Follistatin complexes Myostatin and antagonises Myostatin-mediated inhibition of myogenesis

Follistatin is known to antagonise the function of several members of the TGF-beta family of secreted signalling factors, including Myostatin, the most powerful inhibitor of muscle growth characterised to date. In this study, we compare the expression of Myostatin and Follistatin during chick development and show that they are expressed in the vicinity or in overlapping domains to suggest possible interaction during muscle development. We performed yeast and mammalian two-hybrid studies and show that Myostatin and Follistatin interact directly. We further show that single modules of the Follistatin protein cannot associate with Myostatin suggesting that the entire protein is required for the interaction. We analysed the interaction kinetics of the two proteins and found that Follistatin binds Myostatin with a high affinity of 5.84 x 10(-10) M. We next tested whether Follistatin suppresses Myostatin activity during muscle development. We confirmed our previous observation that treatment of chick limb buds with Myostatin results in a severe decrease in the expression of two key myogenic regulatory genes Pax-3 and MyoD. However, in the presence of Follistatin, the Myostatin-mediated inhibition of Pax-3 and MyoD expression is blocked. We additionally show that Myostatin inhibits terminal differentiation of muscle cells in high-density cell cultures of limb mesenchyme (micromass) and that Follistatin rescues muscle differentiation in a concentration-dependent manner. In summary, our data suggest that Follistatin antagonises Myostatin by direct protein interaction, which prevents Myostatin from executing its inhibitory effect on muscle development. (C) 2004 Elsevier Inc. All rights reserved.

Effects of lactoperoxidase and hydrogen peroxide on rheological properties of yoghurt

The effects of activation of the lactoperoxidase (LPO) system by H2O2-NaSCN and hydrogen peroxide (H2O2) on the accessibility of sulphydryl groups (SH) in skimmed milk, and on the dynamic rheological properties of the resulting yoghurt were investigated. Four different concentrations of each reagent (20-80 mg H2O2-NaSCN/kg milk and 100-400 mg H2O2/kg milk) were compared. Clear negative correlations were noted between the accessibility of SH groups and both LPO activation rate and H2O2 concentration. Also the native PAGE pattern of the heat-treated samples showed that with increase in the H2O2-NaSCN and H2O2 concentrations, the level of interaction between beta-lactoglobulin (beta-Ig) and kappa-casein (K-CN) decreased. The complex modulus (G*) of skimmed milk yoghurts declined gradually with the decrease in the concentration of accessible SH groups accordingly. Tan delta values of yoghurt samples were found to be different from the control, but close to each other, indicating that protein interaction forces taking place in the formation of gel networks of treated yoghurts were different from the control.

Influence of SNPs in nutrient-sensitive candidate genes and gene–diet interactions on blood lipids: the DiOGenes study

Blood lipid response to a given dietary intervention could be determined by the effect of diet, gene variants or gene–diet interactions. The objective of the present study was to investigate whether variants in presumed nutrient-sensitive genes involved in lipid metabolism modified lipid profile after weight loss and in response to a given diet, among overweight European adults participating in the Diet Obesity and Genes study. By multiple linear regressions, 240 SNPs in twenty-four candidate genes were investigated for SNP main and SNP–diet interaction effects on total cholesterol, LDL-cholesterol, HDL-cholesterol and TAG after an 8-week low-energy diet (only main effect), and a 6-month ad libitum weight maintenance diet, with different contents of dietary protein or glycaemic index. After adjusting for multiple testing, a SNP–dietary protein interaction effect on TAG was identified for lipin 1 (LPIN1) rs4315495, with a decrease in TAG of − 0·26 mmol/l per A-allele/protein unit (95 % CI − 0·38, − 0·14, P= 0·000043). In conclusion, we investigated SNP–diet interactions for blood lipid profiles for 240 SNPs in twenty-four candidate genes, selected for their involvement in lipid metabolism pathways, and identified one significant interaction between LPIN1 rs4315495 and dietary protein for TAG concentration.

Analyses of single nucleotide polymorphisms in selected nutrient-sensitive genes in weight-regain prevention: the DIOGENES study

BACKGROUND: Differences in the interindividual response to dietary intervention could be modified by genetic variation in nutrient-sensitive genes. OBJECTIVE: This study examined single nucleotide polymorphisms (SNPs) in presumed nutrient-sensitive candidate genes for obesity and obesity-related diseases for main and dietary interaction effects on weight, waist circumference, and fat mass regain over 6 mo. DESIGN: In total, 742 participants who had lost ≥ 8% of their initial body weight were randomly assigned to follow 1 of 5 different ad libitum diets with different glycemic indexes and contents of dietary protein. The SNP main and SNP-diet interaction effects were analyzed by using linear regression models, corrected for multiple testing by using Bonferroni correction and evaluated by using quantile-quantile (Q-Q) plots. RESULTS: After correction for multiple testing, none of the SNPs were significantly associated with weight, waist circumference, or fat mass regain. Q-Q plots showed that ALOX5AP rs4769873 showed a higher observed than predicted P value for the association with less waist circumference regain over 6 mo (-3.1 cm/allele; 95% CI: -4.6, -1.6; P/Bonferroni-corrected P = 0.000039/0.076), independently of diet. Additional associations were identified by using Q-Q plots for SNPs in ALOX5AP, TNF, and KCNJ11 for main effects; in LPL and TUB for glycemic index interaction effects on waist circumference regain; in GHRL, CCK, MLXIPL, and LEPR on weight; in PPARC1A, PCK2, ALOX5AP, PYY, and ADRB3 on waist circumference; and in PPARD, FABP1, PLAUR, and LPIN1 on fat mass regain for dietary protein interaction. CONCLUSION: The observed effects of SNP-diet interactions on weight, waist, and fat mass regain suggest that genetic variation in nutrient-sensitive genes can modify the response to diet. This trial was registered at clinicaltrials.gov as NCT00390637.

Identification of novel cholesteatoma-related gene expression signatures using full-genome microarrays

This study demonstrates that the expression profile of cholesteatoma is similar to a metastatic tumour and chronically inflamed tissue. Based on the investigated profiles we present novel protein-protein interaction and signal transduction networks, which include cholesteatoma-regulated transcripts and may be of great value for drug targeting and therapy development.

Genetic diversity at the Dhn3 locus in Turkish Hordeum spontaneum populations with comparative structural analyses

We analysed Hordeum spontaneum accessions from 21 different locations to understand the genetic diversity of HsDhn3 alleles and effects of single base mutations on the intrinsically disordered structure of the resulting polypeptide (HsDHN3). HsDHN3 was found to be YSK2-type with a low-frequency 6-aa deletion in the beginning of Exon 1. There is relatively high diversity in the intron region of HsDhn3 compared to the two exon regions. We have found subtle differences in K segments led to changes in amino acids chemical properties. Predictions for protein interaction profiles suggest the presence of a protein-binding site in HsDHN3 that coincides with the K1 segment. Comparison of DHN3 to closely related cereals showed that all of them contain a nuclear localization signal sequence flanking to the K1 segment and a novel conserved region located between the S and K1 segments [E(D/T)DGMGGR]. We found that H. vulgare, H. spontaneum, and Triticum urartu DHN3s have a greater number of phosphorylation sites for protein kinase C than other cereal species, which may be related to stress adaptation. Our results show that the nature and extent of mutations in the conserved segments of K1 and K2 are likely to be key factors in protection of cells.

Combining structure and dynamics: non-denaturing high-pressure effect on lysozyme in solution

Small-angle X-ray scattering (SAXS) and elastic and quasi-elastic neutron scattering techniques were used to investigate the high-pressure-induced changes on interactions, the low-resolution structure and the dynamics of lysozyme in solution. SAXS data, analysed using a global-fit procedure based on a new approach for hydrated protein form factor description, indicate that lysozyme completely maintains its globular structure up to 1500 bar, but significant modi. cations in the protein-protein interaction potential occur at approximately 600-1000 bar. Moreover, the mass density of the protein hydration water shows a clear discontinuity within this pressure range. Neutron scattering experiments indicate that the global and the local lysozyme dynamics change at a similar threshold pressure. A clear evolution of the internal protein dynamics from diffusing to more localized motions has also been probed. Protein structure and dynamics results have then been discussed in the context of protein-water interface and hydration water dynamics. According to SAXS results, the new configuration of water in the first hydration layer induced by pressure is suggested to be at the origin of the observed local mobility changes.

Comparison of the interactomic networks of different species in terms of accessibility

Protein-protein interaction networks were investigated in terms of outward accessibility, which quantifies the effectiveness of each protein in accessing other proteins and is related to the internality of nodes. By comparing the accessibility between 144 ortholog proteins in yeast and the fruit fly, we found that the accessibility tends to be higher among proteins in the fly than in yeast. In addition, z-scores of the accessibility calculated for different species revealed that the protein networks of less evolved species tend to be more random than those of more evolved species. The accessibility was also used to identify the border of the yeast protein interaction network, which was found to be mainly composed of viable proteins.

Seeking for simplicity in complex networks

Complex networks can be understood as graphs whose connectivity properties deviate from those of regular or near-regular graphs, which are understood as being ""simple"". While a great deal of the attention so far dedicated to complex networks has been duly driven by the ""complex"" nature of these structures, in this work we address the identification of their simplicity. The basic idea is to seek for subgraphs whose nodes exhibit similar measurements. This approach paves the way for complementing the characterization of networks, including results suggesting that the protein-protein interaction networks, and to a lesser extent also the Internet, may be getting simpler over time. Copyright (C) EPLA, 2009

Disruption of Saccharomyces cerevisiae by Plantaricin 149 and investigation of its mechanism of action with biomembrane model systems

The action of a synthetic antimicrobial peptide analog of Plantaricin 149 (Pln149a) against Saccharomyces cerevisiae and its interaction with biomembrane model systems were investigated. Pln149a was shown to inhibit S. cerevisiae growth by more than 80% in YPD medium, causing morphological changes in the yeast wall and remaining active and resistant to the yeast proteases even after 24 h of incubation. Different membrane model systems and carbohydrates were employed to better describe the Pln149a interaction with cellular components using circular dichroism and fluorescence spectroscopies, adsorption kinetics and surface elasticity in Langmuir monolayers. These assays showed that Pln149a does not interact with either mono/polysaccharides or zwitterionic LUVs, but is strongly adsorbed to and incorporated into negatively charged surfaces, causing a conformational change in its secondary structure from random-coil to helix upon adsorption. From the concurrent analysis of Pln149a adsorption kinetics and dilatational surface elasticity data, we determined that 2.5 mu M is the critical concentration at which Pln149a will disrupt a negative DPPG monolayer. Furthermore, Pln149a exhibited a carpet-like mechanism of action, in which the peptide initially binds to the membrane, covering its surface and acquiring a helical structure that remains associated to the negatively charged phospholipids. After this electrostatic interaction, another peptide region causes a strain in the membrane, promoting its disruption. (C) 2009 Elsevier B.V. All rights reserved.

Structural stability and reversible unfolding of recombinant porcine S100A12

Porcine S100A12 is a member of the S100 proteins, family of small acidic calcium-binding proteins characterized by the presence of two EF-hand motifs. These proteins are involved in many cellular events such as the regulation of protein phosphorylation, enzymatic activity, protein-protein interaction, Ca(2+) homeostasis, inflammatory processes and intermediate filament polymerization. In addition, members of this family bind Zn(2+) or Ca(2+) with cooperative effect on binding. In this study, the gene sequence encoding porcine S100A12 was obtained by the synthetic gene approach using E. coli codon bias. Additionally, we report a thermodynamic study of the recombinant S100A12 using circular dichroism, fluorescence and isothermal titration calorimetry. The results of urea and temperature induced unfolding and refolding processes indicated a reversible two-state process. Also, the ANS fluorescence studies showed that in presence of divalent ions the protein exposes hydrophobic sites which could facilitate the interaction with other proteins and trigger the physiological responses. (c) 2008 Elsevier B.V. All rights reserved.

Conformational Properties of Angiotensin II and Its Active and Inactive TOAC-Labeled Analogs in the Presence of Micelles. Electron Paramagnetic Resonance, Fluorescence, and Circular Dichroism Studies

The interaction between angiotensin II (AII, DRVYIHPF) and its analogs carrying 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid (TOAC) and detergents-negatively charged sodium dodecyl sulfate (SDS) and zwitterionic N-hexadecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate (HPS)-was examined by means of EPR, CD, and fluorescence. EPR spectra of partially active TOAC(1)-AII and inactive TOAC(3)-AII in aqueous solution indicated fast tumbling, the freedom of motion being greater at the N-terminus. Line broadening occurred upon interaction with micelles. Below SDS critical micelle concentration, broader lines indicated complex formation with tighter molecular packing than in micelles. Small changes in hyperfine splittings evinced TOAC location at the micelle-water interface. The interaction with anionic micelles was more effective than with zwitterionic micelles. Peptide-micelle interaction caused fluorescence increase. The TOAC-promoted intramolecular fluorescence quenching was more, pronounced for TOAC(3)-AII because of the proximity between the nitroxide and Tyr(4). CD spectra showed that although both AII and TOAC(1)-AII presented flexible conformations in water, TOAC(3)-AII displayed conformational restriction because of the TOAC-imposed bend (Schreier et al., Biopolymers 2004, 74, 389). In HPS, conformational changes were observed for the labeled peptides at neutral and basic pH. In SDS, all peptides underwent pH-dependent conformational changes. Although the spectra suggested similar folds for All and TOAC(1)-AII, different conformations were acquired by TOAC(3)-AII. The membrane environment has been hypothesized to shift conformational equilibria so as to stabilize the receptor-bound conformation of ligands. The fact that TOAC(3)-AII is unable to acquire conformations similar to those of native AII and partially active TOAC(1)-AII is probably the explanation for its lack of biological activity. (C) 2009 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 92: 525-537, 2009.

Aplicação de nanopartículas de quitosana com potencial de adjuvante na produção de soro contra o venneno do escorpião Tityus serrulatus

In Brazil, several species of scorpions are known to cause accidents which can lead to death, which are mainly belonging to the genus Tityus. The scorpion Tityus serrulatus is the main responsible for more severe cases. Anti-scorpion serums are routinely produced by various institutions, despite their effectiveness, quality and action depends on how quickly treatment is started. Studies have been developed in the search for appropriate technologies to encapsulate and release recombinant or natives proteins capable of inducing antibody production. In this context, chitosan copolymer which can be obtained from the partial deacetylation of chitin or in some microorganisms and it is biocompatible and biodegradable has been widely used for this purpose. This study aimed to search for a system release from chitosan nanoparticles for peptide / protein of the venom of the scorpion T. serrulatus, able to provide a new model of immunization in animals, in order to obtain a potential novel polyclonal serum, anti-venom T. serrulatus. The chitosan nanoparticles were prepared by ionic gelation with polyanion tripolyphosphate (TPP). After standardizing the concentrations of TPP and chitosan was evaluated the efficiency of incorporation of bovine serum albumin (BSA) and scorpion venom, showed particle size compatible with the intended purpose. The particles showed adequate size around 200nm. The crosslinking was confirmed by absorption spectroscopy in the infrared. After verified the high encapsulation efficiency (EE) for acid bicinconínico method (BCA) protein assay and the particle size distribution, the success of the technique was proven and the potential for in vivo application of nanoparticles. The experimental animals were vaccinated and the antibodies measured by ELISA

Transcriptomics and systems biology analysis in identification of specific pathways involved in cacao resistance and susceptibility to witches' broom disease

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