenase) were identified by MS/MS. The enhanced expression of Dps possibly helped to prevent physical damage to DNA. Although small, the changes in protein expression observed here were probably beneficial in helping the bacteria to adapt to the stress generated by the electromagnetic field.

Omics profiles used to evaluate the gene expression of Exiguobacterium antarcticum B7 during cold adaptation.

BACKGROUND: Exiguobacterium antarcticum strain B7 is a Gram-positive psychrotrophic bacterial species isolated in Antarctica. Although this bacteria has been poorly studied, its genome has already been sequenced. Therefore, it is an appropriate model for the study of thermal adaptation. In the present study, we analyzed the transcriptomes and proteomes of E. antarcticum B7 grown at 0°C and 37°C by SOLiD RNA-Seq, Ion Torrent RNA-Seq and two-dimensional difference gel electrophoresis tandem mass spectrometry (2D-DIGE-MS/MS). RESULTS: We found expression of 2,058 transcripts in all replicates from both platforms and differential expression of 564 genes (absolute log2FC≥1, P-value<0.001) comparing the two temperatures by RNA-Seq. A total of 73 spots were differentially expressed between the two temperatures on 2D-DIGE, 25 of which were identified by MS/MS. Some proteins exhibited patterns of dispersion in the gel that are characteristic of post-translational modifications. CONCLUSIONS: Our findings suggest that the two sequencing platforms yielded similar results and that different omic approaches may be used to improve the understanding of gene expression. To adapt to low temperatures, E. antarcticum B7 expresses four of the six cold-shock proteins present in its genome. The cold-shock proteins were the most abundant in the bacterial proteome at 0°C. Some of the differentially expressed genes are required to preserve transcription and translation, while others encode proteins that contribute to the maintenance of the intracellular environment and appropriate protein folding. The results denote the complexity intrinsic to the adaptation of psychrotrophic organisms to cold environments and are based on two omic approaches. They also unveil the lifestyle of a bacterial species isolated in Antarctica.

Genotype by sex and genotype by age interactions with sedentary behavior: the Portuguese Healthy Family Study.

Sedentary behavior (SB) expression and its underlying causal factors have been progressively studied, as it is a major determinant of decreased health quality. In the present study we applied Genotype x Age (GxAge) and Genotype x Sex (GxSex) interaction methods to determine if the phenotypic expression of different SB traits is influenced by an interaction between genetic architecture and both age and sex. A total of 1345 subjects, comprising 249 fathers, 327 mothers, 334 sons and 325 daughters, from 339 families of The Portuguese Healthy Family Study were included in the analysis. SB traits were assessed by means of a 3-d physical activity recall, the Baecke and IPAQ questionnaires. GxAge and GxSex interactions were analyzed using SOLAR 4.0 software. Sedentary behaviour heritability estimates were not always statistically significant (p>0.05) and ranged from 3% to 27%. The GxSex and GxAge interaction models were significantly better than the single polygenic models for TV (min/day), EEsed (kcal/day), personal computer (PC) usage and physical activty (PA) tertiles. The GxAge model is also significantly better than the polygenic model for Sed (min/day). For EEsed, PA tertiles, PC and Sed, the GxAge interaction was significant because the genetic correlation between SB environments was significantly different from 1. Further, PC and Sed variance heterogeneity among distinct ages were observed. The GxSex interaction was significant for EEsed due to genetic variance heterogeneity between genders and for PC due to a genetic correlation less than 1 across both sexes. Our results suggest that SB expression may be influenced by the interactions between genotype with both sex and age. Further, different sedentary behaviors seem to have distinct genetic architectures and are differentially affected by age and sex.

Genotype by energy expenditure interaction with metabolic syndrome traits: the Portuguese healthy family study.

Moderate-to-high levels of physical activity are established as preventive factors in metabolic syndrome development. However, there is variability in the phenotypic expression of metabolic syndrome under distinct physical activity conditions. In the present study we applied a Genotype X Environment interaction method to examine the presence of GxEE interaction in the phenotypic expression of metabolic syndrome. A total of 958 subjects, from 294 families of The Portuguese Healthy Family study, were included in the analysis. Total daily energy expenditure was assessed using a 3 day physical activity diary. Six metabolic syndrome related traits, including waist circumference, systolic blood pressure, glucose, HDL cholesterol, total cholesterol and triglycerides, were measured and adjusted for age and sex. GxEE examination was performed on SOLAR 4.3.1. All metabolic syndrome indicators were significantly heritable. The GxEE interaction model fitted the data better than the polygenic model (p<0.001) for waist circumference, systolic blood pressure, glucose, total cholesterol and triglycerides. For waist circumference, glucose, total cholesterol and triglycerides, the significant GxEE interaction was due to rejection of the variance homogeneity hypothesis. For waist circumference and glucose, GxEE was also significant by the rejection of the genetic correlation hypothesis. The results showed that metabolic syndrome traits expression is significantly influenced by the interaction established between total daily energy expenditure and genotypes. Physical activity may be considered an environmental variable that promotes metabolic differences between individuals that are distinctively active.

Another record of significant regional variation in toxicity of Tityus serrulatus venom in Brazil: a step towards understanding the possible role of sodium channel modulators.

The scorpion Tityus serrulatus is responsible for the most severe accidents that have been registered in Brazil, mainly in the state of Minas Gerais (MG), being the lung edema (LE), the main cause of death in these accidents. Although an increased in the number of accidents caused to this species in Federal District (Distrito Federal - DF), it seems that this particular species is not responsible for severe scorpionism cases in this region. Given this observation, we tested the toxicity in mice and compared the ability of T. serrulatus venom from DF (Ts-DF) and Minas Gerais State (Ts-MG) to induce LE in rats. The LD50 of Ts-DF venom was 51.6 µg/mouse, almost twice (1.98) higher than that obtained for Ts-MG venom. The ability of venom (0.5 mg/kg) to induce LE in rats was determined by the wet weight differences between treated and untreated lungs, by pulmonary morphological analyses and by pulmonary vascular permeability (PVP) using the Evans blue protocol. Significant differences in the wet weight of lungs and changes in PVP were found in Ts-MG venom treated rats when compared to rats treated with Ts-DF venom or untreated rats (p < 0.001), but no differences occurred when comparing rats treated with Ts-DF venom and untreated rats (p < 0.05). These results were confirmed by evaluation of pulmonary morphology. Comparison of chromatographic profiles obtained from these venoms (Ts-DF and Ts-MG) using the fractal dimension (D) analysis and the molecular mass fingerprint of the chromatographic fractions showed a higher number of components between 35 and 40% acetonitrile in Ts-MG venom than in Ts-DF venom, indicating a higher diversity of sodium channel modulators in that venom.

Interaction of arsenite with a zinc finger CCHC peptide: evidence for formation of an As-Zn-peptide mixed complex.

The interaction of arsenite with a Cys(3)His (CCHC) zinc finger model (34-51) HIV-1 nucleocapsid protein p7 (NCp7) peptide in the absence and presence of Zn(II) was studied using fluorescence spectroscopy, CD (circular dichroism) and ESI-MS (Electrospray Ionization Mass Spectrometry). We found that arsenic forms different complexes with the free peptide and the zinc finger peptide. In the former case the peptide conformation differed greatly from that of the zinc finger, whereas in the second case a mixed As-Zn-peptide complex was formed with partial preservation of zinc finger conformation. An apparent stability constant was estimated for the mixed As-Zn-peptide complex (K=2083 M(-1) and 442 M(-1) at 25°C and pHs 6 and 7, respectively). Our study also shows that the interaction of arsenic with the CCHC motif is facilitated by glutathione (GSH), through formation of a GS-As-peptide conjugate.

New cassane diterpenes from Caesalpinia echinata.

An investigation of the ethanolic extract from stems of Caesalpinia echinata Lam (Leguminosae-Caesalpinioideae) led to the isolation of five new cassane diterpenes along with known lambertianic acid. Their structures were determined based on spectroscopic methods. A preliminary study on leishmanicidal activity demonstrated that compounds 1, 2 and 6 were found to inhibit the growth of amastigote-like forms of Leishmania amazonensis without affecting mononuclear cells obtained from human peripheral blood.