Homocysteine alters the osteoprotegerin/RANKL system in the osteoblast to promote bone loss: pivotal role of the redox regulator forkhead O1

In this study we determined the molecular mechanisms of how homocysteine differentially affects receptor activator of nuclear factor-kappa B ligand (RANKL) and osteoprotegerin (OPG) synthesis in the bone. The results showed that oxidative stress induced by homocysteine deranges insulin-sensitive FOXO1 and MAP kinase signaling cascades to decrease OPG and increase RANKL synthesis in osteoblast cultures. We observed that downregulation of insulin/FOXO1 and p38 MAP kinase signaling mechanisms due to phosphorylation of protein phosphatase 2 A (PP2A) was the key event that inhibited OPG synthesis in homocysteine-treated osteoblast cultures. siRNA knockdown experiments confirmed that FOXO1 is integral to OPG and p38 synthesis. Conversely homocysteine increased RANKL synthesis in osteoblasts through c-Jun/JNK MAP kinase signaling mechanisms independent of FOXO1. In the rat bone milieu, high-methionine diet-induced hyperhomocysteinemia lowered FOXO1 and OPG expression and increased synthesis of proresorptive and inflammatory cytokines such as RANKL, M-CSF, IL-1 alpha, IL-1 beta, G-CSF, GM-CSF, MIP-1 alpha, IFN-gamma, IL-17, and TNF-alpha. Such pathophysiological conditions were exacerbated by ovariectomy. Lowering the serum homocysteine level by a simultaneous supplementation with N-acetylcysteine improved OPG and FOXO1 expression and partially antagonized RANKL and proresorptive cytokine synthesis in the bone milieu. These results emphasize that hyperhomocysteinemia alters the redox regulatory mechanism in the osteoblast by activating PP2A and deranging FOXO1 and MAPK signaling cascades, eventually shifting the OPG:RANKL ratio toward increased osteoclast activity and decreased bone quality (C) 2013 Elsevier Inc. All rights reserved.

Stabilizing effect of electrostatic vs. aromatic interactions in diproline nucleated peptide beta-hairpins

The contribution of Tyr-His vs. Cys-His interacting pairs to the scaffold stability of (D)Pro-(L)Pro nucleated peptide beta-hairpins has been examined. We present direct evidence for the superiority of the Cys-His pairs, mediated by sulphur-imidazole interactions, as added stabilizing agents of the beta-hairpin scaffold.

Structural Integrity of the Greek Key Motif in beta gamma-Crystallins Is Vital for Central Eye Lens Transparency

Background: We highlight an unrecognized physiological role for the Greek key motif, an evolutionarily conserved super-secondary structural topology of the beta gamma-crystallins. These proteins constitute the bulk of the human eye lens, packed at very high concentrations in a compact, globular, short-range order, generating transparency. Congenital cataract (affecting 400,000 newborns yearly worldwide), associated with 54 mutations in beta gamma-crystallins, occurs in two major phenotypes nuclear cataract, which blocks the central visual axis, hampering the development of the growing eye and demanding earliest intervention, and the milder peripheral progressive cataract where surgery can wait. In order to understand this phenotypic dichotomy at the molecular level, we have studied the structural and aggregation features of representative mutations. Methods: Wild type and several representative mutant proteins were cloned, expressed and purified and their secondary and tertiary structural details, as well as structural stability, were compared in solution, using spectroscopy. Their tendencies to aggregate in vitro and in cellulo were also compared. In addition, we analyzed their structural differences by molecular modeling in silico. Results: Based on their properties, mutants are seen to fall into two classes. Mutants A36P, L45PL54P, R140X, and G165fs display lowered solubility and structural stability, expose several buried residues to the surface, aggregate in vitro and in cellulo, and disturb/distort the Greek key motif. And they are associated with nuclear cataract. In contrast, mutants P24T and R77S, associated with peripheral cataract, behave quite similar to the wild type molecule, and do not affect the Greek key topology. Conclusion: When a mutation distorts even one of the four Greek key motifs, the protein readily self-aggregates and precipitates, consistent with the phenotype of nuclear cataract, while mutations not affecting the motif display `native state aggregation', leading to peripheral cataract, thus offering a protein structural rationale for the cataract phenotypic dichotomy ``distort motif, lose central vision''.

Generation of Ligand Specificity and Modes of Oligomerization in beta-Prism I Fold Lectins

beta-Prism I fold lectins constitute one of the five widely occurring structural classes of plant lectins. Each single domain subunit is made up of three Greek key motifs arranged in a threefold symmetric fashion. The threefold symmetry is not reflected in the sequence except in the case of the lectin from banana, a monocot, which carries two sugar-binding sites instead of the one in other lectins of known three-dimensional structure, all from dicots. This is believed to be a consequence of the different evolutionary paths followed by the lectin in monocots and dicots. The galactose-specific lectins among them have two chains produced by posttranslational proteolysis and contain three aromatic residues at the binding site. The extended binding sites of galactose- and mannose-specific lectins have been thoroughly characterized. Ligand binding at the sites involves both conformational selection and induced fit. Molecular plasticity of some of the lectins in the family has been characterized. The plasticity appears to be such as to promote variability in quaternary association which could be dimeric, tetrameric, or octameric. Structural and evolutionary reasons for the variability have been explored, and the relation of oligomerization to ligand binding and conformational selection investigated.

Structure constants of beta deformed super Yang-Mills

We study the structure constants of the N = 1 beta deformed theory perturbatively and at strong coupling. We show that the planar one loop corrections to the structure constants of single trace gauge invariant operators in the scalar sector is determined by the anomalous dimension Hamiltonian. This result implies that 3 point functions of the chiral primaries of the theory do not receive corrections at one loop. We then study the structure constants at strong coupling using the Lunin-Maldacena geometry. We explicitly construct the supergravity mode dual to the chiral primary with three equal U(1) R-charges in the Lunin-Maldacena geometry. We show that the 3 point function of this supergravity mode with semi-classical states representing two other similar chiral primary states but with large U(1) charges to be independent of the beta deformation and identical to that found in the AdS(5) x S-5 geometry. This together with the one-loop result indicate that these structure constants are protected by a non-renormalization theorem. We also show that three point function of U(1) R-currents with classical massive strings is proportional to the R-charge carried by the string solution. This is in accordance with the prediction of the R-symmetry Ward identity.

Thermoelectric properties of beta-FeSi2

We investigate the thermoelectric properties of beta-FeSi2 using first principles electronic structure and Boltzmann transport calculations. We report a high thermopower for both p- and n-type beta-FeSi2 over a wide range of carrier concentration and in addition find the performance for n-type to be higher than for the p-type. Our results indicate that, depending upon temperature, a doping level of 3 x 10(20) to 2 x 10(21) cm(-3) may optimize the thermoelectric performance. (C) 2013 AIP Publishing LLC.

On the absence of shear cracking and grain boundary cavitation in secondary tensile regions of Ti-6Al-4V-0.1B alloy during hot (alpha plus beta)-compression

Deformation instabilities, such as shear cracking and grain boundary cavitation, which are observed in the secondary tensile region of Ti-6Al-4V alloy during compressive deformation in the (+)-phase field, do not form in Ti-6Al-4V-0.1B alloy when processed under the same conditions. This has been attributed to the microstructural modifications, e.g. the absence of grain boundary and adjacent grain boundary retained layers and a lower proportion of 90(o)-misoriented -colonies that occur with boron addition.

Crystallographic texture and microstructure evolution during hot compression of Ti-6Al-4V-0.1B alloy in the (alpha plus beta)-regime

Microstructure and texture are known to undergo drastic modifications due to trace hypoeutectic boron addition (similar to 0.1wt.%) for various titanium alloys e.g. Ti-6Al-4V. The deformation behaviour of such an alloy Ti-6Al-4V-0.1B is investigated in the (+) phase field and compared against that of the base alloy Ti-6Al-4V studied under selfsame conditions. The deformation microstructures for the two alloys display bending and kinking of lamellae in near and softening via globularization of lamella in near phase regimes, respectively. The transition temperature at which pure slip based deformation changes to softening is lower for the boron added alloy. The presence of TiB particles is largely held attributable for the early softening of Ti-6Al-4V-0.1B alloy. The compression texture of both the alloys carry signature of pure phase defamation at lower temperature and phase transformation near the transus temperature. Texture is influenced by a complex interplay of the deformation and transformation processes in the intermediate temperature range. The contribution from phase transformation is prominent for Ti-6Al-4V-0.1B alloy at comparatively lower temperature.

Enhanced superplasticity for (alpha plus beta)-hot rolled Ti-6Al-4V-0.1B alloy by means of dynamic globularization

Thermo-mechanically processed Ti-6Al-4V alloy, with (0.1 wt.%) and without boron addition, has been subjected to tensile test under superplastic deformation conditions (Temperature, T = 850 degrees C and initial strain rate, (epsilon) over dot = 3 x 10(-4) s(-1)). The boron added alloy exhibited higher elongation (similar to 430%) in comparison to the base alloy without boron (similar to 365%). Superior ductility of the boron added alloy has been attributed to an enhanced alpha/beta interfacial boundary sliding. This was caused by riotous dynamic globularization leading to the abundant presence of equiaxed primary alpha grains with refined sizes and narrow distribution in the deforming microstructure. Cavities do occur around TiB particles during deformation; the cavities are, however, extremely localized and do not cause macroscopic cracking. (C) 2014 Elsevier Ltd. All rights reserved.

Specific Sequence of a Beta Turn in Human La Protein May Contribute to Species Specificity of Hepatitis C Virus

Human La protein is known to be an essential host factor for translation and replication of hepatitis C virus (HCV) RNA. Previously, we have demonstrated that residues responsible for interaction of human La protein with the HCV internal ribosomal entry site (IRES) around the initiator AUG within stem-loop IV form a beta-turn in the RNA recognition motif (RRM) structure. In this study, sequence alignment and mutagenesis suggest that the HCV RNA-interacting beta-turn is conserved only in humans and chimpanzees, the species primarily known to be infected by HCV. A 7-mer peptide corresponding to the HCV RNA-interacting region of human La inhibits HCV translation, whereas another peptide corresponding to the mouse La sequence was unable to do so. Furthermore, IRES-mediated translation was found to be significantly high in the presence of recombinant human La protein in vitro in rabbit reticulocyte lysate. We observed enhanced replication with HCV subgenomic and full-length replicons upon overexpression of either human La protein or a chimeric mouse La protein harboring a human La beta-turn sequence in mouse cells. Taken together, our results raise the possibility of creating an immunocompetent HCV mouse model using human-specific cell entry factors and a humanized form of La protein.

Self-assembled dipeptide nanotubes constituted by flexible beta-phenylalanine and conformationally constrained alpha,beta-dehydrophenylalanine residues as drug delivery system

Peptide based self assembled nanostructures have attracted growing interest in recent years due to their numerous potential applications particularly in biomedical sciences. Di-peptide Phe-Phe was shown previously to self-assemble into nanotube like structures. In this work, we studied the affect of peptide backbone length and conformational flexibility on the self assembly process by using two dipeptides based on the Phe-Phe backbone (beta Phe-Phe and beta Phe-Delta Phe): one containing a flexible beta Phe amino acid, and the other containing both a flexible bPhe as well as a backbone constraining Alpha Phe (alpha,beta-dehydrophenylalanine) amino acid. Electron microscopy and X-ray diffraction experiments revealed that these new di-peptides can self-assemble into nanotubes having different properties than the native Phe-Phe nanotubes. These nanotubes were stable over a broad range of temperatures and the introduction of non-natural amino acids provided them with stability against the action of nonspecific proteases. Moreover, these dipeptides showed no cytotoxicity towards HeLa and L929 cells, and were able to encapsulate small drug molecules. We further showed that anticancerous drug mitoxantrone was more efficient in killing HeLa and B6F10 cells when entrapped in nanotubes as compared to free mitoxantrone. Therefore, these beta-phenylalanine and alpha, beta-dehydrophenylalanine containing dipeptide nanotubes may be useful in the development of biocompatible and proteolytically stable drug delivery vehicles.

The Glycosylated Rv1860 Protein of Mycobacterium tuberculosis Inhibits Dendritic Cell Mediated TH1 and TH17 Polarization of T Cells and Abrogates Protective Immunity Conferred by BCG

We previously reported interferon gamma secretion by human CD4(+) and CD8(+) T cells in response to recombinant E. coli-expressed Rv1860 protein of Mycobacterium tuberculosis (MTB) as well as protection of guinea pigs against a challenge with virulent MTB following prime-boost immunization with DNA vaccine and poxvirus expressing Rv1860. In contrast, a Statens Serum Institute Mycobacterium bovis BCG (BCG-SSI) recombinant expressing MTB Rv1860 (BCG-TB1860) showed loss of protective ability compared to the parent BCG strain expressing the control GFP protein (BCG-GFP). Since Rv1860 is a secreted mannosylated protein of MTB and BCG, we investigated the effect of BCG-TB1860 on innate immunity. Relative to BCG-GFP, BCG-TB1860 effected a significant near total reduction both in secretion of cytokines IL-2, IL-12p40, IL-12p70, TNF-alpha, IL-6 and IL-10, and up regulation of co-stimulatory molecules MHC-II, CD40, CD54, CD80 and CD86 by infected bone marrow derived dendritic cells (BMDC), while leaving secreted levels of TGF-beta unchanged. These effects were mimicked by BCG-TB1860His which carried a 6-Histidine tag at the C-terminus of Rv1860, killed sonicated preparations of BCG-TB1860 and purified H37Rv-derived Rv1860 glycoprotein added to BCG-GFP, but not by E. coli-expressed recombinant Rv1860. Most importantly, BMDC exposed to BCG-TB1860 failed to polarize allogeneic as well as syngeneic T cells to secrete IFN-gamma and IL-17 relative to BCG-GFP. Splenocytes from mice infected with BCG-SSI showed significantly less proliferation and secretion of IL-2, IFN-gamma and IL-17, but secreted higher levels of IL-10 in response to in vitro restimulation with BCG-TB1860 compared to BCG-GFP. Spleens from mice infected with BCG-TB1860 also harboured significantly fewer DC expressing MHC-II, IL-12, IL-2 and TNF-alpha compared to mice infected with BCG-GFP. Glycoproteins of MTB, through their deleterious effects on DC may thus contribute to suppress the generation of a TH1- and TH17-dominated adaptive immune response that is vital for protection against tuberculosis.

Process induced electroactive beta-polymorph in PVDF: effect on dielectric and ferroelectric properties

The effects of various processing conditions, like annealing, poling, mechanical rolling and their combinations, on the dielectric and ferroelectric properties of PVDF poly(vinylidene fluoride)] were systematically studied in this work. Further, the effect of processing sequence on the structure and properties was investigated. While all the processing conditions adopted here resulted in phase transformation of the alpha- to electroactive beta-polymorph in PVDF, the fraction of beta-phase developed was observed to be strongly contingent on the adopted process. The transformation of alpha- to electroactive beta-polymorph was determined by X-ray diffraction and FTIR. The neat PVDF showed only beta-phase, whereas mechanically rolled samples exhibited the highest ca. 85% beta-phase in PVDF. Both the permittivity and the loss tangent decreased in the samples which had undergone different processing conditions. The polarization-electric field (P-E) loops for all the samples were evaluated. Interestingly, the energy density, estimated from the electrical displacement-electric field (D-E) loops, was observed to be highest for the poled samples which were initially rolled. The results indicate that various processing conditions can influence the dielectric and the ferroelectric properties differently.

Non-existence of tight neighborly triangulated manifolds with beta(1)=2

All triangulated d-manifolds satisfy the inequality ((f0-d-1)(2)) >= ((d+2)(2))beta(1) for d >= 3. A triangulated d-manifold is called tight neighborly if it attains equality in this bound. For each d >= 3, a (2d + 3)-vertex tight neighborly triangulation of the Sd-1-bundle over S-1 with beta(1) = 1 was constructed by Kuhnel in 1986. In this paper, it is shown that there does not exist a tight neighborly triangulated manifold with beta(1) = 2. In other words, there is no tight neighborly triangulation of (Sd-1 x S-1)(#2) or (Sd-1 (sic) S-1)(#2) for d >= 3. A short proof of the uniqueness of K hnel's complexes for d >= 4 under the assumption beta(1) not equal 0 is also presented.