Hemoglobin interactions with αB crystallin: a direct test of sensitivity to protein instability.

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Tyler J W Clark

Full Text Available As a small stress response protein, human αB crystallin, detects protein destabilization that can alter structure and function to cause self assembly of fibrils or aggregates in diseases of aging. The sensitivity of αB crystallin to protein instability was evaluated using wild-type hemoglobin (HbA and hemoglobin S (HbS, the glutamate-6-valine mutant that forms elongated, filamentous aggregates in sickling red blood cells. The progressive thermal unfolding and aggregation of HbA and HbS in solution at 37°C, 50°C and 55°C was measured as increased light scattering. UV circular dichroism (UVCD was used to evaluate conformational changes in HbA and HbS with time at the selected temperatures. The changes in interactions between αB crystallin and HbA or HbS with temperature were analyzed using differential centrifugation and SDS PAGE at 37°C, 50°C and 55°C. After only 5 minutes at the selected temperatures, differences in the aggregation or conformation of HbA and HbS were not observed, but αB crystallin bound approximately 6% and 25% more HbS than HbA at 37°C, and 50°C respectively. The results confirmed (a the remarkable sensitivity of αB crystallin to structural instabilities at the very earliest stages of thermal unfolding and (b an ability to distinguish the self assembling mutant form of HbS from the wild type HbA in solution.

Small heat shock proteins protect against α-synuclein-induced toxicity and aggregation

International Nuclear Information System (INIS)

Outeiro, Tiago Fleming; Klucken, Jochen; Strathearn, Katherine E.; Liu Fang; Nguyen, Paul; Rochet, Jean-Christophe; Hyman, Bradley T.; McLean, Pamela J.

2006-01-01

Protein misfolding and inclusion formation are common events in neurodegenerative diseases, such as Parkinson's disease (PD), Alzheimer's disease (AD) or Huntington's disease (HD). α-Synuclein (aSyn) is the main protein component of inclusions called Lewy bodies (LB) which are pathognomic of PD, Dementia with Lewy bodies (DLB), and other diseases collectively known as LB diseases. Heat shock proteins (HSPs) are one class of the cellular quality control system that mediate protein folding, remodeling, and even disaggregation. Here, we investigated the role of the small heat shock proteins Hsp27 and αB-crystallin, in LB diseases. We demonstrate, via quantitative PCR, that Hsp27 messenger RNA levels are ∼2-3-fold higher in DLB cases compared to control. We also show a corresponding increase in Hsp27 protein levels. Furthermore, we found that Hsp27 reduces aSyn-induced toxicity by ∼80% in a culture model while αB-crystallin reduces toxicity by ∼20%. In addition, intracellular inclusions were immunopositive for endogenous Hsp27, and overexpression of this protein reduced aSyn aggregation in a cell culture model

Amyloid-like protein inclusions in tobacco transgenic plants.

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Anna Villar-Piqué

Full Text Available The formation of insoluble protein deposits in human tissues is linked to the onset of more than 40 different disorders, ranging from dementia to diabetes. In these diseases, the proteins usually self-assemble into ordered β-sheet enriched aggregates known as amyloid fibrils. Here we study the structure of the inclusions formed by maize transglutaminase (TGZ in the chloroplasts of tobacco transplastomic plants and demonstrate that they have an amyloid-like nature. Together with the evidence of amyloid structures in bacteria and fungi our data argue that amyloid formation is likely a ubiquitous process occurring across the different kingdoms of life. The discovery of amyloid conformations inside inclusions of genetically modified plants might have implications regarding their use for human applications.

The Small Heal Shock Protein αA-Crystallin Is Expressed In Pancreas and Acts as Negative Regulator of Carcinogenesis

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Deng , Mi; Chen , Pei-Chao; Xie , Sisi; Zhao , Junqiong; Gong , Lili; Liu , Jinping; Zhang , Lan; Sun , Shuming; Liu , Jiao; Ma , Haili; Batra , Surinder; Li , David Wan-Cheng

2010-01-01

Abstract The small heat shock protein ?A-crystallin is a structural protein in the ocular lens. In addition, recent studies have also revealed that it is a molecular chaperone, an autokinase and a strong anti-apoptotic regulator. Besides its lenticular distribution, a previous study demonstrates that a detectable level of ?A-crystallin is found in other tissues including thymus and spleen. In the present study, we have re-examined the distribution of ?A-crystallin in various normal...

Structural and Functional Consequences of Chaperone Site Deletion in αA-Crystallin

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Santhoshkumar, Puttur; Karmakar, Srabani; Sharma, Krishna K.

2016-01-01

The chaperone-like activity of αA-crystallin has an important role in maintaining lens transparency. Previously we identified residues 70–88 as a chaperone site in αA-crystallin. In this study, we deleted the chaperone site residues to generate αAΔ70–76 and αAΔ70–88 mutants and investigated if there are additional substrate-binding sites in αA-crystallin. Both mutant proteins when expressed in E. coli formed inclusion bodies, and on solubilizing and refolding, they exhibited similar structural properties, with a 2- to 3-fold increase in molar mass compared to the molar mass of wild-type protein. The deletion mutants were less stable than the wild-type αA-crystallin. Functionally αAΔ70–88 was completely inactive as a chaperone, while αAΔ70–76 demonstrated a 40–50% reduction in anti-aggregation activity against alcohol dehydrogenase (ADH). Deletion of residues 70–88 abolished the ADH binding sites in αA-crystallin at physiological temperature. At 45 °C, cryptic ADH binding site(s) became exposed, which contributed subtly to the chaperone-like activity of αAΔ70–88. Both of the deletion mutants were completely inactive in suppressing aggregation of βL-crystallin at 53 °C. The mutants completely lost the anti-apoptotic property that αA-crystallin exhibits while they protected ARPE-19 (a human retinal pigment epithelial cell line) and primary human lens epithelial (HLE) cells from oxidative stress. Our studies demonstrate that residues 70–88 in αA-crystallin act as a primary substrate binding site and account for the bulk of the total chaperone activity. The β3 and β4 strands in αA-crystallin comprising 70–88 residues play an important role in maintenance of the structure and in preventing aggregation of denaturing proteins. PMID:27524665

Photoaggregation of crystallins (main proteins of eye lens) under the effect of XeCl laser radiation

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Soustov, Lev V.; Chelnokov, Evgeny V.; Bityurin, Nikita M.; Kiselev, A. L.; Nemov, V. V.; Sergeev, Yu. V.; Ostrovsky, Michail A.

2004-07-01

UV light is one of primary factors associated with cataract formation in the eye lens. α-, β-, γ-Crystallins maintain lens transparency, and damage to these proteins plays a major role in cataract formation. The effect of XeCl laser radiation (308 nm) on βL-crystallin solution is studied. The strong dependence of protein aggregation kinetics on both laser fluence (w) and repetition rate (F) is investigated. The kinetics features are similar to those of carbonic anhydrase photoaggregation studied previously.

Diffusion of nanosized sodium inclusions in platinum

International Nuclear Information System (INIS)

Poulsen, J.R.; Horsewell, A.; Eldrup, M.; Johnson, E.; Johansen, A.

1994-01-01

Na inclusions with diameters in the range from 2 nm to 15 nm have been made by ion implantation of Na into 70 nm thick single-crystalline Pt foils followed by annealing. The structure of solid inclusions and the diffusion of molten inclusions have been studied by transmission electron microscopy. At room temperature the inclusions are faceted and crystalline with a BCC structure and they are aligned topotactically with the Pt (FCC) matrix. The diffusion of inclusions in the liquid state was investigated by annealing at temperatures of 1227 K, 1432 K and 1534 K. The results are used to propose a method to produce sources for positron annihilation spectroscopy at high temperatures. (author)

Utilization of crystalline and protein-bound amino acids by growing-finishing pigs

DEFF Research Database (Denmark)

Nørgaard, Jan Værum; Buxant, L.; Clausen, D.

2016-01-01

It was hypothesized that diets containing crystalline AA (CAA) and protein-bound AA had a comparable nitrogen retention rate, even though the CAA-based diet is optimized as having a standardized ileal digestibility (SID) of 100% for the CAA. Two isoenergetic diets were formulated to provide ident...

Microfluidic chips with multi-junctions: an advanced tool in recovering proteins from inclusion bodies.

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Yamaguchi, Hiroshi; Miyazaki, Masaya

2015-01-01

Active recombinant proteins are used for studying the biological functions of genes and for the development of therapeutic drugs. Overexpression of recombinant proteins in bacteria often results in the formation of inclusion bodies, which are protein aggregates with non-native conformations. Protein refolding is an important process for obtaining active recombinant proteins from inclusion bodies. However, the conventional refolding method of dialysis or dilution is time-consuming and recovered active protein yields are often low, and a cumbersome trial-and-error process is required to achieve success. To circumvent these difficulties, we used controllable diffusion through laminar flow in microchannels to regulate the denaturant concentration. This method largely aims at reducing protein aggregation during the refolding procedure. This Commentary introduces the principles of the protein refolding method using microfluidic chips and the advantage of our results as a tool for rapid and efficient recovery of active recombinant proteins from inclusion bodies.

Membrane and inclusion body targeting of lyssavirus matrix proteins.

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Pollin, Reiko; Granzow, Harald; Köllner, Bernd; Conzelmann, Karl-Klaus; Finke, Stefan

2013-02-01

Lyssavirus matrix proteins (M) support virus budding and have accessory functions that may contribute to host cell manipulation and adaptation to specific hosts. Here, we show that rabies virus (RABV) and European Bat Lyssavirus Type 1 (EBLV-1) M proteins differ in targeting and accumulation at cellular membranes. In contrast to RABV M, EBLV-1 M expressed from authentic EBLV-1 or chimeric RABV accumulated at the Golgi apparatus. Chimeric M proteins revealed that Golgi association depends on the integrity of the entire EBLV-1 M protein. Since RABV and EBLV-1 M differ in the use of cellular membranes for particle formation, differential membrane targeting and transport of M might determine the site of virus production. Moreover, both RABV and EBLV-1 M were for the first time detected within the nucleus and in Negri body-like inclusions bodies. Whereas nuclear M may imply hitherto unknown functions of lyssavirus M in host cell manipulation, the presence of M in inclusion bodies may correlate with regulatory functions of M in virus RNA synthesis. The data strongly support a model in which targeting of lyssavirus M proteins to distinctintracellular sites is a key determinant of diverse features in lyssavirus replication, host adaptation and pathogenesis. © 2012 Blackwell Publishing Ltd.

Structure of a Highly Active Cephalopod S-crystallin Mutant: New Molecular Evidence for Evolution from an Active Enzyme into Lens-Refractive Protein.

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Tan, Wei-Hung; Cheng, Shu-Chun; Liu, Yu-Tung; Wu, Cheng-Guo; Lin, Min-Han; Chen, Chiao-Che; Lin, Chao-Hsiung; Chou, Chi-Yuan

2016-08-08

Crystallins are found widely in animal lenses and have important functions due to their refractive properties. In the coleoid cephalopods, a lens with a graded refractive index provides good vision and is required for survival. Cephalopod S-crystallin is thought to have evolved from glutathione S-transferase (GST) with various homologs differentially expressed in the lens. However, there is no direct structural information that helps to delineate the mechanisms by which S-crystallin could have evolved. Here we report the structural and biochemical characterization of novel S-crystallin-glutathione complex. The 2.35-Å crystal structure of a S-crystallin mutant from Octopus vulgaris reveals an active-site architecture that is different from that of GST. S-crystallin has a preference for glutathione binding, although almost lost its GST enzymatic activity. We've also identified four historical mutations that are able to produce a "GST-like" S-crystallin that has regained activity. This protein recapitulates the evolution of S-crystallin from GST. Protein stability studies suggest that S-crystallin is stabilized by glutathione binding to prevent its aggregation; this contrasts with GST-σ, which do not possess this protection. We suggest that a tradeoff between enzyme activity and the stability of the lens protein might have been one of the major driving force behind lens evolution.

Kinetics of inclusion body formation and its correlation with the characteristics of protein aggregates in Escherichia coli.

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Arun K Upadhyay

Full Text Available The objective of the research was to understand the structural determinants governing protein aggregation into inclusion bodies during expression of recombinant proteins in Escherichia coli. Recombinant human growth hormone (hGH and asparaginase were expressed as inclusion bodies in E.coli and the kinetics of aggregate formation was analyzed in details. Asparaginase inclusion bodies were of smaller size (200 nm and the size of the aggregates did not increase with induction time. In contrast, the seeding and growth behavior of hGH inclusion bodies were found to be sequential, kinetically stable and the aggregate size increased from 200 to 800 nm with induction time. Human growth hormone inclusion bodies showed higher resistance to denaturants and proteinase K degradation in comparison to those of asparaginase inclusion bodies. Asparaginase inclusion bodies were completely solubilized at 2-3 M urea concentration and could be refolded into active protein, whereas 7 M urea was required for complete solubilization of hGH inclusion bodies. Both hGH and asparaginase inclusion bodies showed binding with amyloid specific dyes. In spite of its low β-sheet content, binding with dyes was more prominent in case of hGH inclusion bodies than that of asparaginase. Arrangements of protein molecules present in the surface as well as in the core of inclusion bodies were similar. Hydrophobic interactions between partially folded amphiphillic and hydrophobic alpha-helices were found to be one of the main determinants of hGH inclusion body formation. Aggregation behavior of the protein molecules decides the nature and properties of inclusion bodies.

Kinetics of Inclusion Body Formation and Its Correlation with the Characteristics of Protein Aggregates in Escherichia coli

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Upadhyay, Arun K.; Murmu, Aruna; Singh, Anupam; Panda, Amulya K.

2012-01-01

The objective of the research was to understand the structural determinants governing protein aggregation into inclusion bodies during expression of recombinant proteins in Escherichia coli. Recombinant human growth hormone (hGH) and asparaginase were expressed as inclusion bodies in E.coli and the kinetics of aggregate formation was analyzed in details. Asparaginase inclusion bodies were of smaller size (200 nm) and the size of the aggregates did not increase with induction time. In contrast, the seeding and growth behavior of hGH inclusion bodies were found to be sequential, kinetically stable and the aggregate size increased from 200 to 800 nm with induction time. Human growth hormone inclusion bodies showed higher resistance to denaturants and proteinase K degradation in comparison to those of asparaginase inclusion bodies. Asparaginase inclusion bodies were completely solubilized at 2–3 M urea concentration and could be refolded into active protein, whereas 7 M urea was required for complete solubilization of hGH inclusion bodies. Both hGH and asparaginase inclusion bodies showed binding with amyloid specific dyes. In spite of its low β-sheet content, binding with dyes was more prominent in case of hGH inclusion bodies than that of asparaginase. Arrangements of protein molecules present in the surface as well as in the core of inclusion bodies were similar. Hydrophobic interactions between partially folded amphiphillic and hydrophobic alpha-helices were found to be one of the main determinants of hGH inclusion body formation. Aggregation behavior of the protein molecules decides the nature and properties of inclusion bodies. PMID:22479486

RNA aptamers targeted for human αA-crystallin do not bind αB-crystallin, and spare the α-crystallin domain.

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Mallik, Prabhat K; Shi, Hua; Pande, Jayanti

2017-09-16

The molecular chaperones, α-crystallins, belong to the small heat shock protein (sHSP) family and prevent the aggregation and insolubilization of client proteins. Studies in vivo have shown that the chaperone activity of the α-crystallins is raised or lowered in various disease states. Therefore, the development of tools to control chaperone activity may provide avenues for therapeutic intervention, as well as enable a molecular understanding of chaperone function. The major human lens α-crystallins, αA- (HAA) and αB- (HAB), share 57% sequence identity and show similar activity towards some clients, but differing activities towards others. Notably, both crystallins contain the "α-crystallin domain" (ACD, the primary client binding site), like all other members of the sHSP family. Here we show that RNA aptamers selected for HAA, in vitro, exhibit specific affinity to HAA but do not bind HAB. Significantly, these aptamers also exclude the ACD. This study thus demonstrates that RNA aptamers against sHSPs can be designed that show high affinity and specificity - yet exclude the primary client binding region - thereby facilitating the development of RNA aptamer-based therapeutic intervention strategies. Copyright © 2017 Elsevier Inc. All rights reserved.

Glycation precedes lens crystallin aggregation

International Nuclear Information System (INIS)

Swamy, M.S.; Perry, R.E.; Abraham, E.C.

1987-01-01

Non-enzymatic glycosylation (glycation) seems to have the potential to alter the structure of crystallins and make them susceptible to thiol oxidation leading to disulfide-linked high molecular weight (HMW) aggregate formation. They used streptozotocin diabetic rats during precataract and cataract stages and long-term cell-free glycation of bovine lens crystallins to study the relationship between glycation and lens crystallin aggregation. HMW aggregates and other protein components of the water-soluble (WS) and urea-soluble (US) fractions were separated by molecular sieve high performance liquid chromatography. Glycation was estimated by both [ 3 H]NaBH 4 reduction and phenylboronate agarose affinity chromatography. Levels of total glycated protein (GP) in the US fractions were about 2-fold higher than in the WS fractions and there was a linear increase in GP in both WS and US fractions. This increase was parallelled by a corresponding increase in HMW aggregates. Total GP extracted by the affinity method from the US fraction showed a predominance of HMW aggregates and vice versa. Cell-free glycation studies with bovine crystallins confirmed the results of the animals studies. Increasing glycation caused a corresponding increase in protein insolubilization and the insoluble fraction thus formed also contained more glycated protein. It appears that lens protein glycation, HMW aggregate formation, and protein insolubilization are interrelated

Characterization of hypothetical proteins Cpn0146, 0147, 0284 & 0285 that are predicted to be in the Chlamydia pneumoniae inclusion membrane

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Liu Kaiyang

2007-05-01

Full Text Available Abstract Background Although more than 100 Chlamydia pneumoniae hypothetical proteins have been predicted to be inclusion membrane proteins, only a few have been experimentally demonstrated to be in the inclusion membrane. Using antibodies raised with fusion proteins, we characterized four such hypothetical proteins encoded by two gene clusters (Cpn0146-147 and Cpn0284-285 in the C. pneumoniae genome. Results Cpn0146 and 0147 were detected in the inclusion membrane while Cpn0284 and 0285 inside inclusion and mainly associated with reticulate bodies although all four proteins contain an N-terminal bi-lobed hydrophobic region, a signature motif assigned to inclusion membrane proteins. These four hypothetical proteins were only detected in cells infected with C. pneumoniae but not other chlamydial species, with Cpn0147 at 6 hours and Cpn0146, 0284 & 0285 at 24 hours after infection. Cpn0146 & 147 but not Cpn0284 and 285 co-localized with a host cell endoplasmic reticulum marker, a property known to be possessed by some chlamydial inclusion membrane proteins, when expressed in the host cell cytosol via transgenes. However, the endoplasmic reticulum localization of the C. pneumoniae inclusion membrane proteins did not result in inhibition of the subsequent C. pneumoniae infection. Conclusion The hypothetical proteins Cpn0146 & 0147 were localized in the C. pneumoniae inclusion membrane while Cpn0284 & 0285 within the inclusion although all four were predicted to be Inc proteins, suggesting the need to experimentally characterize the predicted Inc proteins.

Viscosity of crystalline proteins in solution, when irradiated with 60 Co

International Nuclear Information System (INIS)

Bernardes, D.M.L.; Mastro, N.L. del

1992-01-01

In order to study 60 Co radiation effects on proteins, an aqueous solution of bovine crystalline was irradiated with doses from O to 25,000 Gy. Changes in viscosity were followed whether in the presence or absence of radiation response modifiers: glutathione (GSH), amino ethyl isothiourea (AET), mercapto ethyl alanine (MEA) e dimethyl sulfoxide (DMSO). Viscosity data at different temperature revealed that aggregate formation was the predominant process induced by radiation. The results showed also that in presence of those substances the radiation effects was diminished. (author)

Preparative Protein Production from Inclusion Bodies and Crystallization: A Seven-Week Biochemistry Sequence

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Peterson, Megan J.; Snyder, W. Kalani; Westerman, Shelley; McFarland, Benjamin J.

2011-01-01

We describe how to produce and purify proteins from E. coli inclusion bodies by adapting versatile, preparative-scale techniques to the undergraduate laboratory schedule. This seven-week sequence of experiments fits into an annual cycle of research activity in biochemistry courses. Recombinant proteins are expressed as inclusion bodies, which are collected, washed, then solubilized in urea. Stepwise dialysis to dilute urea over the course of a week produces refolded protein. Column chromatography is used to purify protein into fractions, which are then analyzed with gel electrophoresis and concentration assays. Students culminate the project by designing crystallization trials in sitting-drop trays. Student evaluation of the experience has been positive, listing 5–12 new techniques learned, which are transferrable to graduate research in academia and industry. PMID:21691428

Lipotoxicity induces hepatic protein inclusions through TBK1-mediated p62/SQSTM1 phosphorylation.

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Cho, Chun-Seok; Park, Hwan-Woo; Ho, Allison; Semple, Ian A; Kim, Boyoung; Jang, Insook; Park, Haeli; Reilly, Shannon; Saltiel, Alan R; Lee, Jun Hee

2017-12-18

Obesity commonly leads to hepatic steatosis, which often provokes lipotoxic injuries to hepatocytes that cause non-alcoholic steatohepatitis (NASH). NASH in turn is associated with the accumulation of insoluble protein aggregates that are composed of ubiquitinated proteins and ubiquitin adaptor p62/sequestosome 1 (SQSTM1). The formation of p62 inclusions in hepatocytes is the critical marker that distinguishes simple fatty liver from NASH and predicts a poor prognostic outcome for subsequent liver carcinogenesis. However, the molecular mechanism by which lipotoxicity induces protein aggregation is currently unknown. Here we show that upon saturated fatty acid-induced lipotoxicity, Tank-binding protein kinase 1 (TBK1) is activated and phosphorylates p62. The TBK1-mediated p62 phosphorylation is important for lipotoxicity-induced aggregation of ubiquitinated proteins and the formation of large protein inclusions in hepatocytes. In addition, cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING), upstream regulators of TBK1, are involved in the lipotoxic activation of TBK1 and subsequent p62 phosphorylation in hepatocytes. Furthermore, TBK1 inhibition prevented formation of the ubiquitin-p62 aggregates, not only in cultured hepatocytes, but also in mouse models of obesity and NASH. These results suggest that lipotoxic activation of TBK1 and subsequent p62 phosphorylation are critical steps in the NASH pathology of protein inclusion accumulation in hepatocytes. This mechanism can provide an explanation for how hypernutrition and obesity promote the development of severe liver pathologies, such as steatohepatitis and liver cancer, by facilitating the formation of p62 inclusions. This article is protected by copyright. All rights reserved. © 2017 by the American Association for the Study of Liver Diseases.

Radiocarbon Dating of the Human Eye Lens Crystallines Reveal Proteins without Carbon Turnover throughout Life

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Lynnerup, Niels; Kjeldsen, Henrik; Heegaard, Steffen; Jacobsen, Christina; Heinemeier, Jan

2008-01-01

Background Lens crystallines are special proteins in the eye lens. Because the epithelial basement membrane (lens capsule) completely encloses the lens, desquamation of aging cells is impossible, and due to the complete absence of blood vessels or transport of metabolites in this area, there is no subsequent remodelling of these fibers, nor removal of degraded lens fibers. Human tissue ultimately derives its 14C content from the atmospheric carbon dioxide. The 14C content of the lens proteins thus reflects the atmospheric content of 14C when the lens crystallines were formed. Precise radiocarbon dating is made possible by comparing the 14C content of the lens crystallines to the so-called bomb pulse, i.e. a plot of the atmospheric 14C content since the Second World War, when there was a significant increase due to nuclear-bomb testing. Since the change in concentration is significant even on a yearly basis this allows very accurate dating. Methodology/Principal Findings Our results allow us to conclude that the crystalline formation in the lens nucleus almost entirely takes place around the time of birth, with a very small, and decreasing, continuous formation throughout life. The close relationship may be further expressed as a mathematical model, which takes into account the timing of the crystalline formation. Conclusions/Significance Such a life-long permanence of human tissue has hitherto only been described for dental enamel. In confront to dental enamel it must be held in mind that the eye lens is a soft structure, subjected to almost continuous deformation, due to lens accommodation, yet its most important constituent, the lens crystalline, is never subject to turnover or remodelling once formed. The determination of the 14C content of various tissues may be used to assess turnover rates and degree of substitution (for example for brain cell DNA). Potential targets may be nervous tissues in terms of senile or pre-senile degradation, as well as other highly

Radiocarbon dating of the human eye lens crystallines reveal proteins without carbon turnover throughout life.

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Niels Lynnerup

Full Text Available BACKGROUND: Lens crystallines are special proteins in the eye lens. Because the epithelial basement membrane (lens capsule completely encloses the lens, desquamation of aging cells is impossible, and due to the complete absence of blood vessels or transport of metabolites in this area, there is no subsequent remodelling of these fibers, nor removal of degraded lens fibers. Human tissue ultimately derives its (14C content from the atmospheric carbon dioxide. The (14C content of the lens proteins thus reflects the atmospheric content of (14C when the lens crystallines were formed. Precise radiocarbon dating is made possible by comparing the (14C content of the lens crystallines to the so-called bomb pulse, i.e. a plot of the atmospheric (14C content since the Second World War, when there was a significant increase due to nuclear-bomb testing. Since the change in concentration is significant even on a yearly basis this allows very accurate dating. METHODOLOGY/PRINCIPAL FINDINGS: Our results allow us to conclude that the crystalline formation in the lens nucleus almost entirely takes place around the time of birth, with a very small, and decreasing, continuous formation throughout life. The close relationship may be further expressed as a mathematical model, which takes into account the timing of the crystalline formation. CONCLUSIONS/SIGNIFICANCE: Such a life-long permanence of human tissue has hitherto only been described for dental enamel. In confront to dental enamel it must be held in mind that the eye lens is a soft structure, subjected to almost continuous deformation, due to lens accommodation, yet its most important constituent, the lens crystalline, is never subject to turnover or remodelling once formed. The determination of the (14C content of various tissues may be used to assess turnover rates and degree of substitution (for example for brain cell DNA. Potential targets may be nervous tissues in terms of senile or pre

Phosphorylation of αB-crystallin: Role in stress, aging and patho-physiological conditions.

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Bakthisaran, Raman; Akula, Kranthi Kiran; Tangirala, Ramakrishna; Rao, Ch Mohan

2016-01-01

αB-crystallin, once thought to be a lenticular protein, is ubiquitous and has critical roles in several cellular processes that are modulated by phosphorylation. Serine residues 19, 45 and 59 of αB-crystallin undergo phosphorylation. Phosphorylation of S45 is mediated by p44/42 MAP kinase, whereas S59 phosphorylation is mediated by MAPKAP kinase-2. Pathway involved in S19 phosphorylation is not known. The review highlights the role of phosphorylation in (i) oligomeric structure, stability and chaperone activity, (ii) cellular processes such as apoptosis, myogenic differentiation, cell cycle regulation and angiogenesis, and (iii) aging, stress, cardiomyopathy-causing αB-crystallin mutants, and in other diseases. Depending on the context and extent of phosphorylation, αB-crystallin seems to confer beneficial or deleterious effects. Phosphorylation alters structure, stability, size distribution and dynamics of the oligomeric assembly, thus modulating chaperone activity and various cellular processes. Phosphorylated αB-crystallin has a tendency to partition to the cytoskeleton and hence to the insoluble fraction. Low levels of phosphorylation appear to be protective, while hyperphosphorylation has negative implications. Mutations in αB-crystallin, such as R120G, Q151X and 464delCT, associated with inherited myofibrillar myopathy lead to hyperphosphorylation and intracellular inclusions. An ongoing study in our laboratory with phosphorylation-mimicking mutants indicates that phosphorylation of R120GαB-crystallin increases its propensity to aggregate. Phosphorylation of αB-crystallin has dual role that manifests either beneficial or deleterious consequences depending on the extent of phosphorylation and interaction with cytoskeleton. Considering that disease-causing mutants of αB-crystallin are hyperphosphorylated, moderation of phosphorylation may be a useful strategy in disease management. This article is part of a Special Issue entitled Crystallin

Development of shear bands in amorphous-crystalline metallic alloys

International Nuclear Information System (INIS)

Pozdnyakov, V.A.

2004-01-01

A theoretical study is made into conditions of shear band evolution in amorphous-crystalline alloys with various morphological types of structural constituents. The condition of shear band evolution in thin amorphous alloys in the interior of the crystalline matrix is obtained. It is shown that a scale effect exists which manifests itself in suppression of the process of localized plastic flow with amorphous alloy thickness decreasing down to the limit. The analysis of the condition for shear band evolution in an amorphous alloy with nanocrystalline inclusions is accomplished. The relationship of a critical stress of shear band evolution to a volume fraction of disperse crystal inclusions is obtained. A consideration is also given to the evolution of shear bands in the material containing amorphous and crystalline areas of micro meter size. For the alloy with the structure of this type conditions for propagation of localized flows by a relay race type mechanism are determined [ru

Radiocarbon dating of the human eye lens crystallines reveal proteins without carbon turnover throughout life

DEFF Research Database (Denmark)

Lynnerup, Niels; Kjeldsen, Henrik; Heegaard, Steffen

2008-01-01

, there is no subsequent remodelling of these fibers, nor removal of degraded lens fibers. Human tissue ultimately derives its (14)C content from the atmospheric carbon dioxide. The (14)C content of the lens proteins thus reflects the atmospheric content of (14)C when the lens crystallines were formed. Precise radiocarbon...

Protein misfolding specifies recruitment to cytoplasmic inclusion bodies.

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Bersuker, Kirill; Brandeis, Michael; Kopito, Ron R

2016-04-25

Inclusion bodies (IBs) containing aggregated disease-associated proteins and polyubiquitin (poly-Ub) conjugates are universal histopathological features of neurodegenerative diseases. Ub has been proposed to target proteins to IBs for degradation via autophagy, but the mechanisms that govern recruitment of ubiquitylated proteins to IBs are not well understood. In this paper, we use conditionally destabilized reporters that undergo misfolding and ubiquitylation upon removal of a stabilizing ligand to examine the role of Ub conjugation in targeting proteins to IBs that are composed of an N-terminal fragment of mutant huntingtin, the causative protein of Huntington's disease. We show that reporters are excluded from IBs in the presence of the stabilizing ligand but are recruited to IBs after ligand washout. However, we find that Ub conjugation is not necessary to target reporters to IBs. We also report that forced Ub conjugation by the Ub fusion degradation pathway is not sufficient for recruitment to IBs. Finally, we find that reporters and Ub conjugates are stable at IBs. These data indicate that compromised folding states, rather than conjugation to Ub, can specify recruitment to IBs. © 2016 Bersuker et al.

Crystallization and evaluation of hen egg-white lysozyme crystals for protein pH titration in the crystalline state.

Science.gov (United States)

Iwai, Wakari; Yagi, Daichi; Ishikawa, Takuya; Ohnishi, Yuki; Tanaka, Ichiro; Niimura, Nobuo

2008-05-01

To observe the ionized status of the amino acid residues in proteins at different pH (protein pH titration in the crystalline state) by neutron diffraction, hen egg-white lysozyme was crystallized over a wide pH range (2.5-8.0). Crystallization phase diagrams at pH 2.5, 6.0 and 7.5 were determined. At pH diagram, and at pH > 4.5 the border shifted to the right (higher precipitant concentration). The qualities of these crystals were characterized using the Wilson plot method. The qualities of all crystals at different pH were more or less equivalent (B-factor values within 25-40). It is expected that neutron diffraction analysis of these crystals of different pH provides equivalent data in quality for discussions of protein pH titration in the crystalline state of hen egg-white lysozyme.

Role of the disaggregase ClpB in processing of proteins aggregated as inclusion bodies.

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Zblewska, Kamila; Krajewska, Joanna; Zolkiewski, Michal; Kędzierska-Mieszkowska, Sabina

2014-08-01

Overproduction of heterologous proteins in bacterial systems often results in the formation of insoluble inclusion bodies (IBs), which is a major impediment in biochemical research and biotechnology. In principle, the activity of molecular chaperones could be employed to gain control over the IB formation and to improve the recombinant protein yields, but the potential of each of the major bacterial chaperones (DnaK/J, GroEL/ES, and ClpB) to process IBs has not been fully established yet. We investigated the formation of inclusion bodies (IBs) of two aggregation-prone proteins, VP1LAC and VP1GFP, overproduced in Escherichiacoli in the presence and absence of the chaperone ClpB. We found that both ClpB isoforms, ClpB95 and ClpB80 accumulated in E. coli cells during the production of IBs. The amount of IB proteins increased in the absence of ClpB. ClpB supported the resolubilization and reactivation of the aggregated VP1LAC and VP1GFP in E. coli cells. The IB disaggregation was optimal in the presence of both ClpB95 and ClpB80. Our results indicate an essential role of ClpB in controlling protein aggregation and inclusion body formation in bacteria. Copyright © 2014 Elsevier Inc. All rights reserved.

Employment of colorimetric enzyme assay for monitoring expression and solubility of GST fusion proteins targeted to inclusion bodies.

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Mačinković, Igor S; Abughren, Mohamed; Mrkic, Ivan; Grozdanović, Milica M; Prodanović, Radivoje; Gavrović-Jankulović, Marija

2013-12-01

High levels of recombinant protein expression can lead to the formation of insoluble inclusion bodies. These complex aggregates are commonly solubilized in strong denaturants, such as 6-8M urea, although, if possible, solubilization under milder conditions could facilitate subsequent refolding and purification of bioactive proteins. Commercially available GST-tag assays are designed for quantitative measurement of GST activity under native conditions. GST fusion proteins accumulated in inclusion bodies are considered to be undetectable by such assays. In this work, solubilization of recombinantly produced proteins was performed in 4M urea. The activity of rGST was assayed in 2M urea and it was shown that rGST preserves 85% of its activity under such denaturing conditions. A colorimetric GST activity assay with 1-chloro-2, 4-dinitrobenzene (CDNB) was examined for use in rapid detection of expression targeted to inclusion bodies and for the identification of inclusion body proteins which can be solubilized in low concentrations of chaotropic agents. Applicability of the assay was evaluated by tracking protein expression of two GST-fused allergens of biopharmaceutical value in E. coli, GST-Der p 2 and GST-Mus a 5, both targeted to inclusion bodies. Copyright © 2013 Elsevier B.V. All rights reserved.

Lens proteome map and alpha-crystallin profile of the catfish Rita rita.

Science.gov (United States)

Mohanty, Bimal Prasanna; Bhattacharjee, Soma; Das, Manas Kumar

2011-02-01

Crystallins are a diverse group of proteins that constitute nearly 90% of the total soluble proteins of the vertebrate eye lens and these tightly packed crystallins are responsible for transparency of the lens. These proteins have been studied in different model and non-model species for understanding the modifications they undergo with ageing that lead to cataract, a disease of protein aggregation. In the present investigation, we studied the lens crystallin profile of the tropical freshwater catfish Rita rita. Profiles of lens crystallins were analyzed and crystallin proteome maps of Rita rita were generated for the first time. alphaA-crystallins, member of the alpha-crystallin family, which are molecular chaperons and play crucial role in maintaining lens transparency were identified by 1- and 2-D immunoblot analysis with anti-alphaA-crystallin antibody. Two protein bands of 19-20 kDa were identified as alphaA-crystallins on 1-D immunoblots and these bands separated into 10 discrete spots on 2-D immunoblot. However, anti-alphaB-crystallin and antiphospho-alphaB-crystallin antibodies were not able to detect any immunoreactive bands on 1- and 2-D immunoblots, indicating alphaB-crystallin was either absent or present in extremely low concentration in Rita rita lens. Thus, Rita rita alpha-crystallins are more like that of the catfish Clarias batrachus and the mammal kangaroo in its alphaA- and alphaB-crystallin content (contain low amount from 5-9% of alphaB-crystallin) and unlike the dogfish, zebrafish, human, bovine and mouse alpha-crystallins (contain higher amount of alphaB-crystallin from 25% in mouse and bovine to 85% in dogfish). Results of the present study can be the baseline information for stimulating further investigation on Rita rita lens crystallins for comparative lens proteomics. Comparing and contrasting the alpha-crystallins of the dogfish and Rita rita may provide valuable information on the functional attributes of alphaA- and alphaB-isoforms, as

Specific chlamydial inclusion membrane proteins associate with active Src family kinases in microdomains that interact with the host microtubule network.

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Mital, Jeffrey; Miller, Natalie J; Fischer, Elizabeth R; Hackstadt, Ted

2010-09-01

Chlamydiae are Gram-negative obligate intracellular bacteria that cause diseases with significant medical and economic impact. Chlamydia trachomatis replicates within a vacuole termed an inclusion, which is extensively modified by the insertion of a number of bacterial effector proteins known as inclusion membrane proteins (Incs). Once modified, the inclusion is trafficked in a dynein-dependent manner to the microtubule-organizing centre (MTOC), where it associates with host centrosomes. Here we describe a novel structure on the inclusion membrane comprised of both host and bacterial proteins. Members of the Src family of kinases are recruited to the chlamydial inclusion in an active form. These kinases display a distinct, localized punctate microdomain-like staining pattern on the inclusion membrane that colocalizes with four chlamydial inclusion membrane proteins (Incs) and is enriched in cholesterol. Biochemical studies show that at least two of these Incs stably interact with one another. Furthermore, host centrosomes associate with these microdomain proteins in C. trachomatis-infected cells and in uninfected cells exogenously expressing one of the chlamydial effectors. Together, the data suggest that a specific structure on the C. trachomatis inclusion membrane may be responsible for the known interactions of chlamydiae with the microtubule network and resultant effects on centrosome stability.

Refolding in high hydrostatic pressure of recombinant proteins from inclusion bodies in Escherichia Coli

International Nuclear Information System (INIS)

Balduino, Keli Nunes

2009-01-01

The expression of proteins as inclusion bodies in bacteria is a widely used alternative for production of recombinant protein. However, the aggregation is a problem often encountered during refolding of these proteins. High hydrostatic pressure are able to solubilise the inclusion bodies in the presence of low concentrations of denaturant reagents, encouraging refolding protein with high efficiency and reduce costs. This work aims to refolding of recombinant proteins expressed in Escherichia coli from inclusion bodies using high hydrostatic pressure. Three toxins, all featuring five or more disulfide bonds were studied: NXH8, Natterin 2 and Bothropstoxin 1. Suspensions of inclusion bodies of the three proteins were pressurized to 2000 bars for 16 hours. The buffers were optimized for refolding of the three proteins. The buffer used in the refolding of NXH8 was 50 mM Tris HCl, pH 9.0 with proportion of 1GSH: 4GSSG at a concentration of 6 mM and 2 M GdnHCl. Inclusion bodies were used in O.D. (A600nm) of 0.5. After refolding process, dialysis was performed at pH 7.0. The final yield of obtaining soluble NXH8 was 40% (28,6 mg of soluble NXH8/L of culture medium). The refolding of Bothropstoxin 1 was obtained in refolding buffer of Tris HCl 50 mM, pH 7,5 with proportion of 2 GSH: GSSG 3 and concentration of 3 mM and 1 M GdnHCl. Use with a suspension of O.D. (A600nm) of 0.5. The final yield of recovery of Bothropstoxin 1 refolded was 32% (9,2 mg of refolded Bothropstoxin 1/L of culture medium). The refolding of Natterin 2 was performed in the refolding buffer: 20 mM Tris HCl pH 9.0 at a ratio of 2 GSH: 3GSSG and concentration of 10 mM and 1 M GdnHCl and inclusion bodies O.D. (A600nm) of 6.0. The yield of Natterin 2 refolded was 20% (3,7 mg/L of culture medium). Physico-chemical and biological analysis were performed by SDS-PAGE, western blot, scanning electron microscopy, biological tests in vivo and in vitro and structural. The analysis conducted in NXH8 did not show

Small heat shock protein αA-crystallin prevents photoreceptor degeneration in experimental autoimmune uveitis.

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Narsing A Rao

Full Text Available The small heat shock protein, αA-crystallin null (αA-/- mice are known to be more prone to retinal degeneration than the wild type mice in Experimental Autoimmune Uveoretinitis (EAU. In this report we demonstrate that intravenous administration of αA preserves retinal architecture and prevents photoreceptor damage in EAU. Interestingly, only αA and not αB-crystallin (αB, a closely related small heat shock protein works, pointing to molecular specificity in the observed retinal protection. The possible involvement of αA in retinal protection through immune modulation is corroborated by adaptive transfer experiments, (employing αA-/- and wild type mice with EAU as donors and Rag2-/- as the recipient mice, which indicate that αA protects against the autoimmune challenge by modulating the systemic B and T cell immunity. We show that αA administration causes marked reduction in Th1 cytokines (TNF-α, IL-12 and IFN-γ, both in the retina and in the spleen; notably, IL-17 was only reduced in the retina suggesting local intervention. Importantly, expression of Toll-like receptors and their associated adaptors is also inhibited suggesting that αA protection, against photoreceptor loss in EAU, is associated with systemic suppression of both the adaptive and innate immune responses.

Small heat shock protein αA-crystallin prevents photoreceptor degeneration in experimental autoimmune uveitis.

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Rao, Narsing A; Saraswathy, Sindhu; Pararajasegaram, Geeta; Bhat, Suraj P

2012-01-01

The small heat shock protein, αA-crystallin null (αA-/-) mice are known to be more prone to retinal degeneration than the wild type mice in Experimental Autoimmune Uveoretinitis (EAU). In this report we demonstrate that intravenous administration of αA preserves retinal architecture and prevents photoreceptor damage in EAU. Interestingly, only αA and not αB-crystallin (αB), a closely related small heat shock protein works, pointing to molecular specificity in the observed retinal protection. The possible involvement of αA in retinal protection through immune modulation is corroborated by adaptive transfer experiments, (employing αA-/- and wild type mice with EAU as donors and Rag2-/- as the recipient mice), which indicate that αA protects against the autoimmune challenge by modulating the systemic B and T cell immunity. We show that αA administration causes marked reduction in Th1 cytokines (TNF-α, IL-12 and IFN-γ), both in the retina and in the spleen; notably, IL-17 was only reduced in the retina suggesting local intervention. Importantly, expression of Toll-like receptors and their associated adaptors is also inhibited suggesting that αA protection, against photoreceptor loss in EAU, is associated with systemic suppression of both the adaptive and innate immune responses.

A study on effective thermal conductivity of crystalline layers in layer melt crystallization

International Nuclear Information System (INIS)

Kim, Kwang-Joo; Ulrich, Joachim

2002-01-01

An effective thermal conductivity in layer melt crystallization was explored based on a model considering inclusions inside a crystalline layer during crystal growth, molecular diffusion of inclusions migration due to temperature gradient and heat generation due to recrystallization of inclusions in the crystalline layer. The effective thermal conductivity increases with time, in general, as a result of compactness of the layer. Lower cooling temperature, i.e. greater supercooling, results in a more porous layer with lower effective thermal conductivity. A similar result is seen for the parameter of melt temperature, but less pronounced. A high concentration of the melt results in a high effective thermal conductivity while low concentration yields low effective thermal conductivity. At higher impurity levels in the melt phase, constitutional supercooling becomes more pronounced and unstable growth morphologies occur more easily. Cooling rate and Reynolds number also affect the effective thermal conductivity. The predictions of an effective thermal conductivity agree with the experimental data. The model was applied to estimate the thermal conductivities of the crystalline layer during layer melt crystallization. (author)

α-Crystallin localizes to the leading edges of migrating lens epithelial cells

International Nuclear Information System (INIS)

Maddala, Rupalatha; Vasantha Rao, P.

2005-01-01

α-crystallin (αA and αB) is a major lens protein, which belongs to the small heat-shock family of proteins and binds to various cytoskeletal proteins including actin, vimentin and desmin. In this study, we investigated the cellular localization of αA and αB-crystallins in migrating epithelial cells isolated from porcine lens. Immunofluorescence localization and confocal imaging of αB-crystallin in confluent and in migrating subconfluent cell cultures revealed a distinct pattern of subcellular distribution. While αB-crystallin localization was predominantly cytoplasmic in confluent cultures, it was strongly localized to the leading edges of cell membrane or the lamellipodia in migrating cells. In accordance with this pattern, we found abundant levels of αB-crystallin in membrane fractions compared to cytosolic and nuclear fractions in migrating lens epithelial cells. αA-crystallin, which has 60% sequence identity to αB-crystallin, also exhibited a distribution profile localizing to the leading edge of the cell membrane in migrating lens epithelial cells. Localization of αB-crystallin to the lamellipodia appears to be dependent on phosphorylation of residue serine-59. An inhibitor of p38 MAP kinase (SB202190), but not the ERK kinase inhibitor PD98059, was found to diminish localization of αB-crystallin to the lamellipodia, and this effect was found to be associated with reduced levels of Serine-59 phosphorylated αB-crystallin in SB202190-treated migrating lens epithelial cells. αB-crystallin localization to the lamellipodia was also altered by the treatment with RGD (Arg-Ala-Asp) peptide, dominant negative N17 Rac1 GTPase, cytochalasin D and Src kinase inhibitor (PP2), but not by the Rho kinase inhibitor Y-27632 or the myosin II inhibitor, blebbistatin. Additionally, in migrating lens epithelial cells, αB-crystallin exhibited a clear co-localization with the actin meshwork, β-catenin, WAVE-1, a promoter of actin nucleation, Abi-2, a component of WAVE

Investigation of the phase morphology of bacterial PHA inclusion bodies by contrast variation SANS

International Nuclear Information System (INIS)

Russell, R.A.; Holden, P.J.; Garvey, C.J.; Wilde, K.L.; Hammerton, K.M.; Foster, L.J.

2006-01-01

Under growth-limiting conditions, many bacteria are able to metabolise excess organic acids into polyhydroxyalkanoates (PHA) and store these polymers as intracellular inclusions until the return of favourable conditions. Various models have been proposed for the macromolecular organisation of the boundary layer surrounding the polymer, and contrast-variation small-angle neutron scattering (SANS) was used to study its organisation. Inclusions formed by Pseudomonas oleovorans under hydrogenating conditions showed lowest scattering intensity at ca. 20% D 2 O. The inclusions consist of protein and membrane lipids in the boundary layer and polyhydroxyoctanoate (lipid) in the inclusion body. At 20% D 2 O the contributions of lipids were contrast matched with the solvent, indicating that lipids contributed the bulk of the scattering intensity observed at other D 2 O/H 2 O ratios. These results are inconsistent with a model of the boundary layer which proposed outer and inner layers of crystalline protein lattice sandwiching a membrane lipid membrane layer [E.S. Stuart, R.W. Lenz, R.C. Fuller, Can J Microbiol 41(Suppl 1) (1995) 84-93], and is more consistent with a model consisting of a lipid monolayer containing embedded proteins [U. Pieper-furst, M.H. Madkour, F. Mayer, A. Steinbuchel, J. Bacteriol. 176 (1994) 4328-4337.] By altering the H/D content of the precursors, we were able to collect SANS data from preparations of both deuterated and H/D copolymer inclusions, where initial PHA produced was hydrogenated followed by deuteration. Deuterated inclusions showed minimum intensity above 90% D 2 O/H 2 O whereas the sequentially produced copolymer (assumed to be in a core/shell arrangement) displayed minimum scattering some 20% lower, which is consistent with the increased hydrogenation of the boundary layer expected from its synthesis during supply of hydrogenated followed by deuterated precursors

Purification of inclusion bodies using PEG precipitation under denaturing conditions to produce recombinant therapeutic proteins from Escherichia coli.

Science.gov (United States)

Chen, Huanhuan; Li, Ninghuan; Xie, Yueqing; Jiang, Hua; Yang, Xiaoyi; Cagliero, Cedric; Shi, Siwei; Zhu, Chencen; Luo, Han; Chen, Junsheng; Zhang, Lei; Zhao, Menglin; Feng, Lei; Lu, Huili; Zhu, Jianwei

2017-07-01

It has been documented that the purification of inclusion bodies from Escherichia coli by size exclusion chromatography (SEC) may benefit subsequent refolding and recovery of recombinant proteins. However, loading volume and the high cost of the column limits its application in large-scale manufacturing of biopharmaceutical proteins. We report a novel process using polyethylene glycol (PEG) precipitation under denaturing conditions to replace SEC for rapid purification of inclusion bodies containing recombinant therapeutic proteins. Using recombinant human interleukin 15 (rhIL-15) as an example, inclusion bodies of rhIL-15 were solubilized in 7 M guanidine hydrochloride, and rhIL-15 was precipitated by the addition of PEG 6000. A final concentration of 5% (w/v) PEG 6000 was found to be optimal to precipitate target proteins and enhance recovery and purity. Compared to the previously reported S-200 size exclusion purification method, PEG precipitation was easier to scale up and achieved the same protein yields and quality of the product. PEG precipitation also reduced manufacturing time by about 50 and 95% of material costs. After refolding and further purification, the rhIL-15 product was highly pure and demonstrated a comparable bioactivity with a rhIL-15 reference standard. Our studies demonstrated that PEG precipitation of inclusion bodies under denaturing conditions holds significant potential as a manufacturing process for biopharmaceuticals from E. coli protein expression systems.

Packaging protein drugs as bacterial inclusion bodies for therapeutic applications

Directory of Open Access Journals (Sweden)

Villaverde Antonio

2012-06-01

Full Text Available Abstract A growing number of insights on the biology of bacterial inclusion bodies (IBs have revealed intriguing utilities of these protein particles. Since they combine mechanical stability and protein functionality, IBs have been already exploited in biocatalysis and explored for bottom-up topographical modification in tissue engineering. Being fully biocompatible and with tuneable bio-physical properties, IBs are currently emerging as agents for protein delivery into mammalian cells in protein-replacement cell therapies. So far, IBs formed by chaperones (heat shock protein 70, Hsp70, enzymes (catalase and dihydrofolate reductase, grow factors (leukemia inhibitory factor, LIF and structural proteins (the cytoskeleton keratin 14 have been shown to rescue exposed cells from a spectrum of stresses and restore cell functions in absence of cytotoxicity. The natural penetrability of IBs into mammalian cells (reaching both cytoplasm and nucleus empowers them as an unexpected platform for the controlled delivery of essentially any therapeutic polypeptide. Production of protein drugs by biopharma has been traditionally challenged by IB formation. However, a time might have arrived in which recombinant bacteria are to be engineered for the controlled packaging of therapeutic proteins as nanoparticulate materials (nanopills, for their extra- or intra-cellular release in medicine and cosmetics.

Online analysis of protein inclusion bodies produced in E. coli by monitoring alterations in scattered and reflected light.

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Ude, Christian; Ben-Dov, Nadav; Jochums, André; Li, Zhaopeng; Segal, Ester; Scheper, Thomas; Beutel, Sascha

2016-05-01

The online monitoring of recombinant protein aggregate inclusion bodies during microbial cultivation is an immense challenge. Measurement of scattered and reflected light offers a versatile and non-invasive measurement technique. Therefore, we investigated two methods to detect the formation of inclusion bodies and monitor their production: (1) online 180° scattered light measurement (λ = 625 nm) using a sensor platform during cultivation in shake flask and (2) online measurement of the light reflective interference using a porous Si-based optical biosensor (SiPA). It could be shown that 180° scattered light measurement allows monitoring of alterations in the optical properties of Escherichia coli BL21 cells, associated with the formation of inclusion bodies during cultivation. A reproducible linear correlation between the inclusion body concentration of the non-fluorescent protein human leukemia inhibitory factor (hLIF) carrying a thioredoxin tag and the shift ("Δamp") in scattered light signal intensity was observed. This was also observed for the glutathione-S-transferase-tagged green fluorescent protein (GFP-GST). Continuous online monitoring of reflective interference spectra reveals a significant increase in the bacterium refractive index during hLIF production in comparison to a non-induced reference that coincide with the formation of inclusion bodies. These online monitoring techniques could be applied for fast and cost-effective screening of different protein expression systems.

The molecular chaperone function of α-crystallin is impaired by UV photolysis

International Nuclear Information System (INIS)

Borkman, R.F.; McLaughlin, J.

1995-01-01

Buffer solutions of the lens protein γ-crystallin and the enzymes aldolase and liver alcohol dehydrogenase became turbid and formed solid precipitate upon exposure to an elevated temperature of 63 o C or to UV radiation at 308 nm. When α-crystallin was added to the protein solutions in stoichiometric amounts, heat or UV irradiation did not cause turbidity, or turbidity developed much less rapidly than in the absence of α-crystallin. Hence, normal α-crystallin functioned as a ''molecular chaperone,'' providing protection against both UV and heat-induced protein aggregation. When α-crystallin was preirradiated with UV at 308 nm, its ability to function as a chaperone vis-a-vis both UV and heat-induced aggregation was significantly impaired, but only at relatively high UV doss. (author)

Influence of nano-inclusions' grain boundaries on crack propagation modes in materials

International Nuclear Information System (INIS)

Karakasidis, T.E.; Charitidis, C.A.

2011-01-01

The effect of nano-inclusions on materials' strength and toughness has attracted great interest in recent years. It has been shown that tuning the morphological and microstructural features of materials can tailor their fracture modes. The existence of a characteristic size of inclusions that favours the fracture mode (i.e. transgranular or intergranular) has been experimentally observed but also predicted by a 2D model based on energetic arguments which relates the crack propagation mode to the ratio of the interface area between the crystalline inclusion and the matrix with the area of the crystallite inclusion in a previous work. In the present work, a 3D model is proposed in order to extend the 2D model and take into account the influence of the size of grain boundary zone on the toughening/hardening behavior of the material as it was observed experimentally in the literature. The model relates crack propagation mode to the ratio of the volume of the grain boundary zone between the crystalline inclusion and the matrix with the volume of the nano-inclusion. For a ratio below a critical value, transgranular propagation is favoured while for larger values, intergranular propagation is favoured. We also demonstrate that the extent of the grain boundary region also can significantly affect this critical value. The results of the model are in agreement with the literature experimental observations related to the toughening/hardening behavior as a function of the size of crystalline inclusions as well as the width of the grain boundary regions.

Role of αA-crystallin-derived αA66-80 peptide in guinea pig lens crystallin aggregation and insolubilization.

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Raju, Murugesan; Mooney, Brian P; Thakkar, Kavi M; Giblin, Frank J; Schey, Kevin L; Sharma, K Krishna

2015-03-01

Earlier we reported that low molecular weight (LMW) peptides accumulate in aging human lens tissue and that among the LMW peptides, the chaperone inhibitor peptide αA66-80, derived from α-crystallin protein, is one of the predominant peptides. We showed that in vitro αA66-80 induces protein aggregation. The current study was undertaken to determine whether LMW peptides are also present in guinea pig lens tissue subjected to hyperbaric oxygen (HBO) in vivo. The nuclear opacity induced by HBO in guinea pig lens is the closest animal model for studying age-related cataract formation in humans. A LMW peptide profile by mass spectrometry showed the presence of an increased amount of LMW peptides in HBO-treated guinea pig lenses compared to age-matched controls. Interestingly, the mass spectrometric data also showed that the chaperone inhibitor peptide αA66-80 accumulates in HBO-treated guinea pig lens. Following incubation of synthetic chaperone inhibitor peptide αA66-80 with α-crystallin from guinea pig lens extracts, we observed a decreased ability of α-crystallin to inhibit the amorphous aggregation of the target protein alcohol dehydrogenase and the formation of large light scattering aggregates, similar to those we have observed with human α-crystallin and αA66-80 peptide. Further, time-lapse recordings showed that a preformed complex of α-crystallin and αA66-80 attracted additional crystallin molecules to form even larger aggregates. These results demonstrate that LMW peptide-mediated cataract development in aged human lens and in HBO-induced lens opacity in the guinea pig may have common molecular pathways. Copyright © 2015 Elsevier Ltd. All rights reserved.

The Effect of Attractive Interactions and Macromolecular Crowding on Crystallins Association.

Directory of Open Access Journals (Sweden)

Jiachen Wei

Full Text Available In living systems proteins are typically found in crowded environments where their effective interactions strongly depend on the surrounding medium. Yet, their association and dissociation needs to be robustly controlled in order to enable biological function. Uncontrolled protein aggregation often causes disease. For instance, cataract is caused by the clustering of lens proteins, i.e., crystallins, resulting in enhanced light scattering and impaired vision or blindness. To investigate the molecular origins of cataract formation and to design efficient treatments, a better understanding of crystallin association in macromolecular crowded environment is needed. Here we present a theoretical study of simple coarse grained colloidal models to characterize the general features of how the association equilibrium of proteins depends on the magnitude of intermolecular attraction. By comparing the analytic results to the available experimental data on the osmotic pressure in crystallin solutions, we identify the effective parameters regimes applicable to crystallins. Moreover, the combination of two models allows us to predict that the number of binding sites on crystallin is small, i.e. one to three per protein, which is different from previous estimates. We further observe that the crowding factor is sensitive to the size asymmetry between the reactants and crowding agents, the shape of the protein clusters, and to small variations of intermolecular attraction. Our work may provide general guidelines on how to steer the protein interactions in order to control their association.

Refolding of proteins from inclusion bodies: rational design and recipes.

Science.gov (United States)

Basu, Anindya; Li, Xiang; Leong, Susanna Su Jan

2011-10-01

The need to develop protein biomanufacturing platforms that can deliver proteins quickly and cost-effectively is ever more pressing. The rapid rate at which genomes can now be sequenced demands efficient protein production platforms for gene function identification. There is a continued need for the biotech industry to deliver new and more effective protein-based drugs to address new diseases. Bacterial production platforms have the advantage of high expression yields, but insoluble expression of many proteins necessitates the development of diverse and optimised refolding-based processes. Strategies employed to eliminate insoluble expression are reviewed, where it is concluded that inclusion bodies are difficult to eliminate for various reasons. Rational design of refolding systems and recipes are therefore needed to expedite production of recombinant proteins. This review article discusses efforts towards rational design of refolding systems and recipes, which can be guided by the development of refolding screening platforms that yield both qualitative and quantitative information on the progression of a given refolding process. The new opportunities presented by light scattering technologies for developing rational protein refolding buffer systems which in turn can be used to develop new process designs armed with better monitoring and controlling functionalities are discussed. The coupling of dynamic and static light scattering methodologies for incorporation into future bioprocess designs to ensure delivery of high-quality refolded proteins at faster rates is also discussed.

Crystallization and evaluation of hen egg-white lysozyme crystals for protein pH titration in the crystalline state

International Nuclear Information System (INIS)

Iwai, Wakari; Yagi, Daichi; Ishikawa, Takuya; Ohnishi, Yuki; Tanaka, Ichiro; Niimura, Nobuo

2008-01-01

Hen egg-white lysozyme was crystallized over a wide pH range (2.5–8.0) and the quality of the crystals was characterized. Crystallization phase diagrams at pH 2.5, 6.0 and 7.5 were determined To observe the ionized status of the amino acid residues in proteins at different pH (protein pH titration in the crystalline state) by neutron diffraction, hen egg-white lysozyme was crystallized over a wide pH range (2.5–8.0). Crystallization phase diagrams at pH 2.5, 6.0 and 7.5 were determined. At pH < 4.5 the border between the metastable region and the nucleation region shifted to the left (lower precipitant concentration) in the phase diagram, and at pH > 4.5 the border shifted to the right (higher precipitant concentration). The qualities of these crystals were characterized using the Wilson plot method. The qualities of all crystals at different pH were more or less equivalent (B-factor values within 25–40). It is expected that neutron diffraction analysis of these crystals of different pH provides equivalent data in quality for discussions of protein pH titration in the crystalline state of hen egg-white lysozyme

Rab11-family of interacting protein 2 associates with chlamydial inclusions through its Rab-binding domain and promotes bacterial multiplication.

Science.gov (United States)

Leiva, Natalia; Capmany, Anahí; Damiani, María Teresa

2013-01-01

Chlamydia trachomatis, an obligate intracellular pathogen, survives within host cells in a special compartment named 'inclusion' and takes advantage of host vesicular transport pathways for its growth and replication. Rab GTPases are key regulatory proteins of intracellular trafficking. Several Rabs, among them Rab11 and Rab14, are implicated in chlamydial development. FIP2, a member of the Rab11-Family of Interacting Proteins, presents at the C-terminus a Rab-binding domain that interacts with both Rab11 and Rab14. In this study, we determined and characterized the recruitment of endogenous and GFP-tagged FIP2 to the chlamydial inclusions. The recruitment of FIP2 is specific since other members of the Rab11-Family of Interacting Proteins do not associate with the chlamydial inclusions. The Rab-binding domain of FIP2 is essential for its association. Our results indicate that FIP2 binds to Rab11 at the chlamydial inclusion membrane through its Rab-binding domain. The presence of FIP2 at the chlamydial inclusion favours the recruitment of Rab14. Furthermore, our results show that FIP2 promotes inclusion development and bacterial replication. In agreement, the silencing of FIP2 decreases the bacterial progeny. C. trachomatis likely recruits FIP2 to hijack host intracellular trafficking to redirect vesicles full of nutrients towards the inclusion. © 2012 Blackwell Publishing Ltd.

Catalytically-active inclusion bodies-Carrier-free protein immobilizates for application in biotechnology and biomedicine.

Science.gov (United States)

Krauss, Ulrich; Jäger, Vera D; Diener, Martin; Pohl, Martina; Jaeger, Karl-Erich

2017-09-20

Bacterial inclusion bodies (IBs) consist of unfolded protein aggregates and represent inactive waste products often accumulating during heterologous overexpression of recombinant genes in Escherichia coli. This general misconception has been challenged in recent years by the discovery that IBs, apart from misfolded polypeptides, can also contain substantial amounts of active and thus correctly or native-like folded protein. The corresponding catalytically-active inclusion bodies (CatIBs) can be regarded as a biologically-active sub-micrometer sized biomaterial or naturally-produced carrier-free protein immobilizate. Fusion of polypeptide (protein) tags can induce CatIB formation paving the way towards the wider application of CatIBs in synthetic chemistry, biocatalysis and biomedicine. In the present review we summarize the history of CatIBs, present the molecular-biological tools that are available to induce CatIB formation, and highlight potential lines of application. In the second part findings regarding the formation, architecture, and structure of (Cat)IBs are summarized. Finally, an overview is presented about the available bioinformatic tools that potentially allow for the prediction of aggregation and thus (Cat)IB formation. This review aims at demonstrating the potential of CatIBs for biotechnology and hopefully contributes to a wider acceptance of this promising, yet not widely utilized, protein preparation. Copyright © 2017 Elsevier B.V. All rights reserved.

Nanosized f.c.c. thallium inclusions in aluminium

International Nuclear Information System (INIS)

Johnson, E.; Johansen, A.; Thoft, N.B.; Andersen, H.H.; Sarholt-Kristensen, L.

1993-01-01

Ion implantation of pure aluminium with thallium induces the formation of nanosized crystalline inclusions of thallium with a f.c.c. structure. The size of the inclusions depends on the implantation conditions and subsequent annealing treatments and is typically in the range from 1 to 10 nm. The inclusions are aligned topotactically with the aluminium matrix with a cube-cube orientation relationship and they have a truncated octahedral shape bounded by {111} and {001} planes. The lattice parameter of the f.c.c. thallium inclusions is 0.484 ± 0.002 nm, which is slightly but significantly larger than in the high-pressure f.c.c. thallium phase known to be stable above 3.8 GPa. (Author)

Micromechanical modeling of the elasto-viscoplastic bahavior of semi-crystalline polymers

NARCIS (Netherlands)

Dommelen, van J.A.W.; Parks, D.M.; Boyce, M.C.; Brekelmans, W.A.M.; Baaijens, F.P.T.

2003-01-01

A micromechanically-based constitutive model for the elasto-viscoplastic deformationand texture evolution of semi-crystalline polymers is developed. The modelidealizes the microstructure to consist of an aggregate of two-phase layered compositeinclusions. A new framework for the composite inclusion

Effect of glycation on α-crystallin structure and chaperone-like function

Science.gov (United States)

Kumar, P. Anil; Kumar, M. Satish; Reddy, G. Bhanuprakash

2007-01-01

The chaperone-like activity of α-crystallin is considered to play an important role in the maintenance of the transparency of the eye lens. However, in the case of aging and in diabetes, the chaperone function of α-crystallin is compromized, resulting in cataract formation. Several post-translational modifications, including non-enzymatic glycation, have been shown to affect the chaperone function of α-crystallin in aging and in diabetes. A variety of agents have been identified as the predominant sources for the formation of AGEs (advanced glycation end-products) in various tissues, including the lens. Nevertheless, glycation of α-crystallin with various sugars has resulted in divergent results. In the present in vitro study, we have investigated the effect of glucose, fructose, G6P (glucose 6-phosphate) and MGO (methylglyoxal), which represent the major classes of glycating agents, on the structure and chaperone function of α-crystallin. Modification of α-crystallin with all four agents resulted in the formation of glycated protein, increased AGE fluorescence, protein cross-linking and HMM (high-molecular-mass) aggregation. Interestingly, these glycation-related profiles were found to vary with different glycating agents. For instance, CML [Nϵ-(carboxymethyl)lysine] was the predominant AGE formed upon glycation of α-crystallin with these agents. Although fructose and MGO caused significant conformational changes, there were no significant structural perturbations with glucose and G6P. With the exception of MGO modification, glycation with other sugars resulted in decreased chaperone activity in aggregation assays. However, modification with all four sugars led to the loss of chaperone activity as assessed using an enzyme inactivation assay. Glycation-induced loss of α-crystallin chaperone activity was associated with decreased hydrophobicity. Furthermore, α-crystallin isolated from glycated TSP (total lens soluble protein) had also increased AGE

Active tuberculosis patients have high levels of IgA anti-alpha-crystallin and isocitrate lyase proteins.

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Talavera-Paulín, M; García-Morales, L; Ruíz-Sánchez, B P; Caamal-Ley, Á D; Hernández-Solis, A; Ramírez-Casanova, E; Cicero-Sabido, R; Espitia, C; Helguera-Repetto, C; González-Y-Merchand, J A; Flores-Mejía, R; Estrada-Parra, S; Estrada-García, I; Chacón-Salinas, R; Wong-Baeza, I; Serafín-López, J

2016-12-01

Mexico City, Mexico. To identify proteins synthetised by Mycobacterium tuberculosis in hypoxic culture, which resemble more closely a granuloma environment than aerobic culture, and to determine if they are recognised by antibodies from patients with active pulmonary tuberculosis (PTB). Soluble extracts from M. tuberculosis H37Rv cultured under aerobic or hypoxic conditions were analysed using two-dimensional polyacrylamide gel electrophoresis, and proteins over-expressed under hypoxia were identified by mass spectrometry. The presence of immunoglobulin (Ig) G, IgA and IgM antibodies against these proteins was determined in the serum of 42 patients with active PTB and 42 healthy controls. We selected three M. tuberculosis H37Rv proteins (alpha-crystallin protein [Acr, Rv2031c], universal stress protein Rv2623 and isocitrate lyase [ICL, RV0467]) that were over-expressed under hypoxia. Titres of anti-Acr and anti-ICL IgA antibodies were higher in patients than in healthy controls, with an area under the receiver operating characteristic curve of 0.71 for anti-ICL IgA antibodies. ICL could be used in combination with other M. tuberculosis antigens to improve the sensitivity and specificity of current serological TB diagnostic methods.

Different alpha crystallin expression in human age-related and congenital cataract lens epithelium.

Science.gov (United States)

Yang, Jing; Zhou, Sheng; Guo, Minfei; Li, Yuting; Gu, Jianjun

2016-05-28

The purpose of this study was to investigate the different expressions of αA-crystallin and αB-crystallin in human lens epithelium of age-related and congenital cataracts. The central part of the human anterior lens capsule approximately 5 mm in diameter together with the adhering epithelial cells, were harvested and processed within 6 hours after cataract surgery from age-related and congenital cataract patients or from normal eyes of fresh cadavers. The mRNA and soluble protein levels of αA-crystallin and αB-crystallin in the human lens epithelium were detected by real-time PCR and western blots, respectively. The mRNA and soluble protein expressions of αA-crystallin and αB-crystallin in the lens epithelium were both reduced in age-related and congenital cataract groups when compared with the normal control group. However, the degree of α-crystallin loss in the lens epithelium was highly correlated with different cataract types. The α-crystallin expression of the lens epithelium was greatly reduced in the congenital cataract group but only moderately decreased in the age-related cataract group. The reduction of αA-crystallin soluble protein levels in the congenital cataract group was approximately 2.4 fold decrease compared with that of the age-related cataract group, while an mRNA fold change of 1.67 decrease was observed for the age-related cataract group. Similarly, the reduction of soluble protein levels of αB-crystallin in the congenital cataract group was approximately a 1.57 fold change compared with that of the age-related cataract group. A 1.75 fold change for mRNA levels compared with that of the age-related cataract group was observed. The results suggest that the differential loss of α-crystallin in the human lens epithelium could be associated with the different mechanisms of cataractogenesis in age-related versus congenital cataracts, subsequently resulting in different clinical presentations.

Crystallization and preliminary X-ray crystallographic investigations on a βγ-crystallin domain of absent in melanoma 1 (AIM1), a protein from Homo sapiens

International Nuclear Information System (INIS)

Aravind, Penmatsa; Rajini, Bheemreddy; Sharma, Yogendra; Sankaranarayanan, Rajan

2006-01-01

The crystallization and preliminary X-ray diffraction analysis of AIM1g1, a βγ-crystallin domain of absent in melanoma (AIM1) protein from H. sapiens, is reported. AIM1g1 is a single βγ-crystallin domain from the protein absent in melanoma 1 (AIM1), which appears to play a role in the suppression of melanomas. This domain is known to bind calcium and its structure would help in identifying calcium-coordinating sites in vertebrate crystallins, which have hitherto been believed to have lost this ability during evolution. Crystallization of this domain was performed by the hanging-drop vapour-diffusion method. Crystals diffracted to a maximum resolution of 1.86 Å and were found to belong to space group P6 1 or P6 5 , with unit-cell parameters a = b = 54.98, c = 59.73 Å. Solvent-content analysis indicated the presence of one monomer per asymmetric unit

Crystallization and preliminary X-ray crystallographic investigations on a βγ-crystallin domain of absent in melanoma 1 (AIM1), a protein from Homo sapiens

Energy Technology Data Exchange (ETDEWEB)

Aravind, Penmatsa; Rajini, Bheemreddy; Sharma, Yogendra; Sankaranarayanan, Rajan, E-mail: sankar@ccmb.res.in [Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad 500 007 (India)

2006-03-01

The crystallization and preliminary X-ray diffraction analysis of AIM1g1, a βγ-crystallin domain of absent in melanoma (AIM1) protein from H. sapiens, is reported. AIM1g1 is a single βγ-crystallin domain from the protein absent in melanoma 1 (AIM1), which appears to play a role in the suppression of melanomas. This domain is known to bind calcium and its structure would help in identifying calcium-coordinating sites in vertebrate crystallins, which have hitherto been believed to have lost this ability during evolution. Crystallization of this domain was performed by the hanging-drop vapour-diffusion method. Crystals diffracted to a maximum resolution of 1.86 Å and were found to belong to space group P6{sub 1} or P6{sub 5}, with unit-cell parameters a = b = 54.98, c = 59.73 Å. Solvent-content analysis indicated the presence of one monomer per asymmetric unit.

The role of macromolecular crowding in the evolution of lens crystallins with high molecular refractive index

International Nuclear Information System (INIS)

Zhao, Huaying; Magone, M Teresa; Schuck, Peter

2011-01-01

Crystallins are present in the lens at extremely high concentrations in order to provide transparency and generate a high refractive power of the lens. The crystallin families prevalent in the highest density lens tissues are γ-crystallins in vertebrates and S-crystallins in cephalopods. As shown elsewhere, in parallel evolution, both have evolved molecular refractive index increments 5–10% above those of most proteins. Although this is a small increase, it is statistically very significant and can be achieved only by very unusual amino acid compositions. In contrast, such a molecular adaptation to aid in the refractive function of the lens did not occur in crystallins that are preferentially located in lower density lens tissues, such as vertebrate α-crystallin and taxon-specific crystallins. In the current work, we apply a model of non-interacting hard spheres to examine the thermodynamic contributions of volume exclusion at lenticular protein concentrations. We show that the small concentration decrease afforded by the higher molecular refractive index increment of crystallins can amplify nonlinearly to produce order of magnitude differences in chemical activities, and lead to reduced osmotic pressure and the reduced propensity for protein aggregation. Quantitatively, this amplification sets in only at protein concentrations as high as those found in hard lenses or the nucleus of soft lenses, in good correspondence to the observed crystallin properties in different tissues and different species. This suggests that volume exclusion effects provide the evolutionary driving force for the unusual refractive properties and the unusual amino acid compositions of γ-crystallins and S-crystallins

Production and purification of avian antibodies (IgYs from inclusion bodies of a recombinant protein central in NAD+ metabolism

Directory of Open Access Journals (Sweden)

Paula A. Moreno-González

2013-08-01

Full Text Available The use of hens for the production of polyclonal antibodies reduces animal intervention and moreover yields a higher quantity of antibodies than other animal models.  The phylogenetic distance between bird and mammal antigens, often leads to more specific avian antibodies than their mammalian counterparts.Since a large amount of antigen is required for avian antibody production, the use of recombinant proteins for this procedure has been growing faster over the last years. Nevertheless, recombinant protein production through heterologous systems frequently prompts the protein to precipitate, forming insoluble aggregates of limited utility (inclusion bodies. A methodology for the production of avian polyclonal antibodies, using recombinant protein from inclusion bodies is presented in this article.In order to produce the antigen, a recombinant Nicotinamide mononucleotide adenylyltransferase from Giardia intestinalis (His-GiNMNAT was expressed in Escherichia coli.  The protein was purified through solubilization from inclusion bodies prior to its renaturalization.  Antibodies were purified from egg yolk of immunized hens by water dilution, followed by ammonium sulfate precipitation and thiophilic affinity chromatography.The purified antibodies were tested against His-GiNMNAT protein in Western blot essays. From one egg yolk, 14.4 mg of highly pure IgY were obtained; this antibody was able to detect 15ng of His-GiNMNAT.  IgY specificity was improved by means of antigen affinity purification, allowing its use for parasite protein recognition.

Upon Infection the Cellular WD Repeat-containing Protein 5 (WDR5) Localizes to Cytoplasmic Inclusion Bodies and Enhances Measles Virus Replication.

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Ma, Dzwokai; George, Cyril X; Nomburg, Jason; Pfaller, Christian K; Cattaneo, Roberto; Samuel, Charles E

2017-12-13

Replication of negative-strand RNA viruses occurs in association with discrete cytoplasmic foci called inclusion bodies. Whereas inclusion bodies represent a prominent subcellular structure induced by viral infection, our knowledge of the cellular protein components involved in inclusion body formation and function is limited. Using measles virus-infected HeLa cells, we found that the WD repeat-containing protein 5 (WDR5), a subunit of histone H3 lysine 4 methyltransferases, was selectively recruited to virus-induced inclusion bodies. Furthermore, WDR5 was found in complexes containing viral proteins associated with RNA replication. WDR5 was not detected with mitochondria, stress granules, or other known secretory or endocytic compartments of infected cells. WDR5 deficiency decreased both viral protein production and infectious virus yields. Interferon production was modestly increased in WDR5 deficient cells. Thus, our study identifies WDR5 as a novel viral inclusion body-associated cellular protein and suggests a role for WDR5 in promoting viral replication. IMPORTANCE Measles virus is a human pathogen that remains a global concern with more than 100,000 measles-related deaths annually despite the availability of an effective vaccine. As measles continues to cause significant morbidity and mortality, understanding the virus-host interactions at the molecular level that affect virus replication efficiency is important for development and optimization of treatment procedures. Measles virus is an RNA virus that encodes six genes and replicates in the cytoplasm of infected cells in discrete cytoplasmic replication bodies, though little is known of the biochemical nature of these structures. Here we show that the cellular protein WDR5 is enriched in the cytoplasmic viral replication factories and enhances virus growth. WDR5-containing protein complex includes viral proteins responsible for viral RNA replication. Thus, we have identified WDR5 as a host factor that

Evidence of yttrium silicate inclusions in YSZ-porcelain veneers.

Science.gov (United States)

Stoner, Brian R; Griggs, Jason A; Neidigh, John; Piascik, Jeffrey R

2014-04-01

This report introduces the discovery of crystalline defects that can form in the porcelain veneering layer when in contact with yttria-stabilized zirconia (YSZ). The focus was on dental prostheses and understanding the defects that form in the YSZ/porcelain system; however the data reported herein may have broader implications toward the use and stability of YSZ-based ceramics in general. Specimens were cut from fully sintered YSZ plates and veneering porcelain was applied (X-ray (EDAX) was used for microstructural and elemental analysis. EDAX, for chemical analysis and transmission electron diffraction (TED) for structural analysis were both performed in the transmission electron microscope (TEM). Additionally, in order to spatially resolve Y-rich precipitates, micro-CT scans were conducted at varying depths within the porcelain veneer. Local EDAX (SEM) was performed in the regions of visible inclusions and showed significant increases in yttrium concentration. TEM specimens also showed apparent inclusions in the porcelain and selected area electron diffraction was performed on these regions and found the inclusions to be crystalline and identified as either yttrium-silicate (Y2 SiO5 ) or yttrium-disilicate (Y2 Si2 O7 ). Micro-CT data showed that yttrium-silicate precipitates were distributed throughout the thickness of the porcelain veneer. Future studies are needed to determine whether many of the premature failures associated with this materials system may be the result of crystalline flaws that form as a result of high temperature yttrium diffusion near the surfaces of YSZ. © 2013 Wiley Periodicals, Inc.

Alternative preparation of inclusion bodies excludes interfering non-protein contaminants and improves the yield of recombinant proinsulin.

Science.gov (United States)

Mackin, Robert B

2014-01-01

The goal of simple, high-yield expression and purification of recombinant human proinsulin has proven to be a considerable challenge. First, proinsulin forms inclusion bodies during bacterial expression. While this phenomenon can be exploited as a capture step, conventionally prepared inclusion bodies contain significant amounts of non-protein contaminants that interfere with subsequent chromatographic purification. Second, the proinsulin molecules within the inclusion bodies are incorrectly folded, and likely cross-linked to one another, making it difficult to quantify the amount of expressed proinsulin. Third, proinsulin is an intermediate between the initial product of ribosomal translation (preproinsulin) and the final product secreted by pancreatic beta cells (insulin). Therefore, to be efficiently produced in bacteria, it must be produced as an N-terminally extended fusion protein, which has to be converted to authentic proinsulin during the purification scheme. To address all three of these problems, while simultaneously streamlining the procedure and increasing the yield of recombinant proinsulin, we have made three substantive modifications to our previous method for producing proinsulin:.•Conditions for the preparation of inclusion bodies have been altered so contaminants that interfere with semi-preparative reversed-phase chromatography are excluded while the proinsulin fusion protein is retained at high yield.•Aliquots are taken following important steps in the procedure and the quantity of proinsulin-related polypeptide in the sample is compared to the amount present prior to that step.•Final purification is performed using a silica-based reversed-phase matrix in place of a polystyrene-divinylbenzene-based matrix.

Location of catalase in crystalline peroxisomes of methanol-grown Hansenula polymorpha

NARCIS (Netherlands)

Keizer, Ineke; Roggenkamp, Rainer; Harder, Willem; Veenhuis, Marten

1992-01-01

We have studied the intraperoxisomal location of catalase in peroxisomes of methanol-grown Hansenula polymorpha by (immuno)cytochemical means. In completely crystalline peroxisomes, in which the crystalline matrix is composed of octameric alcohol oxidase (AO) molecules, most of the catalase protein

Autophagy and UPR in alpha-crystallin mutant knock-in mouse models of hereditary cataracts.

Science.gov (United States)

Andley, Usha P; Goldman, Joshua W

2016-01-01

Knock-in mice provide useful models of congenital and age-related cataracts caused by α-crystallin mutations. R49C αA-crystallin and R120G αB-crystallin mutations are linked with hereditary cataracts. Knock-in αA-R49C+/- heterozygotes develop cataracts by 1-2months, whereas homozygote mice have cataracts at birth. The R49C mutation drastically reduces lens protein water solubility and causes cell death in knock-in mouse lenses. Mutant crystallin cannot function as a chaperone, which leads to protein aggregation and lens opacity. Protein aggregation disrupts the lens fiber cell structure and normal development and causes cell death in epithelial and fiber cells. We determined what aspects of the wild-type phenotype are age-dependently altered in the mutant lens. Wild-type, heterozygote (αA-R49C+/-), and homozygote (αA-R49C+/+) mouse lenses were assessed pre- and postnatally for lens morphology (electron microscopy, immunohistochemistry), and autophagy or unfolded protein response markers (immunoblotting). Morphology was altered by embryonic day 17 in R49C+/+ lenses; R49C+/- lens morphology was unaffected at this stage. Active autophagy in the lens epithelium of mutant lenses was indicated by the presence of autophagosomes using electron microscopy. Protein p62 levels, which are degraded specifically by autophagy, increased in αA-R49C mutant versus wild-type lenses, suggesting autophagy inhibition in the mutant lenses. The unfolded protein response marker XBP-1 was upregulated in adult lenses of αB-R120G+/+ mice, suggesting its role in lens opacification. Mutated crystallins alter lens morphology, autophagy, and stress responses. Therapeutic modulation of autophagic pathways may improve protein degradation in cataractous lenses and reduce lens opacity. This article is part of a Special Issue entitled Crystallin Biochemistry in Health and Disease. Copyright © 2015 Elsevier B.V. All rights reserved.

IC-tagged proteins are able to interact with each other and perform complex reactions when integrated into muNS-derived inclusions.

Science.gov (United States)

Brandariz-Nuñez, Alberto; Otero-Romero, Iria; Benavente, Javier; Martinez-Costas, Jose M

2011-09-20

We have recently developed a versatile tagging system (IC-tagging) that causes relocation of the tagged proteins to ARV muNS-derived intracellular globular inclusions. In the present study we demonstrate (i) that the IC-tag can be successfully fused either to the amino or carboxyl terminus of the protein to be tagged and (ii) that IC-tagged proteins are able to interact between them and perform complex reactions that require such interactions while integrated into muNS inclusions, increasing the versatility of the IC-tagging system. Also, our studies with the DsRed protein add some light on the structure/function relationship of the evolution of DsRed chromophore. Copyright © 2011 Elsevier B.V. All rights reserved.

Isolation of cell-free bacterial inclusion bodies.

Science.gov (United States)

Rodríguez-Carmona, Escarlata; Cano-Garrido, Olivia; Seras-Franzoso, Joaquin; Villaverde, Antonio; García-Fruitós, Elena

2010-09-17

Bacterial inclusion bodies are submicron protein clusters usually found in recombinant bacteria that have been traditionally considered as undesirable products from protein production processes. However, being fully biocompatible, they have been recently characterized as nanoparticulate inert materials useful as scaffolds for tissue engineering, with potentially wider applicability in biomedicine and material sciences. Current protocols for inclusion body isolation from Escherichia coli usually offer between 95 to 99% of protein recovery, what in practical terms, might imply extensive bacterial cell contamination, not compatible with the use of inclusion bodies in biological interfaces. Using an appropriate combination of chemical and mechanical cell disruption methods we have established a convenient procedure for the recovery of bacterial inclusion bodies with undetectable levels of viable cell contamination, below 10⁻¹ cfu/ml, keeping the particulate organization of these aggregates regarding size and protein folding features. The application of the developed protocol allows obtaining bacterial free inclusion bodies suitable for use in mammalian cell cultures and other biological interfaces.

Comparative study to develop a single method for retrieving wide class of recombinant proteins from classical inclusion bodies.

Science.gov (United States)

Padhiar, Arshad Ahmed; Chanda, Warren; Joseph, Thomson Patrick; Guo, Xuefang; Liu, Min; Sha, Li; Batool, Samana; Gao, Yifan; Zhang, Wei; Huang, Min; Zhong, Mintao

2018-03-01

The formation of inclusion bodies (IBs) is considered as an Achilles heel of heterologous protein expression in bacterial hosts. Wide array of techniques has been developed to recover biochemically challenging proteins from IBs. However, acquiring the active state even from the same protein family was found to be an independent of single established method. Here, we present a new strategy for the recovery of wide sub-classes of recombinant protein from harsh IBs. We found that numerous methods and their combinations for reducing IB formation and producing soluble proteins were not effective, if the inclusion bodies were harsh in nature. On the other hand, different practices with mild solubilization buffers were able to solubilize IBs completely, yet the recovery of active protein requires large screening of refolding buffers. With the integration of previously reported mild solubilization techniques, we proposed an improved method, which comprised low sarkosyl concentration, ranging from 0.05 to 0.1% coupled with slow freezing (- 1 °C/min) and fast thaw (room temperature), resulting in greater solubility and the integrity of solubilized protein. Dilution method was employed with single buffer to restore activity for every sub-class of recombinant protein. Results showed that the recovered protein's activity was significantly higher compared with traditional solubilization/refolding approach. Solubilization of IBs by the described method was proved milder in nature, which restored native-like conformation of proteins within IBs.

(alpha)B-crystallin in cerebrospinal fluid of patients with multiple sclerosis

DEFF Research Database (Denmark)

Støvring, Birgitte; Vang, Ole; Christiansen, Michael

2005-01-01

Background: aB-crystallin is a chaperone protein and a potential myelin antigen to human T cells in Multiple Sclerosis (MS). In this study we investigate the existence of aB-crystallin in the cerebrospinal fluid (CSF) of patients with clinical symptoms of MS and control individuals without...

Reverse-phase HPLC analysis of human alpha crystallin.

Science.gov (United States)

Swamy, M S; Abraham, E C

1991-03-01

A rapid and highly sensitive reverse-phase HPLC (RP-HPLC) method was used to separate crystallin subunits from human alpha crystallin. Three distinct peaks were separated; by electrophoretic and immunological analyses the first and second peaks were identified as alpha B and alpha A respectively. On the other hand, peak 3 appeared to be a modified form of alpha crystallin. The ratio of alpha A and alpha B proteins was 3:1 in 1 day old lenses which gradually changed to 2:1 in 17 year old lenses and to 1:1 in the 50 and 82 year old whole lenses and 82 year old lens cortex, with a concomitant increase in the modified alpha, suggesting that alpha A subunits are relatively more involved in aggregation. Analysis of the 82 year old lens nucleus also supported this conclusion. The RP-HPLC analysis of the HMW aggregate fraction showed substantial enrichment of the modified alpha. The alpha A and alpha B subunits independently reassociated to form polymeric alpha crystallin whereas the modified alpha reassociated to form HMW aggregates as shown by molecular sieve HPLC. Hence it appears that the HMW aggregate peak was constituted by modified alpha crystallin. Only in the peak 3 material the 280 nm absorbance was about 2-fold higher than what was expected from the actual protein content. The data suggest that the changes induced by post-translational modifications may have some role in the formation of modified alpha. The present RP-HPLC method is useful in separating these modified alpha from the unmodified alpha A and alpha B subunits.

αB-crystallin is essential for the TGF-β2-mediated epithelial to mesenchymal transition of lens epithelial cells.

Science.gov (United States)

Nahomi, Rooban B; Pantcheva, Mina B; Nagaraj, Ram H

2016-05-15

Transforming growth factor (TGF)-β2-mediated pathways play a major role in the epithelial to mesenchymal transition (EMT) of lens epithelial cells (LECs) during secondary cataract formation, which is also known as posterior capsule opacification (PCO). Although αB-crystallin is a major protein in LEC, its role in the EMT remains unknown. In a human LEC line (FHL124), TGF-β2 treatment resulted in changes in the EMT-associated proteins at the mRNA and protein levels. This was associated with nuclear localization of αB-crystallin, phosphorylated Smad2 (pSmad2) (S245/250/255), pSmad3 (S423/425), Smad4 and Snail and the binding of αB-crystallin to these transcription factors, all of which were reduced by the down-regulation of αB-crystallin. Expression of the functionally defective R120G mutant of αB-crystallin reduced TGF-β2-induced EMT in LECs of αB-crystallin knockout (KO) mice. Treatment of bovine lens epithelial explants and mouse LEC with TGF-β2 resulted in changes in the EMT-associated proteins at the mRNA and protein levels. This was accompanied by increase in phosphorylation of p44/42 mitogen-activated protein kinases (MAPK) (T202/Y204), p38 MAPK (T180/Y182), protein kinase B (Akt) (S473) and Smad2 when compared with untreated cells. These changes were significantly reduced in αB-crystallin depleted or knocked out LEC. The removal of the fibre cell mass from the lens of wild-type (WT) mice resulted in the up-regulation of EMT-associated genes in the capsule-adherent epithelial cells, which was reduced in the αB-crystallin KO mice. Together, our data show that αB-crystallin plays a central role in the TGF-β2-induced EMT of LEC. αB-Crystallin could be targeted to prevent PCO and pathological fibrosis in other tissues. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

Gamma crystallins of the human eye lens.

Science.gov (United States)

Vendra, Venkata Pulla Rao; Khan, Ismail; Chandani, Sushil; Muniyandi, Anbukkarasi; Balasubramanian, Dorairajan

2016-01-01

Protein crystallins co me in three types (α, β and γ) and are found predominantly in the eye, and particularly in the lens, where they are packed into a compact, plastic, elastic, and transparent globule of proper refractive power range that aids in focusing incoming light on to the retina. Of these, the γ-crystallins are found largely in the nuclear region of the lens at very high concentrations (>400 mg/ml). The connection between their structure and inter-molecular interactions and lens transparency is an issue of particular interest. We review the origin and phylogeny of the gamma crystallins, their special structure involving the use of Greek key supersecondary structural motif, and how they aid in offering the appropriate refractive index gradient, intermolecular short range attractive interactions (aiding in packing them into a transparent ball), the role that several of the constituent amino acid residues play in this process, the thermodynamic and kinetic stability and how even single point mutations can upset this delicate balance and lead to intermolecular aggregation, forming light-scattering particles which compromise transparency. We cite several examples of this, and illustrate this by cloning, expressing, isolating and comparing the properties of the mutant protein S39C of human γS-crystallin (associated with congenital cataract-microcornea), with those of the wild type molecule. In addition, we note that human γ-crystallins are also present in other parts of the eye (e.g., retina), where their functions are yet to be understood. There are several 'crucial' residues in and around the Greek key motifs which are essential to maintain the compact architecture of the crystallin molecules. We find that a mutation that replaces even one of these residues can lead to reduction in solubility, formation of light-scattering particles and loss of transparency in the molecular assembly. Such a molecular understanding of the process helps us construct the

Crystalline amino acids and nitrogen emission

NARCIS (Netherlands)

Verstegen, M.W.A.; Jongbloed, A.W.

2003-01-01

Reductions in dietary protein level and supplementation with certain crystalline amino acids is a well-established method of formulating diets to achieve a more ideal amino acid pattern and to reduce nitrogen excretion. Up to 35% reduction in nitrogen excretion may be achieved by supplementing pig

Inclusion bodies as potential vehicles for recombinant protein delivery into epithelial cells

Science.gov (United States)

2012-01-01

Background We present the potential of inclusion bodies (IBs) as a protein delivery method for polymeric filamentous proteins. We used as cell factory a strain of E. coli, a conventional host organism, and keratin 14 (K14) as an example of a complex protein. Keratins build the intermediate filament cytoskeleton of all epithelial cells. In order to build filaments, monomeric K14 needs first to dimerize with its binding partner (keratin 5, K5), which is then followed by heterodimer assembly into filaments. Results K14 IBs were electroporated into SW13 cells grown in culture together with a “reporter” plasmid containing EYFP labeled keratin 5 (K5) cDNA. As SW13 cells do not normally express keratins, and keratin filaments are built exclusively of keratin heterodimers (i.e. K5/K14), the short filamentous structures we obtained in this study can only be the result of: a) if both IBs and plasmid DNA are transfected simultaneously into the cell(s); b) once inside the cells, K14 protein is being released from IBs; c) released K14 is functional, able to form heterodimers with EYFP-K5. Conclusions Soluble IBs may be also developed for complex cytoskeletal proteins and used as nanoparticles for their delivery into epithelial cells. PMID:22624805

Activation of an immune-regulatory macrophage response and inhibition of lung inflammation in a mouse model of COPD using heat-shock protein alpha B-crystallin-loaded PLGA microparticles

NARCIS (Netherlands)

van Noort, J.M.; Bsibsi, M.; Nacken, P.J.; Gerritsen, W.H.; Amor, S.; Holtman, I.R.; Boddeke, E.; van Ark, I.; Leusink-Muis, T.; Folkerts, G.; Hennink, W.E.; Amidi, M.

2013-01-01

As an extracellular protein, the small heat-shock protein alpha B-crystallin (HSPB5) has anti-inflammatory effects in several mouse models of inflammation. Here, we show that these effects are associated with the ability of HSPB5 to activate an immune-regulatory response in macrophages via

Activation of an immune-regulatory macrophage response and inhibition of lung inflammation in a mouse model of COPD using heat-shock protein alpha B-crystallin-loaded PLGA microparticles

NARCIS (Netherlands)

van Noort, Johannes M.; Bsibsi, Malika; Nacken, Peter J.; Gerritsen, Wouter H.; Amor, Sandra; Holtman, Inge R.; Boddeke, Erik; van Ark, Ingrid; Leusink-Muis, Thea; Folkerts, Gert; Hennink, Wim E.; Amidi, Maryam

As an extracellular protein, the small heat-shock protein alpha B-crystallin (HSPB5) has anti-inflammatory effects in several mouse models of inflammation. Here, we show that these effects are associated with the ability of HSPB5 to activate an immune-regulatory response in macrophages via

Study of α-crystallin structure by small-angle neutron scattering with contrast variation

International Nuclear Information System (INIS)

Krivandin, A.V.; Muranov, K.O.; Polyanskij, N.B.; Ostrovskij, M.A.; Murugova, T.N.; Kuklin, A.I.; Aksenov, V.L.

2010-01-01

The structure of the oligomeric protein α-crystallin from the bovine eye lens has been investigated by small-angle neutron scattering (SANS) by the contrast variation method (volume fraction of D 2 O was 0, 23, 68 and 90%). Experiments were carried out on YuMO spectrometer (IBR-2 reactor, JINR). From the SANS curves the match point for α-crystallin (43% D 2 O) and its average scattering length density at this point (2.4·10 10 cm -2 ) have been obtained. The radius of gyration and distance distribution functions for α-crystallin have been calculated as well. On the basis of these calculations it was concluded that α-crystallin has a homogeneous distribution of the scattering density in domains inaccessible for water penetration and all parts of this protein undergo a uniform deuteration. The latter indicates that all α-crystallin subunits have an equal accessibility for water and presumably for some other low molecular weight substances. These conclusions on the α-crystallin structure (a homogeneous distribution of the scattering density and an equal accessibility of all subunits for low molecular weight substances) should be taken into account in the time of elaboration of α-crystallin quaternary structure models

Chlamydia trachomatis inclusion membrane protein CT850 interacts with the dynein light chain DYNLT1 (Tctex1).

Science.gov (United States)

Mital, Jeffrey; Lutter, Erika I; Barger, Alexandra C; Dooley, Cheryl A; Hackstadt, Ted

2015-06-26

Chlamydia trachomatis actively subverts the minus-end directed microtubule motor, dynein, to traffic along microtubule tracks to the Microtubule Organizing Center (MTOC) where it remains within a membrane bound replicative vacuole for the duration of its intracellular development. Unlike most substrates of the dynein motor, disruption of the dynactin cargo-linking complex by over-expression of the p50 dynamitin subunit does not inhibit C. trachomatis transport. A requirement for chlamydial protein synthesis to initiate this process suggests that a chlamydial product supersedes a requirement for p50 dynamitin. A yeast 2-hybrid system was used to screen the chlamydia inclusion membrane protein CT850 against a HeLa cell cDNA library and identified an interaction with the dynein light chain DYNLT1 (Tctex1). This interaction was at least partially dependent upon an (R/K-R/K-X-X-R/K) motif that is characteristic of DYNLT1 binding domains. CT850 expressed ectopically in HeLa cells localized at the MTOC and this localization is similarly dependent upon the predicted DYNLT1 binding domain. Furthermore, DYNLT1 is enriched at focal concentrations of CT850 on the chlamydial inclusion membrane that are known to interact with dynein and microtubules. Depletion of DYNLT1 disrupts the characteristic association of the inclusion membrane with centrosomes. Collectively, the results suggest that CT850 interacts with DYNLT1 to promote appropriate positioning of the inclusion at the MTOC. Published by Elsevier Inc.

Interactive domains in the molecular chaperone human alphaB crystallin modulate microtubule assembly and disassembly.

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Joy G Ghosh

2007-06-01

Full Text Available Small heat shock proteins regulate microtubule assembly during cell proliferation and in response to stress through interactions that are poorly understood.Novel functions for five interactive sequences in the small heat shock protein and molecular chaperone, human alphaB crystallin, were investigated in the assembly/disassembly of microtubules and aggregation of tubulin using synthetic peptides and mutants of human alphaB crystallin.The interactive sequence (113FISREFHR(120 exposed on the surface of alphaB crystallin decreased microtubule assembly by approximately 45%. In contrast, the interactive sequences, (131LTITSSLSSDGV(142 and (156ERTIPITRE(164, corresponding to the beta8 strand and the C-terminal extension respectively, which are involved in complex formation, increased microtubule assembly by approximately 34-45%. The alphaB crystallin peptides, (113FISREFHR(120 and (156ERTIPITRE(164, inhibited microtubule disassembly by approximately 26-36%, and the peptides (113FISREFHR(120 and (131LTITSSLSSDGV(142 decreased the thermal aggregation of tubulin by approximately 42-44%. The (131LTITSSLSSDGV(142 and (156ERTIPITRE(164 peptides were more effective than the widely used anti-cancer drug, Paclitaxel, in modulating tubulinmicrotubule dynamics. Mutagenesis of these interactive sequences in wt human alphaB crystallin confirmed the effects of the alphaB crystallin peptides on microtubule assembly/disassembly and tubulin aggregation. The regulation of microtubule assembly by alphaB crystallin varied over a narrow range of concentrations. The assembly of microtubules was maximal at alphaB crystallin to tubulin molar ratios between 1:4 and 2:1, while molar ratios >2:1 inhibited microtubule assembly.Interactive sequences on the surface of human alphaB crystallin collectively modulate microtubule assembly through a dynamic subunit exchange mechanism that depends on the concentration and ratio of alphaB crystallin to tubulin. These are the first

The thermal structural transition of alpha-crystallin modulates subunit interactions and increases protein solubility.

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Giuseppe Maulucci

Full Text Available BACKGROUND: Alpha crystallin is an oligomer composed of two types of subunits, alpha-A and alpha-B crystallin, and is the major constituent of human lens. The temperature induced condensation of alpha-crystallin, the main cause for eye lens opacification (cataract, is a two step-process, a nucleation followed by an aggregation phase, and a protective effect towards the aggregation is exhibited over the alpha crystallin phase transition temperature (Tc = 318.16 K. METHODS/RESULTS: To investigate if a modulation of the subunit interactions over Tc could trigger the protective mechanism towards the aggregation, we followed, by using simultaneously static and dynamic light scattering, the temperature induced condensation of alpha-crystallin. By developing a mathematical model able to uncouple the nucleation and aggregation processes, we find a previously unobserved transition in the nucleation rate constant. Its temperature dependence allows to determine fundamental structural parameters, the chemical potential (Δμ and the interfacial tension (γ of the aggregating phase, that characterize subunit interactions. CONCLUSIONS/GENERAL SIGNIFICANCE: The decrease of both Δμ and γ at Tc, and a relative increase in solubility, reveal a significative decrease in the strenght of alpha-crystallin subunits interactions, which protects from supramolecolar condensation in hypertermic conditions. On the whole, we suggest a general approach able to understand the structural and kinetic mechanisms involved in aggregation-related diseases and in drugs development and testing.

A simple strategy for the purification of native recombinant full-length human RPL10 protein from inclusion bodies.

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Pereira, Larissa M; Silva, Luana R; Alves, Joseane F; Marin, Nélida; Silva, Flavio Sousa; Morganti, Ligia; Silva, Ismael D C G; Affonso, Regina

2014-09-01

The L10 ribosomal protein (RPL10) plays a role in the binding of the 60 S and 40 S ribosomal subunits and in mRNA translation. The evidence indicates that RPL10 also has multiple extra-ribosomal functions, including tumor suppression. Recently, the presence of RPL10 in prostate and ovarian cancers was evaluated, and it was demonstrated to be associated with autistic disorders and premature ovarian failure. In the present work, we successfully cloned and expressed full-length human RPL10 (hRPL10) protein and isolated inclusion bodies containing this protein that had formed under mild growth conditions. The culture produced 376mg of hRPL10 protein per liter of induced bacterial culture, of which 102.4mg was present in the soluble fraction, and 25.6mg was recovered at approximately 94% purity. These results were obtained using a two-step process of non-denaturing protein extraction from pelleted inclusion bodies. We studied the characteristics of this protein using circular dichroism spectroscopy and by monitoring the changes induced by the presence or absence of zinc ions using fluorescence spectrometry. The results demonstrated that the protein obtained using these non-conventional methods retained its secondary and tertiary structure. The conformational changes induced by the incorporation of zinc suggested that this protein could interact with Jun or the SH3 domain of c-yes. The results suggested that the strategy used to obtain hRPL10 is simple and could be applied to obtaining other proteins that are susceptible to degradation. Copyright © 2014 Elsevier Inc. All rights reserved.

Electrophoretic variation in low molecular weight lens crystallins from inbred strains of rats.

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Donner, M E; Skow, L C; Kunz, H W; Gill, T J

1985-10-01

Analysis of rat lens soluble proteins by analytical isoelectric focusing detected two inherited electrophoretic differences in low molecular weight (LM) crystallins from inbred strains of rats (Rattus norvegicus). The polymorphic lens crystallins were shown to be similar to a genetically variant LM crystallin, LEN-1, previously described in mice (Mus musculus) and encoded on chromosome 1, at a locus linked to Pep-3 (dipeptidase). Linkage analysis demonstrated that the rat crystallin locus was loosely linked to Pep-3 at a recombination distance of 38 +/- 4.5 U. These data suggest the conservation of a large chromosomal region during the evolution of Rodentia and support the hypothesis that the gamma-crystallins are evolving more rapidly than alpha- or beta-crystallins.

Towards revealing the structure of bacterial inclusion bodies.

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Wang, Lei

2009-01-01

Protein aggregation is a widely observed phenomenon in human diseases, biopharmaceutical production, and biological research. Protein aggregates are generally classified as highly ordered, such as amyloid fibrils, or amorphous, such as bacterial inclusion bodies. Amyloid fibrils are elongated filaments with diameters of 6-12 nm, they are comprised of residue-specific cross-beta structure, and display characteristic properties, such as binding with amyloid-specific dyes. Amyloid fibrils are associated with dozens of human pathological conditions, including Alzheimer disease and prion diseases. Distinguished from amyloid fibrils, bacterial inclusion bodies display apparent amorphous morphology. Inclusion bodies are formed during high-level recombinant protein production, and formation of inclusion bodies is a major concern in biotechnology. Despite of the distinctive morphological difference, bacterial inclusion bodies have been found to have some amyloid-like properties, suggesting that they might contain structures similar to amyloid-like fibrils. Recent structural data further support this hypothesis, and this review summarizes the latest progress towards revealing the structural details of bacterial inclusion bodies.

Apatite nano-crystalline surface modification of poly(lactide-co-glycolide) sintered microsphere scaffolds for bone tissue engineering: implications for protein adsorption.

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Jabbarzadeh, Ehsan; Nair, Lakshmi S; Khan, Yusuf M; Deng, Meng; Laurencin, Cato T

2007-01-01

A number of bone tissue engineering approaches are aimed at (i) increasing the osteconductivity and osteoinductivity of matrices, and (ii) incorporating bioactive molecules within the scaffolds. In this study we examined the growth of a nano-crystalline mineral layer on poly(lactide-co-glycolide) (PLAGA) sintered microsphere scaffolds for tissue engineering. In addition, the influence of the mineral precipitate layer on protein adsorption on the scaffolds was studied. Scaffolds were mineralized by incubation in simulated body fluid (SBF). Scanning electron microscopy (SEM) analysis revealed that mineralized scaffolds possess a rough surface with a plate-like nanostructure covering the surface of microspheres. The results of protein adsorption and release studies showed that while the protein release pattern was similar for PLAGA and mineralized PLAGA scaffolds, precipitation of the mineral layer on PLAGA led to enhanced protein adsorption and slower protein release. Mineralization of tissue-engineered surfaces provides a method for both imparting bioactivity and controlling levels of protein adsorption and release.

Crystalline and Crystalline International Disposal Activities

Energy Technology Data Exchange (ETDEWEB)

Viswanathan, Hari S. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Chu, Shaoping [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Dittrich, Timothy M. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Hyman, Jeffrey De' Haven [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Karra, Satish [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Makedonska, Nataliia [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Reimus, Paul William [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

2017-03-06

This report presents the results of work conducted between September 2015 and July 2016 at Los Alamos National Laboratory in the crystalline disposal and crystalline international disposal work packages of the Used Fuel Disposition Campaign (UFDC) for DOE-NE’s Fuel Cycle Research and Development program. Los Alamos focused on two main activities during this period: Discrete fracture network (DFN) modeling to describe flow and radionuclide transport in complex fracture networks that are typical of crystalline rock environments, and a comprehensive interpretation of three different colloid-facilitated radionuclide transport experiments conducted in a fractured granodiorite at the Grimsel Test Site in Switzerland between 2002 and 2013. Chapter 1 presents the results of the DFN work and is divided into three main sections: (1) we show results of our recent study on the correlation between fracture size and fracture transmissivity (2) we present an analysis and visualization prototype using the concept of a flow topology graph for characterization of discrete fracture networks, and (3) we describe the Crystalline International work in support of the Swedish Task Force. Chapter 2 presents interpretation of the colloidfacilitated radionuclide transport experiments in the crystalline rock at the Grimsel Test Site.

Role of alpha-crystallin, early-secreted antigenic target 6-kDa protein and culture filtrate protein 10 as novel diagnostic markers in osteoarticular tuberculosis

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Nazia Rizvi

2016-07-01

Full Text Available Osteoarticular tuberculosis constitutes about 3% of all tuberculosis cases. Early and accurate diagnosis of tuberculosis is a challenging problem especially in the case of osteoarticular tuberculosis owing to the lower number of bacilli. However, an accurate and timely diagnosis of the disease results in an improved efficacy of the given treatment. Besides the limitations of conventional methods, nowadays molecular diagnostic techniques have emerged as a major breakthrough for the early diagnosis of tuberculosis with high sensitivity and specificity. Alpha-crystallin is a dominantly expressed protein responsible for the long viability of the pathogen during the latent phase under certain stress conditions such as hypoxia and nitric oxide stress. Two other proteins—early secreted antigenic target-6 and culture filtrate protein-10—show high expression in the active infective phase of Mycobacterium tuberculosis. In this article, we focus on the different proteins expressed dominantly in latent/active tuberculosis, and which may be further used as prognostic biomarkers for diagnosing tuberculosis, both in latent and active phases.

The P2 of Wheat yellow mosaic virus rearranges the endoplasmic reticulum and recruits other viral proteins into replication-associated inclusion bodies.

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Sun, Liying; Andika, Ida Bagus; Shen, Jiangfeng; Yang, Di; Chen, Jianping

2014-06-01

Viruses commonly modify host endomembranes to facilitate biological processes in the viral life cycle. Infection by viruses belonging to the genus Bymovirus (family Potyviridae) has long been known to induce the formation of large membranous inclusion bodies in host cells, but their assembly and biological roles are still unclear. Immunoelectron microscopy of cells infected with the bymovirus Wheat yellow mosaic virus (WYMV) showed that P1, P2 and P3 are the major viral protein constituents of the membranous inclusions, whereas NIa-Pro (nuclear inclusion-a protease) and VPg (viral protein genome-linked) are probable minor components. P1, P2 and P3 associated with the endoplasmic reticulum (ER), but only P2 was able to rearrange ER and form large aggregate structures. Bioinformatic analyses and chemical experiments showed that P2 is an integral membrane protein and depends on the active secretory pathway to form aggregates of ER membranes. In planta and in vitro assays demonstrated that P2 interacts with P1, P3, NIa-Pro or VPg and recruits these proteins into the aggregates. In vivo RNA labelling using WYMV-infected wheat protoplasts showed that the synthesis of viral RNAs occurs in the P2-associated inclusions. Our results suggest that P2 plays a major role in the formation of membranous compartments that house the genomic replication of WYMV. © 2013 BSPP AND JOHN WILEY & SONS LTD.

Alpha B- and βA3-crystallins containing d-aspartic acids exist in a monomeric state.

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Sakaue, Hiroaki; Takata, Takumi; Fujii, Norihiko; Sasaki, Hiroshi; Fujii, Noriko

2015-01-01

Crystallin stability and subunit-subunit interaction are essential for eye lens transparency. There are three types of crystallins in lens, designated as α-, β-, and γ-crystallins. Alpha-crystallin is a hetero-polymer of about 800kDa, consisting of 35-40 subunits of two different αA- and αB-subunits, each of 20kDa. The β/γ-crystallin superfamily comprises oligomeric β-crystallin (2-6 subunits) and monomeric γ-crystallin. Since lens proteins have very long half-lives, they undergo numerous post-translational modifications including racemization, isomerization, deamidation, oxidation, glycation, and truncation, which may decrease crystallin solubility and ultimately cause cataract formation. Racemization and isomerization of aspartyl (Asp) residues have been detected only in polymeric α- and oligomeric β-crystallin, while the situation in monomeric γ-crystallin has not been studied. Here, we investigated the racemization and isomerization of Asp in the γ-crystallin fraction of elderly donors. The results show that Asp residues of γS-, γD- and γC-crystallins were not racemized and isomerized. However, strikingly, we found that a portion of αB-crystallin and βA3-crystallin moved to the lower molecular weight fraction which is the same size of γ-crystallin. In those fractions, Asp-96 of αB-crystallin and Asp-37 of βA3-crystallin were highly inverted, which do not occur in the native lens higher molecular weight fraction. Our results indicate the possibility that the inversion of Asp residues may induce dissociation of αB- and βA3-crystallins from the polymeric and oligomeric states. This is the first report that stereoinversion of amino acids disturbs lens protein assembly in aged human lens. Copyright © 2014 Elsevier B.V. All rights reserved.

Concentration dependence of transmission losses in UV-laser irradiated bovine α-, βH-, βL- and γ-crystallin solutions

International Nuclear Information System (INIS)

Hott, J.L.; Borkman, R.F.

1993-01-01

Experiments with calf lens protein fractions in aqueous buffer solutions at room temperature showed that β H -, β L - and γ-crystallin fractions became opaque following ultraviolet exposure at 308 nm, while the α-crystallin fraction remained transparent. Transmission loss, due to UV-irradiation, for all of the crystallin samples was studied in the concentration range of 0.1 mg/mL to 1.0 mg/mL, and for α- and γ-crystallin, in the range up to 5 mg/mL. With increased concentrations of β H -, β L -and γ-crystalline, the rate of opacification increased. However, with α-crystallin, the loss of transmission was negligible for all of the concentrations and irradiation times studied. Opacification of the crystallins was accompanied by formation of higher molecular weight insoluble proteins as detected by SDS-PAGE. (Author)

Crystalline and Crystalline International Disposal Activities

Energy Technology Data Exchange (ETDEWEB)

Viswanathan, Hari S. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Chu, Shaoping [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Reimus, Paul William [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Makedonska, Nataliia [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Hyman, Jeffrey De' Haven [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Karra, Satish [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Dittrich, Timothy M. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

2015-12-21

This report presents the results of work conducted between September 2014 and July 2015 at Los Alamos National Laboratory in the crystalline disposal and crystalline international disposal work packages of the Used Fuel Disposition Campaign (UFDC) for DOE-NE’s Fuel Cycle Research and Development program.

Fluid inclusion investigations in Nagra's boreholes of Northern Switzerland

International Nuclear Information System (INIS)

Mullis, J.

1987-01-01

Fluid inclusions studied in quartz and calcite from 6 Nagra-boreholes and the oil exploration well Pfaffnau in northern Switzerland were used to evaluate the evolution of fluid composition and trapping conditions. The studied boreholes covered representative sections of the Tertiary, Mesozoic and Permo-Carboniferous sedimentary rocks in addition to the underlying crystalline basement. Two fluid groups were identified, one containing salt-poor NaCl-(KCl) fluids of Upper Carboniferous age, and a second group containing salt-rich to salt-poor CaCl 2 -NaCl-(MgCl 2 ) fluids trapped between Permian and Tertiary times. The inclusions of the first group were trapped at relatively high temperatures (100 degrees to 350 degrees C, rarely up to 400 degrees C) and low pressures (probably below 1 kbar). Their widespread occurrence along healed fractures in rock forming minerals of the crystalline basement and in detrital quartz grains of the Permo-Carboniferous and Triassic sandstones imply a tectonic/thermic event on a regional scale during late Variscan orogeny. The second group of fluids comprises early salt-rich and late salt-poor inclusions. The inclusions were trapped between 30 degrees and 140 degrees C and are detected preferentially in fissure minerals and porefilling cements. The widespread occurrence of the salt-poor fluids along healed fractures inside the host minerals imply major tectonic events especially during Tertiary times. These tectonic events are probably responsible for increased fluid migration and possibly also for brine dilution. The two fluid groups display a considerable similarity with the high-temperature/low-saliniferous Variscan fluid system of the Black Forest and Oberpfalz. (author) 44 refs., 5 figs., 2 tabs

cAMP-dependent phosphorylation of bovine lens alpha-crystallin

International Nuclear Information System (INIS)

Spector, A.; Chiesa, R.; Sredy, J.; Garner, W.

1985-01-01

This communication reports that the A1 and B1 chains of bovine lens alpha-crystallin are phosphorylated. The conclusion is based on the following evidence: (i) When soluble preparations from lens cortex are incubated with [gamma- 32 P]ATP, a cAMP-dependent labeling of a high molecular weight protein is obtained. (ii) After NaDodSO 4 /PAGE, the label is found in two bands with Mr 22,000 and 20,000, corresponding to the B and A chains of alpha-crystallin, respectively. (iii) Isoelectric focusing indicates that the radioactivity is almost exclusively in bands with pI values of 5.58 and 6.70, corresponding to the A1 and B1 chains, respectively. (iv) Similar results are obtained in experiments of [ 32 P]orthophosphate incorporation in lens organ culture. (v) Analyses of the digested protein indicate the label is exclusively in phosphoserine. (vi) 31 P NMR analyses of native, proteolytically digested, and urea-treated alpha-crystallin gives a chemical shift of 4.6 ppm relative to 85% H 3 PO 4 at pH 7.4, suggesting that the phosphate is covalently bound to a serine in the protein. An abundance of approximately one phosphate per four or five monomer units was found. (vii) Similar results were obtained by chemical analyses of independently prepared alpha-crystallin samples. The results are consistent with the view that the A1 and B1 chains arise as result of the phosphorylation of directly synthesized A2 and B2 polypeptides. It is suggested that this metabolically controlled phosphorylation may be associated with the terminal differentiation of the lens epithelial cell and the intracellular organization of the lens fiber cell

A Transition Metal-Binding, Trimeric βγ-Crystallin from Methane-Producing Thermophilic Archaea, Methanosaeta thermophila.

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Srivastava, Shanti Swaroop; Jamkhindikar, Aditya Anand; Raman, Rajeev; Jobby, Maroor K; Chadalawada, Swathi; Sankaranarayanan, Rajan; Sharma, Yogendra

2017-03-07

βγ-Crystallins are important constituents of the vertebrate eye lens, whereas in microbes, they are prevalent as Ca 2+ -binding proteins. In archaea, βγ-crystallins are conspicuously confined to two methanogens, viz., Methanosaeta and Methanosarcina. One of these, i.e., M-crystallin from Methanosarcina acetivorans, has been shown to be a typical Ca 2+ -binding βγ-crystallin. Here, with the aid of a high-resolution crystal structure and isothermal titration calorimetry, we report that "Methallin", a βγ-crystallin from Methanosaeta thermophila, is a trimeric, transition metal-binding protein. It binds Fe, Ni, Co, or Zn ion with nanomolar affinity, which is consistent even at 55 °C, the optimal temperature for the methanogen's growth. At the center of the protein trimer, the metal ion is coordinated by six histidines, two from each protomer, leading to an octahedral geometry. Small-angle X-ray scattering analysis confirms that the trimer seen in the crystal lattice is a biological assembly; this assembly dissociates to monomers upon removal of the metal ion. The introduction of two histidines (S17H/S19H) into a homologous βγ-crystallin, Clostrillin, allows it to bind nickel at the introduced site, though with micromolar affinity. However, because of the lack of a compatible interface, nickel binding could not induce trimerization, affirming that Methallin is a naturally occurring trimer for high-affinity transition metal binding. While βγ-crystallins are known to bind Ca 2+ and form homodimers and oligomers, the transition metal-binding, trimeric Methallin is a new paradigm for βγ-crystallins. The distinct features of Methallin, such as nickel or iron binding, are also possible imprints of biogeochemical changes during the period of its origin.

Thermal, crystallinity and morphological studies of the filled RBD palm kernel oil polyurethane foam

International Nuclear Information System (INIS)

Khairiah Badri; Sahrim Ahmad; Sarani Zakaria

2000-01-01

The synthesis of RBD palm kernel oil (PKO) polyurethane polyol and the polyurethane foam has well been documented. However, less study has been put in discovering the thermal properties and crystallinity of the foam. It is also an initiative to investigate the effect of oil palm empty fruit bunch (EFB) and sorbitol as fillers in the polyurethane (PU) foam to these properties. Thermogravimetric (TGA) investigation of the PKO PU foam was performed to study their decompositions. The semi-crystalline nature of EFB-filled PU was confirmed by x-ray diffratogram and DSC thermogram of glass transition temperature, T g . The x-ray diffraction (XRD) study of the unfilled PU showed a broad amorphous halo, indicative of absence of crystallinity in the polymer, which has been explained as due to strong hydrogen bonding in the hard phase. Overall crystallinity decreases with an increase in the polyester content in agreement with the XRD results. The crystallinity however, increases with the inclusion of EFB in the polyurethane system. This study was followed by the observation of the surface morphologies of the PKO PU foam with and without fillers. The scanning electron micrographs verified the finding on the improved k-factor values. (Author)

Integrated continuous processing of proteins expressed as inclusion bodies: GCSF as a case study.

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Kateja, Nikhil; Agarwal, Harshit; Hebbi, Vishwanath; Rathore, Anurag S

2017-07-01

Affordability of biopharmaceuticals continues to be a challenge, particularly in developing economies. This has fuelled advancements in manufacturing that can offer higher productivity and better economics without sacrificing product quality in the form of an integrated continuous manufacturing platform. While platform processes for monoclonal antibodies have existed for more than a decade, development of an integrated continuous manufacturing process for bacterial proteins has received relatively scant attention. In this study, we propose an end-to-end integrated continuous downstream process (from inclusion bodies to unformulated drug substance) for a therapeutic protein expressed in Escherichia coli as inclusion body. The final process consisted of a continuous refolding in a coiled flow inverter reactor directly coupled to a three-column periodic counter-current chromatography for capture of the product followed by a three-column con-current chromatography for polishing. The continuous bioprocessing train was run uninterrupted for 26 h to demonstrate its capability and the resulting output was analyzed for the various critical quality attributes, namely product purity (>99%), high molecular weight impurities (<0.5%), host cell proteins (<100 ppm), and host cell DNA (<10 ppb). All attributes were found to be consistent over the period of operation. The developed assembly offers smaller facility footprint, higher productivity, fewer hold steps, and significantly higher equipment and resin utilization. The complexities of process integration in the context of continuous processing have been highlighted. We hope that the study presented here will promote development of highly efficient, universal, end-to-end, fully continuous platforms for manufacturing of biotherapeutics. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:998-1009, 2017. © 2016 American Institute of Chemical Engineers.

Improved hydroxypropyl methylcellulose (HPMC) films through incorporation of amylose-sodium palmitate inclusion complexes

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Polymer film blends of hydroxypropyl methylcellulose (HPMC) and amylose-sodium palmitate inclusion complexes (Na-Palm) were produced with no plasticizer, and were observed to have improved physical and gas barrier properties as compared with pure HPMC. The crystalline amylose helices incorporating t...

Crosslinking and photoreaction of ozone-oxidized calf-lens alpha-crystallin

International Nuclear Information System (INIS)

Fujimori, E.

1982-01-01

Direct-photo-oxidation, singlet oxygen-oxidation, or photosensitized oxidation can modify lens crystallins, causing an increase in blue fluorescence and covalent crosslinking. A relationship between these changes has not been elucidated. We now report results from experiments with ozone oxidation. When calf-lens alpha-crystallin is treated with zone oxidation. When calf-lens alpha-crystallin is treated with ozone, new absorption, fluorescence, and phosphorescence, which are characteristic of the oxidized product of tryptophan (N-formylkynurenine), appear at 320, 435, and 445 nm, respectively. In addition, in this ozonization of alpha-crystallin, its polypeptides are crosslinked by nondisulfide bonds. Irradiation of ozone-treated alpha-crystallin with near-ultraviolet (365 nm) light increases crosslinking and reduces the 320 nm absorbance with a concomitant appearance of a new absorption at about 420 nm. This photoproduct exhibits an intense fluorescence around 450 nm and a weak phosphorescence at 510 nm, with excitation peaks at 400, 415, and 422 nm. These findings are essentially the same as those observed in photo-oxidized alpha-crystallin, suggesting the involvement of the same tryptophan oxidized product in the modification of the lens protein

BAG3 directly interacts with mutated alphaB-crystallin to suppress its aggregation and toxicity.

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Akinori Hishiya

Full Text Available A homozygous disruption or genetic mutation of the bag3 gene causes progressive myofibrillar myopathy in mouse and human skeletal and cardiac muscle disorder while mutations in the small heat shock protein αB-crystallin gene (CRYAB are reported to be responsible for myofibrillar myopathy. Here, we demonstrate that BAG3 directly binds to wild-type αB-crystallin and the αB-crystallin mutant R120G, via the intermediate domain of BAG3. Peptides that inhibit this interaction in an in vitro binding assay indicate that two conserved Ile-Pro-Val regions of BAG3 are involved in the interaction with αB-crystallin, which is similar to results showing BAG3 binding to HspB8 and HspB6. BAG3 overexpression increased αB-crystallin R120G solubility and inhibited its intracellular aggregation in HEK293 cells. BAG3 suppressed cell death induced by αB-crystallin R120G overexpression in differentiating C2C12 mouse myoblast cells. Our findings indicate a novel function for BAG3 in inhibiting protein aggregation caused by the genetic mutation of CRYAB responsible for human myofibrillar myopathy.

BAG3 Directly Interacts with Mutated alphaB-Crystallin to Suppress Its Aggregation and Toxicity

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Hishiya, Akinori; Salman, Mortada Najem; Carra, Serena; Kampinga, Harm H.; Takayama, Shinichi

2011-01-01

A homozygous disruption or genetic mutation of the bag3 gene causes progressive myofibrillar myopathy in mouse and human skeletal and cardiac muscle disorder while mutations in the small heat shock protein αB-crystallin gene (CRYAB) are reported to be responsible for myofibrillar myopathy. Here, we demonstrate that BAG3 directly binds to wild-type αB-crystallin and the αB-crystallin mutant R120G, via the intermediate domain of BAG3. Peptides that inhibit this interaction in an in vitro binding assay indicate that two conserved Ile-Pro-Val regions of BAG3 are involved in the interaction with αB-crystallin, which is similar to results showing BAG3 binding to HspB8 and HspB6. BAG3 overexpression increased αB-crystallin R120G solubility and inhibited its intracellular aggregation in HEK293 cells. BAG3 suppressed cell death induced by αB-crystallin R120G overexpression in differentiating C2C12 mouse myoblast cells. Our findings indicate a novel function for BAG3 in inhibiting protein aggregation caused by the genetic mutation of CRYAB responsible for human myofibrillar myopathy. PMID:21423662

Capillary gel electrophoresis for the quantification and purity determination of recombinant proteins in inclusion bodies.

Science.gov (United States)

Espinosa-de la Garza, Carlos E; Perdomo-Abúndez, Francisco C; Campos-García, Víctor R; Pérez, Néstor O; Flores-Ortiz, Luis F; Medina-Rivero, Emilio

2013-09-01

In this work, a high-resolution CGE method for quantification and purity determination of recombinant proteins was developed, involving a single-component inclusion bodies (IBs) solubilization solution. Different recombinant proteins expressed as IBs were used to show method capabilities, using recombinant interferon-β 1b as the model protein for method validation. Method linearity was verified in the range from 0.05 to 0.40 mg/mL and a determination coefficient (r(2) ) of 0.99 was obtained. The LOQs and LODs were 0.018 and 0.006 mg/mL, respectively. RSD for protein content repeatability test was 2.29%. In addition, RSD for protein purity repeatability test was 4.24%. Method accuracy was higher than 90%. Specificity was confirmed, as the method was able to separate recombinant interferon-β 1b monomer from other aggregates and impurities. Sample content and purity was demonstrated to be stable for up to 48 h. Overall, this method is suitable for the analysis of recombinant proteins in IBs according to the attributes established on the International Conference for Harmonization guidelines. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Crystalline Silica Primer

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,

1992-01-01

Crystalline silica is the scientific name for a group of minerals composed of silicon and oxygen. The term crystalline refers to the fact that the oxygen and silicon atoms are arranged in a threedimensional repeating pattern. This group of minerals has shaped human history since the beginning of civilization. From the sand used for making glass to the piezoelectric quartz crystals used in advanced communication systems, crystalline silica has been a part of our technological development. Crystalline silica's pervasiveness in our technology is matched only by its abundance in nature. It's found in samples from every geologic era and from every location around the globe. Scientists have known for decades that prolonged and excessive exposure to crystalline silica dust in mining environments can cause silicosis, a noncancerous lung disease. During the 1980's, studies were conducted that suggested that crystalline silica also was a carcinogen. As a result of these findings, crystalline silica has been regulated under the Occupational Safety and Health Administration's (OSHA) Hazard Communication Standard (HCS). Under HCS, OSHAregulated businesses that use materials containing 0.1% or more crystalline silica must follow Federal guidelines concerning hazard communication and worker training. Although the HCS does not require that samples be analyzed for crystalline silica, mineral suppliers or OSHAregulated

Dating the time of birth: A radiocarbon calibration curve for human eye-lens crystallines

DEFF Research Database (Denmark)

Kjeldsen, Henrik; Heinemeier, Jan; Heegaard, Steffen

2010-01-01

Radiocarbon bomb-pulse dating has been used to measure the formation age of human eye-lens crystallines. Lens crystallines are special proteins in the eye-lens that consist of virtually inert tissue. The experimental data show that the radiocarbon ages to a large extent reflect the time of birth...

Pointlike Inclusion Interactions in Tubular Membranes

NARCIS (Netherlands)

Vahid Belarghou, A.; Idema, T.

2016-01-01

Membrane tubes and tubular networks are ubiquitous in living cells. Inclusions like proteins are vital for both the stability and the dynamics of such networks. These inclusions interact via the curvature deformations they impose on the membrane. We analytically study the resulting membrane

Application of preparative disk gel electrophoresis for antigen purification from inclusion bodies.

Science.gov (United States)

Okegawa, Yuki; Koshino, Masanori; Okushima, Teruya; Motohashi, Ken

2016-02-01

Specific antibodies are a reliable tool to examine protein expression patterns and to determine the protein localizations within cells. Generally, recombinant proteins are used as antigens for specific antibody production. However, recombinant proteins from mammals and plants are often overexpressed as insoluble inclusion bodies in Escherichia coli. Solubilization of these inclusion bodies is desirable because soluble antigens are more suitable for injection into animals to be immunized. Furthermore, highly purified proteins are also required for specific antibody production. Plastidic acetyl-CoA carboxylase (ACCase: EC 6.4.1.2) from Arabidopsis thaliana, which catalyzes the formation of malonyl-CoA from acetyl-CoA in chloroplasts, formed inclusion bodies when the recombinant protein was overexpressed in E. coli. To obtain the purified protein to use as an antigen, we applied preparative disk gel electrophoresis for protein purification from inclusion bodies. This method is suitable for antigen preparation from inclusion bodies because the purified protein is recovered as a soluble fraction in electrode running buffer containing 0.1% sodium dodecyl sulfate that can be directly injected into immune animals, and it can be used for large-scale antigen preparation (several tens of milligrams). Copyright © 2015 Elsevier Inc. All rights reserved.

Dating the time of birth: A radiocarbon calibration curve for human eye-lens crystallines

International Nuclear Information System (INIS)

Kjeldsen, Henrik; Heinemeier, Jan; Heegaard, Steffen; Jacobsen, Christina; Lynnerup, Niels

2010-01-01

Radiocarbon bomb-pulse dating has been used to measure the formation age of human eye-lens crystallines. Lens crystallines are special proteins in the eye-lens that consist of virtually inert tissue. The experimental data show that the radiocarbon ages to a large extent reflect the time of birth, in accordance with expectations. Moreover, it has been possible to develop an age model for the formation of the eye-lens crystallines. From this model a radiocarbon calibration curve for lens crystallines has been calculated. As a consequence, the time of birth of humans can be determined with an accuracy of a few years by radiocarbon dating.

Dating the time of birth: A radiocarbon calibration curve for human eye-lens crystallines

Energy Technology Data Exchange (ETDEWEB)

Kjeldsen, Henrik, E-mail: kjeldsen@phys.au.d [AMS 14C Dating Centre, Department of Physics and Astronomy, University of Aarhus, Aarhus (Denmark); Heinemeier, Jan [AMS 14C Dating Centre, Department of Physics and Astronomy, University of Aarhus, Aarhus (Denmark); Heegaard, Steffen [Eye Pathology Section, Department of Neuroscience and Pharmacology, University of Copenhagen, Copenhagen (Denmark); Jacobsen, Christina; Lynnerup, Niels [Department of Forensic Medicine, University of Copenhagen, Copenhagen (Denmark)

2010-04-15

Radiocarbon bomb-pulse dating has been used to measure the formation age of human eye-lens crystallines. Lens crystallines are special proteins in the eye-lens that consist of virtually inert tissue. The experimental data show that the radiocarbon ages to a large extent reflect the time of birth, in accordance with expectations. Moreover, it has been possible to develop an age model for the formation of the eye-lens crystallines. From this model a radiocarbon calibration curve for lens crystallines has been calculated. As a consequence, the time of birth of humans can be determined with an accuracy of a few years by radiocarbon dating.

Immunohistochemical identification of messenger RNA-related proteins in basophilic inclusions of adult-onset atypical motor neuron disease.

Science.gov (United States)

Fujita, Kengo; Ito, Hidefumi; Nakano, Satoshi; Kinoshita, Yoshimi; Wate, Reika; Kusaka, Hirofumi

2008-10-01

This report concerns an immunohistochemical investigation on RNA-related proteins in the basophilic inclusions (BIs) from patients with adult-onset atypical motor neuron disease. Formalin-fixed, paraffin-embedded sections of the motor cortex and the lumbar spinal cord were examined. The BIs appeared blue in color with H&E and Nissl stain, and pink with methylgreen-pyronin stain. Ribonuclease pretreatment abolished the methylgreen-pyronin staining, suggesting that the BIs contained RNA. Immunohistochemically, the BIs were distinctly labeled with the antibodies against poly(A)-binding protein 1, T cell intracellular antigen 1, and ribosomal protein S6. These proteins are essential constituents of stress granules. In contrast, the BIs were not immunoreactive for ribosomal protein L28 and decapping enzyme 1, which are core components of transport ribonucleoprotein particles and processing bodies, respectively. Moreover, the BIs were not immunopositive for TDP-43. Our results imply that translation attenuation could be involved in the processes of BI formation in this disorder.

Annealing studies of Bi and Kr inclusions in Al

Energy Technology Data Exchange (ETDEWEB)

Bjoern Thoft, N

1995-04-01

This report contains the results of experimental investigations of melting, solidification and growth of Bi and Kr inclusions made by ion implantation into aluminium. The experimental techniques used of for this study were x-ray diffraction, transmission electron microscopy, Rutherford backscattering, ion channeling, and grazing-incidence small-angle x-ray scattering. The x-ray diffraction signal from crystalline Bi inclusions in Al has been recorded as a function of temperature during heating to temperatures above the bulk melting point and cooling to room temperature. Data from these measurements have been fitted using models (developed by Pawlow and Wronski) for the size-dependent melting temperature of small particles, and size distributions for the inclusions have been determined in this way. Transmission electron microscopy has confirmed the melting and solidification of the Bi inclusions in the temperature ranges, in which these processes were observed by x-ray diffraction, establishing the facts that the inclusions melt below the bulk melting point and that a large supercooling is seen. Information about the amount and depth distribution of the Bi confined in the Al matrix has been derived from Rutherford backscattering measurements. Melting and solidification of Bi inclusions have been observed by means of ion channeling. The results of the investigations of bismuth inclusions in aluminium are compared to previous, similar results for lead inclusions in aluminium. Finally, preliminary experiments have confirmed that growth of Kr inclusions in Al can be observed using grazing-incidence small-angle scattering. (au) (13 tabs., 46 ills., 77 refs.).

Annealing studies of Bi and Kr inclusions in Al

International Nuclear Information System (INIS)

Bjoern Thoft, N.

1995-04-01

This report contains the results of experimental investigations of melting, solidification and growth of Bi and Kr inclusions made by ion implantation into aluminium. The experimental techniques used of for this study were x-ray diffraction, transmission electron microscopy, Rutherford backscattering, ion channeling, and grazing-incidence small-angle x-ray scattering. The x-ray diffraction signal from crystalline Bi inclusions in Al has been recorded as a function of temperature during heating to temperatures above the bulk melting point and cooling to room temperature. Data from these measurements have been fitted using models (developed by Pawlow and Wronski) for the size-dependent melting temperature of small particles, and size distributions for the inclusions have been determined in this way. Transmission electron microscopy has confirmed the melting and solidification of the Bi inclusions in the temperature ranges, in which these processes were observed by x-ray diffraction, establishing the facts that the inclusions melt below the bulk melting point and that a large supercooling is seen. Information about the amount and depth distribution of the Bi confined in the Al matrix has been derived from Rutherford backscattering measurements. Melting and solidification of Bi inclusions have been observed by means of ion channeling. The results of the investigations of bismuth inclusions in aluminium are compared to previous, similar results for lead inclusions in aluminium. Finally, preliminary experiments have confirmed that growth of Kr inclusions in Al can be observed using grazing-incidence small-angle scattering. (au) (13 tabs., 46 ills., 77 refs.)

Reassessment of inclusion body-based production as a versatile opportunity for difficult-to-express recombinant proteins.

Science.gov (United States)

Hoffmann, Daniel; Ebrahimi, Mehrdad; Gerlach, Doreen; Salzig, Denise; Czermak, Peter

2017-11-10

The production of recombinant proteins in the microbial host Escherichia coli often results in the formation of cytoplasmic protein inclusion bodies (IBs). Proteins forming IBs are often branded as difficult-to-express, neglecting that IBs can be an opportunity for their production. IBs are resistant to proteolytic degradation and contain up to 90% pure recombinant protein, which does not interfere with the host metabolism. This is especially advantageous for host-toxic proteins like antimicrobial peptides (AMPs). IBs can be easily isolated by cell disruption followed by filtration and/or centrifugation, but conventional techniques for the recovery of soluble proteins from IBs are laborious. New approaches therefore simplify protein recovery by optimizing the production process conditions, and often include mild resolubilization methods that either increase the yield after refolding or avoid the necessity of refolding all together. For the AMP production, the IB-based approach is ideal, because these peptides often have simple structures and are easy to refold. The intentional IB production of almost every protein can be achieved by fusing recombinant proteins to pull-down tags. This review discusses the techniques available for IB-based protein production before considering technical approaches for the isolation of IBs from E. coli lysates followed by efficient protein resolubilization which ideally omits further refolding. The techniques are evaluated in terms of their suitability for the process-scale production and downstream processing of recombinant proteins and are discussed for AMP production as an example.

Multiple origin of diurnality in geckos: evidence from eye lens crystallins

Science.gov (United States)

Röll, Beate

2001-05-01

The large lizard family Gekkonidae comprises about 90 genera (1000 species). While most geckos are nocturnal, the members of about 15 genera are diurnal. All of these species are 'tertiarily' diurnal, i.e. they are descended from 'secondarily' nocturnal ancestors. They have adapted to a diurnal lifestyle in quite different ways, as can be deduced by the crystallin proteins in their lenses. Evaluation of the heterogeneous lens crystallin compositions of diurnal geckos reveals that there are at least three lineages that regained diurnality independently.

Impact of change of matrix crystallinity and polymorphism on ovalbumin release from lipid-based implants.

Science.gov (United States)

Duque, Luisa; Körber, Martin; Bodmeier, Roland

2018-05-30

The objectives of this study were to prepare lipid-based implants by hot melt extrusion (HME) for the prolonged release of ovalbumin (OVA), and to relate protein release to crystallinity and polymorphic changes of the lipid matrix. Two lipids, glycerol tristearate and hydrogenated palm oil, with different composition and degree of crystallinity were studied. Solid OVA was dispersed within the lipid matrixes, which preserved its stability during extrusion. This was partially attributed to a protective effect of the lipidic matrix. The incorporation of OVA decreased the mechanical strength of the implants prepared with the more crystalline matrix, glycerol tristearate, whereas it remained comparable for the hydrogenated palm oil because of stronger physical and non-covalent interactions between the protein and this lipid. This was also the reason for the faster release of OVA from the glycerol tristearate matrix when compared to the hydrogenated palm oil (8 vs. 28 weeks). Curing induced and increased crystallinity, and changes in the release rate, especially for the more crystalline matrix. In this case, both an increase and a decrease in release, were observed depending on the tempering condition. Curing at higher temperatures induced a melt-mediated crystallization and solid state transformation of the glycerol tristearate matrix and led to rearrangements of the inner structure with the formation of larger pores, which accelerated the release. In contrast, changes in the hydrogenated palm oil under the same curing conditions were less noticeable leading to a more robust formulation, because of less polymorphic changes over time. This study helps to understand the effect of lipid matrix composition and crystallinity degree on the performance of protein-loaded implants, and to establish criteria for the selection of a lipid carrier depending on the release profile desired. Copyright © 2018. Published by Elsevier B.V.

Alpha-crystallins are involved in specific interactions with the murine gamma D/E/F-crystallin-encoding gene.

Science.gov (United States)

Pietrowski, D; Durante, M J; Liebstein, A; Schmitt-John, T; Werner, T; Graw, J

1994-07-08

The promoter of the murine gamma E-crystallin (gamma E-Cry) encoding gene (gamma E-cry) was analyzed for specific interactions with lenticular proteins in a gel-retardation assay. A 21-bp fragment immediately downstream of the transcription initiation site (DOTIS) is demonstrated to be responsible for specific interactions with lens extracts. The DOTIS-binding protein(s) accept only the sense DNA strand as target; anti-sense or double-stranded DNA do not interact with these proteins. The DOTIS sequence element is highly conserved among the murine gamma D-, gamma E- and gamma F-cry and is present at comparable positions in the orthologous rat genes. Only a weak or even no protein-binding activity is observed if a few particular bases are changed, as in the rat gamma A-, gamma C- and gamma E-cry elements. DOTIS-binding proteins were found in commercially available bovine alpha-Cry preparations. The essential participation of alpha-Cry in the DNA-binding protein complex was confirmed using alpha-Cry-specific monoclonal antibody. The results reported here point to a novel function of alpha-Cry besides the structural properties in the lens.

Crystal Structure of Chicken γS-Crystallin Reveals Lattice Contacts with Implications for Function in the Lens and the Evolution of the βγ-Crystallins.

Science.gov (United States)

Sagar, Vatsala; Chaturvedi, Sumit K; Schuck, Peter; Wistow, Graeme

2017-07-05

Previous attempts to crystallize mammalian γS-crystallin were unsuccessful. Native L16 chicken γS crystallized avidly while the Q16 mutant did not. The X-ray structure for chicken γS at 2.3 Å resolution shows the canonical structure of the superfamily plus a well-ordered N arm aligned with a β sheet of a neighboring N domain. L16 is also in a lattice contact, partially shielded from solvent. Unexpectedly, the major lattice contact matches a conserved interface (QR) in the multimeric β-crystallins. QR shows little conservation of residue contacts, except for one between symmetry-related tyrosines, but molecular dipoles for the proteins with QR show striking similarities while other γ-crystallins differ. In γS, QR has few hydrophobic contacts and features a thin layer of tightly bound water. The free energy of QR is slightly repulsive and analytical ultracentrifugation confirms no dimerization in solution. The lattice contacts suggest how γ-crystallins allow close packing without aggregation in the crowded environment of the lens. Published by Elsevier Ltd.

Immunohistochemical Detection of a Unique Protein within Cells of Snakes Having Inclusion Body Disease, a World-Wide Disease Seen in Members of the Families Boidae and Pythonidae

Science.gov (United States)

Chang, Li-Wen; Fu, Ann; Wozniak, Edward; Chow, Marjorie; Duke, Diane G.; Green, Linda; Kelley, Karen; Hernandez, Jorge A.; Jacobson, Elliott R.

2013-01-01

Inclusion body disease (IBD) is a worldwide disease in captive boa constrictors (boa constrictor) and occasionally in other snakes of the families Boidae and Pythonidae. The exact causative agent(s) and pathogenesis are not yet fully understood. Currently, diagnosis of IBD is based on the light microscopic identification of eosinophilic intracytoplasmic inclusion bodies in hematoxylin and eosin stained tissues or blood smears. An antigenically unique 68 KDa protein was identified within the IBD inclusion bodies, called IBD protein. A validated immuno-based ante-mortem diagnostic test is needed for screening snakes that are at risk of having IBD. In this study, despite difficulties in solubilizing semi-purified inclusion bodies, utilizing hybridoma technology a mouse anti-IBD protein monoclonal antibody (MAB) was produced. The antigenic specificity of the antibody was confirmed and validated by western blots, enzyme-linked immunosorbent assay, immuno-transmission electron microscopy, and immunohistochemical staining. Paraffin embedded tissues of IBD positive and negative boa constrictors (n=94) collected from 1990 to 2011 were tested with immunohistochemical staining. In boa constrictors, the anti-IBDP MAB had a sensitivity of 83% and specificity of 100% in detecting IBD. The antibody also cross-reacted with IBD inclusion bodies in carpet pythons (Morelia spilota) and a ball python (python regius). This validated antibody can serve as a tool for the development of ante-mortem immunodiagnostic tests for IBD. PMID:24340066

Immunohistochemical detection of a unique protein within cells of snakes having inclusion body disease, a world-wide disease seen in members of the families Boidae and Pythonidae.

Directory of Open Access Journals (Sweden)

Li-Wen Chang

Full Text Available Inclusion body disease (IBD is a worldwide disease in captive boa constrictors (boa constrictor and occasionally in other snakes of the families Boidae and Pythonidae. The exact causative agent(s and pathogenesis are not yet fully understood. Currently, diagnosis of IBD is based on the light microscopic identification of eosinophilic intracytoplasmic inclusion bodies in hematoxylin and eosin stained tissues or blood smears. An antigenically unique 68 KDa protein was identified within the IBD inclusion bodies, called IBD protein. A validated immuno-based ante-mortem diagnostic test is needed for screening snakes that are at risk of having IBD. In this study, despite difficulties in solubilizing semi-purified inclusion bodies, utilizing hybridoma technology a mouse anti-IBD protein monoclonal antibody (MAB was produced. The antigenic specificity of the antibody was confirmed and validated by western blots, enzyme-linked immunosorbent assay, immuno-transmission electron microscopy, and immunohistochemical staining. Paraffin embedded tissues of IBD positive and negative boa constrictors (n=94 collected from 1990 to 2011 were tested with immunohistochemical staining. In boa constrictors, the anti-IBDP MAB had a sensitivity of 83% and specificity of 100% in detecting IBD. The antibody also cross-reacted with IBD inclusion bodies in carpet pythons (Morelia spilota and a ball python (python regius. This validated antibody can serve as a tool for the development of ante-mortem immunodiagnostic tests for IBD.

Protective Effects of Acetylation on the Pathological Reactions of the Lens Crystallins with Homocysteine Thiolactone.

Directory of Open Access Journals (Sweden)

Zeinab Moafian

Full Text Available Various post-translational lens crystallins modifications result in structural and functional insults, contributing to the development of lens opacity and cataract disorders. Lens crystallins are potential targets of homocysteinylation, particularly under hyperhomocysteinemia which has been indicated in various eye diseases. Since both homocysteinylation and acetylation primarily occur on protein free amino groups, we applied different spectroscopic methods and gel mobility shift analysis to examine the possible preventive role of acetylation against homocysteinylation. Lens crystallins were extensively acetylated in the presence of acetic anhydride and then subjected to homocysteinylation in the presence of homocysteine thiolactone (HCTL. Extensive acetylation of the lens crystallins results in partial structural alteration and enhancement of their stability, as well as improvement of α-crystallin chaperone-like activity. In addition, acetylation partially prevents HCTL-induced structural alteration and aggregation of lens crystallins. Also, acetylation protects against HCTL-induced loss of α-crystallin chaperone activity. Additionally, subsequent acetylation and homocysteinylation cause significant proteolytic degradation of crystallins. Therefore, further experimentation is required in order to judge effectively the preventative role of acetylation on the structural and functional insults induced by homocysteinylation of lens crystallins.

Crystallization and preliminary X-ray diffraction analysis of XAC1151, a small heat-shock protein from Xanthomonas axonopodis pv. citri belonging to the α-crystallin family

Energy Technology Data Exchange (ETDEWEB)

Hilario, Eduardo; Teixeira, Elaine Cristina; Pedroso, Gisele Audrei; Bertolini, Maria Célia [Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, Araraquara-SP (Brazil); Medrano, Francisco Javier, E-mail: fjmedrano@yahoo.com [Departamento de Cristalografia de Proteínas, Centro de Biologia Molecular Estrutural, Laboratório Nacional de Luz Síncrotron, Caixa Postal 6192, CEP 13084-971, Campinas-SP (Brazil); Departamento de Bioquímica e Tecnologia Química, Instituto de Química, Universidade Estadual Paulista, Araraquara-SP (Brazil)

2006-05-01

XAC1151, a small heat-shock protein from X. axonopodis pv. citri belonging to the α-crystallin family, was crystallized using the sitting-drop vapour-diffusion method in the presence of ammonium phosphate. X-ray diffraction data were collected to 1.65 Å resolution using a synchrotron-radiation source. The hspA gene (XAC1151) from Xanthomonas axonopodis pv. citri encodes a protein of 158 amino acids that belongs to the small heat-shock protein (sHSP) family of proteins. These proteins function as molecular chaperones by preventing protein aggregation. The protein was crystallized using the sitting-drop vapour-diffusion method in the presence of ammonium phosphate. X-ray diffraction data were collected to 1.65 Å resolution using a synchrotron-radiation source. The crystal belongs to the rhombohedral space group R3, with unit-cell parameters a = b = 128.7, c = 55.3 Å. The crystal structure was solved by molecular-replacement methods. Structure refinement is in progress.

Inclusion bodies are a site of ebolavirus replication.

Science.gov (United States)

Hoenen, Thomas; Shabman, Reed S; Groseth, Allison; Herwig, Astrid; Weber, Michaela; Schudt, Gordian; Dolnik, Olga; Basler, Christopher F; Becker, Stephan; Feldmann, Heinz

2012-11-01

Inclusion bodies are a characteristic feature of ebolavirus infections in cells. They contain large numbers of preformed nucleocapsids, but their biological significance has been debated, and they have been suggested to be aggregates of viral proteins without any further biological function. However, recent data for other viruses that produce similar structures have suggested that inclusion bodies might be involved in genome replication and transcription. In order to study filovirus inclusion bodies, we fused mCherry to the ebolavirus polymerase L, which is found in inclusion bodies. The resulting L-mCherry fusion protein was functional in minigenome assays and incorporated into virus-like particles. Importantly, L-mCherry fluorescence in transfected cells was readily detectable and distributed in a punctate pattern characteristic for inclusion bodies. A recombinant ebolavirus encoding L-mCherry instead of L was rescued and showed virtually identical growth kinetics and endpoint titers to those for wild-type virus. Using this virus, we showed that the onset of inclusion body formation corresponds to the onset of viral genome replication, but that viral transcription occurs prior to inclusion body formation. Live-cell imaging further showed that inclusion bodies are highly dynamic structures and that they can undergo dramatic reorganization during cell division. Finally, by labeling nascent RNAs using click technology we showed that inclusion bodies are indeed the site of viral RNA synthesis. Based on these data we conclude that, rather than being inert aggregates of nucleocapsids, ebolavirus inclusion bodies are in fact complex and dynamic structures and an important site at which viral RNA replication takes place.

Novel mesostructured inclusions in the epidermal lining of Artemia franciscana ovisacs show optical activity

Directory of Open Access Journals (Sweden)

Elena Hollergschwandtner

2017-10-01

Full Text Available Background Biomineralization, e.g., in sea urchins or mollusks, includes the assembly of mesoscopic superstructures from inorganic crystalline components and biopolymers. The resulting mesocrystals inspire biophysicists and material scientists alike, because of their extraordinary physical properties. Current efforts to replicate mesocrystal synthesis in vitro require understanding the principles of their self-assembly in vivo. One question, not addressed so far, is whether intracellular crystals of proteins can assemble with biopolymers into functional mesocrystal-like structures. During our electron microscopy studies into Artemia franciscana (Crustacea: Branchiopoda, we found initial evidence of such proteinaceous mesostructures. Results EM preparations with high-pressure freezing and accelerated freeze substitution revealed an extraordinary intracellular source of mesostructured inclusions in both the cyto-and nucleoplasm of the epidermal lining of ovisacs of A. franciscana. Confocal reflection microscopy not only confirmed our finding; it also revealed reflective, light dispersing activity of these flake-like structures, their positioning and orientation with respect to the ovisac inside. Both the striation of alternating electron dense and electron-lucent components and the sharp edges of the flakes indicate self-assembly of material of yet unknown origin under supposed participation of crystallization. However, selected area electron diffraction could not verify the status of crystallization. Energy dispersive X-ray analysis measured a marked increase in nitrogen within the flake-like inclusion, and the almost complete absence of elements that are typically involved in inorganic crystallization. This rise in nitrogen could possibility be related to higher package density of proteins, achieved by mesostructure assembly. Conclusions The ovisac lining of A. franciscana is endowed with numerous mesostructured inclusions that have not been

Structural Evolution of Human Recombinant alfaB-Crystallin under UV Irradiation

DEFF Research Database (Denmark)

Sugiyama, Masaaki; Fujii, Noriko; Morimoto, Yukio

2008-01-01

External stresses cause certain proteins to lose their regular structure and aggregate. In order to clarify this abnormal aggregation process, a structural evolution of human recombinant aB-crystallin under UV irradiation was observed with in situ small-angle neutron scattering. The abnormal...

Bacterial inclusion bodies as potential synthetic devices for pathogen recognition and a therapeutic substance release.

Science.gov (United States)

Talafová, Klaudia; Hrabárová, Eva; Chorvát, Dušan; Nahálka, Jozef

2013-02-07

Adhesins of pathogens recognise the glycans on the host cell and mediate adherence. They are also crucial for determining the tissue preferences of pathogens. Currently, glyco-nanomaterials provide potential tool for antimicrobial therapy. We demonstrate that properly glyco-tailored inclusion bodies can specifically bind pathogen adhesins and release therapeutic substances. In this paper, we describe the preparation of tailored inclusion bodies via the conjugation of indicator protein aggregated to form inclusion bodies with soluble proteins. Whereas the indicator protein represents a remedy, the soluble proteins play a role in pathogen recognition. For conjugation, glutaraldehyde was used as linker. The treatment of conjugates with polar lysine, which was used to inactivate the residual glutaraldehyde, inhibited unwanted hydrophobic interactions between inclusion bodies. The tailored inclusion bodies specifically interacted with the SabA adhesin from Helicobacter pylori aggregated to form inclusion bodies that were bound to the sialic acids decorating the surface of human erythrocytes. We also tested the release of indicator proteins from the inclusion bodies using sortase A and Ssp DNAB intein self-cleaving modules, respectively. Sortase A released proteins in a relatively short period of time, whereas the intein cleavage took several weeks. The tailored inclusion bodies are promising "nanopills" for biomedical applications. They are able to specifically target the pathogen, while a self-cleaving module releases a soluble remedy. Various self-cleaving modules can be enabled to achieve the diverse pace of remedy release.

Bacterial Inclusion Bodies: Discovering Their Better Half.

Science.gov (United States)

Rinas, Ursula; Garcia-Fruitós, Elena; Corchero, José Luis; Vázquez, Esther; Seras-Franzoso, Joaquin; Villaverde, Antonio

2017-09-01

Bacterial inclusion bodies (IBs) are functional, non-toxic amyloids occurring in recombinant bacteria showing analogies with secretory granules of the mammalian endocrine system. The scientific interest in these mesoscale protein aggregates has been historically masked by their status as a hurdle in recombinant protein production. However, progressive understanding of how the cell handles the quality of recombinant polypeptides and the main features of their intriguing molecular organization has stimulated the interest in inclusion bodies and spurred their use in diverse technological fields. The engineering and tailoring of IBs as functional protein particles for materials science and biomedicine is a good example of how formerly undesired bacterial byproducts can be rediscovered as promising functional materials for a broad spectrum of applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

Protein synthesis in x-irradiated rabbit lens

International Nuclear Information System (INIS)

Garadi, R.; Foltyn, A.R.; Giblin, F.J.; Reddy, V.N.

1984-01-01

The present study deals with the incorporation of 35 S methionine into lens crystallins as a function of time after x-irradiation. Crystallin synthesis is first affected approximately 4 weeks following x-irradiation. This coincides with the time period at which the ratio of the two cations in the lens is affected, as shown in earlier studies. A greater decrease in 35 S-methionine incorporation into crystallins is observed between 5-7 weeks following x-irradiation in good agreement with a cation imbalance at these time intervals. These studies also revealed for the first time that the change in cation distribution can affect not only crystallin synthesis, but also the synthesis of certain polypeptides of lens membranes. No alteration in protein synthesis could be detected in lens epithelium even after 7 weeks following irradiation. In addition to the effect of Na+ and K+ levels on protein synthesis, an impaired transport of amino acids into the x-rayed lens was also found to be a factor in the observed reduction in synthesis of the crystallin, cytoskeletal and membrane proteins of the fiber cells. It is concluded that Na+/K+ ratio as well as the availability of amino acids in the lens are important factors in protein synthesis of x-ray cataracts

Enhancement of norfloxacin solubility via inclusion complexation with β-cyclodextrin and its derivative hydroxypropyl-β-cyclodextrin

Directory of Open Access Journals (Sweden)

Gabriel Onn Kit Loh

2016-08-01

Full Text Available The objectives of the study were to investigate the effects of β-cyclodextrin (βCD and hydroxypropyl-β-cyclodextrin (HPβCD on the solubility and dissolution rate of norfloxacin prepared using three different methods, at drug to cyclodextrin weight ratios of 1:1, 1:2, 1:4 and 1:8. All the methods increased the solubility and dissolution rate of norfloxacin via inclusion complexation with βCD and HPβCD. Norfloxacin was converted from crystalline to amorphous form through inclusion complexation. Solvent evaporation method was the most effective method in terms of norfloxacin solubilisation, while inclusion complex of HPβCD has higher solubility than βCD complex when prepared using the same procedure.

Nuclear magnetic resonance studies of amino acids and proteins. Side-chain mobility of methionine in the crystalline amonio acid and in crystallne sperm whale (Physeter catodon) myoglobin

International Nuclear Information System (INIS)

Keniry, M.A.; Rothgeb, T.M.; Smith, R.L.; Gutowsky, H.S.; Oldfield, E.

1983-01-01

Deuterium ( 2 H) nuclear magnetic resonance (NMR) spectra and spin-lattice relaxation times (T 1 ) were obtained of L-[epsilon- 2 H 3 ]methionine, L-[epsilon- 2 H 3 ]methionine in a D,L lattice, and [S-methyl- 2 H 3 ]methionine in the crystalline solid state, as a function of temperature, in addition to obtaining 2 H T 1 and line-width results as a function of temperature on [epsilon- 2 H 3 ]methionine-labeled sperm whale (Physeter catodon) myoglobins by using the method of magnetic ordering. Also recorded were 13 C cross-polarization ''magic-angle'' sample-spinning NMR spectra of [epsilon- 13 C]methionine-labeled crystalline cyanoferrimyoglobin (at 37.7 MHz, corresponding to a magnetic field strength of 3.52 T) and of the same protein in aqueous solution

Water-structuring technology with the molecular chaperone proteins: indicated application of the α-crystallin domains and imidazole-containing peptidomimetics in cosmetic skin care systems or dermatological therapeutic drug carrier formulations.

Science.gov (United States)

Babizhayev, Mark A; Nikolayev, Gennady M; Nikolayeva, Juliana G; Yegorov, Yegor E

2011-01-01

Changes in structural proteins and hydration during aging are responsible for altered skin morphologic and mechanical properties manifested as wrinkling, sagging, loss of elasticity, and apparent dryness. Impairment in protein hydration may add to the ultrastructural, mechanical, and biochemical changes in structural proteins in the aged skin. At Innovative Vision Products, Inc., we have pioneered a molecular chaperone protein-activated therapeutic or cosmetic platform to enable simultaneous analysis of water-binding and structuring characteristics for biology-related or skin aging and skin disease-related pathways. This cutting-edge technology has changed the hydration of proteins in photoaged skin which so that they are more compact and interact with water to limited degree. The mechanisms of skin diseases, aging, and cellular and signaling pathways mediated by targeting with molecular chaperone protein(s) are considered. Skin lesions that are growing, spreading, or pigmented, and those that occur on exposed areas of skin are likely to be treated by these emerging pharmacological chaperones that could have cosmetic or dermatological benefits. Examples of such chaperones are anti-/trans-glycation-imidazole-containing peptidomimetic(s) (natural L-carnosine derivatives and mimetics) combined with the molecular chaperone protein α-crystallin derived from a natural source, brine shrimp (Artemia franciscana) cysts, or with recombinant human αA-crystallin. This patented biotechnology represents an efficient tool with which to mitigate the consequences of free radical-induced skin damage. The article is organized to provide in one place all of the relevant technical information, such as high-performance nuclear magnetic resonance and electron spin resonance application tools, and to describe the entire process from sample preparation to data analysis, which is moving from biological studies to biotechnology batches of the product. The proposed biotechnology results in

Formation of distinct inclusion bodies by inhibition of ubiquitin-proteasome and autophagy-lysosome pathways

International Nuclear Information System (INIS)

Lee, Junho; Yang, Kyu-Hwan; Joe, Cheol O.; Kang, Seok-Seong

2011-01-01

Research highlights: → Distinct inclusion bodies are developed by inhibition of UPP and ALP. → The inclusion bodies differ in morphology, localization and formation process. → The inclusion bodies are distinguishable by the localization of TSC2. → Inhibition of both UPP and ALP simultaneously induces those inclusion bodies. -- Abstract: Accumulation of misfolded proteins is caused by the impairment of protein quality control systems, such as ubiquitin-proteasome pathway (UPP) and autophagy-lysosome pathway (ALP). In this study, the formation of inclusion bodies was examined after the blockade of UPP and/or ALP in A549 cells. UPP inhibition induced a single and large inclusion body localized in microtubule-organizing center. Interestingly, however, ALP inhibition generated dispersed small inclusion bodies in the cytoplasm. Tuberous sclerosis complex 2 was selectively accumulated in the inclusion bodies of UPP-inhibited cells, but not those of ALP-inhibited cells. Blockade of transcription and translation entirely inhibited the formation of inclusion body induced by UPP inhibition, but partially by ALP inhibition. Moreover, the simultaneous inhibition of two protein catabolic pathways independently developed two distinct inclusion bodies within a single cell. These findings clearly demonstrated that dysfunction of each catabolic pathway induced formation and accumulation of unique inclusion bodies on the basis of morphology, localization and formation process in A549 cells.

Formation of distinct inclusion bodies by inhibition of ubiquitin-proteasome and autophagy-lysosome pathways.

Science.gov (United States)

Lee, Junho; Yang, Kyu-Hwan; Joe, Cheol O; Kang, Seok-Seong

2011-01-14

Accumulation of misfolded proteins is caused by the impairment of protein quality control systems, such as ubiquitin-proteasome pathway (UPP) and autophagy-lysosome pathway (ALP). In this study, the formation of inclusion bodies was examined after the blockade of UPP and/or ALP in A549 cells. UPP inhibition induced a single and large inclusion body localized in microtubule-organizing center. Interestingly, however, ALP inhibition generated dispersed small inclusion bodies in the cytoplasm. Tuberous sclerosis complex 2 was selectively accumulated in the inclusion bodies of UPP-inhibited cells, but not those of ALP-inhibited cells. Blockade of transcription and translation entirely inhibited the formation of inclusion body induced by UPP inhibition, but partially by ALP inhibition. Moreover, the simultaneous inhibition of two protein catabolic pathways independently developed two distinct inclusion bodies within a single cell. These findings clearly demonstrated that dysfunction of each catabolic pathway induced formation and accumulation of unique inclusion bodies on the basis of morphology, localization and formation process in A549 cells. Copyright © 2010 Elsevier Inc. All rights reserved.

[Solubilization Specificities Interferon beta-1b from Inclusion Bodies].

Science.gov (United States)

Zhuravko, A S; Kononova, N V; Bobruskin, A I

2015-01-01

A new solubilization method of recombinant interferon beta-1b (IFNβ-1b) from the inclusion bodies was developed. This method allows to extract the target protein selectively in the solutions of different alcohols, such as ethanol, propanol and isopropanol. It was shown that the more effective IFNβ-1b solubilization was achieved in the 55% propanol solution. This method allowed to extract the target protein from inclusion bodies around 85-90%, and significantly reduced Escherichia coli content in the solubilizate, in comparison with standard methods.

Crystalline color superconductivity

International Nuclear Information System (INIS)

Alford, Mark; Bowers, Jeffrey A.; Rajagopal, Krishna

2001-01-01

In any context in which color superconductivity arises in nature, it is likely to involve pairing between species of quarks with differing chemical potentials. For suitable values of the differences between chemical potentials, Cooper pairs with nonzero total momentum are favored, as was first realized by Larkin, Ovchinnikov, Fulde, and Ferrell (LOFF). Condensates of this sort spontaneously break translational and rotational invariance, leading to gaps which vary periodically in a crystalline pattern. Unlike the original LOFF state, these crystalline quark matter condensates include both spin-zero and spin-one Cooper pairs. We explore the range of parameters for which crystalline color superconductivity arises in the QCD phase diagram. If in some shell within the quark matter core of a neutron star (or within a strange quark star) the quark number densities are such that crystalline color superconductivity arises, rotational vortices may be pinned in this shell, making it a locus for glitch phenomena

Dynamic recruitment of active proteasomes into polyglutamine initiated inclusion bodies

NARCIS (Netherlands)

Schipper-Krom, Sabine; Juenemann, Katrin; Jansen, Anne H.; Wiemhoefer, Anne; van den Nieuwendijk, Rianne; Smith, Donna L.; Hink, Mark A.; Bates, Gillian P.; Overkleeft, Hermen; Ovaa, Huib; Reits, Eric

2014-01-01

Neurodegenerative disorders such as Huntington's disease are hallmarked by neuronal intracellular inclusion body formation. Whether proteasomes are irreversibly recruited into inclusion bodies in these protein misfolding disorders is a controversial subject. In addition, it has been proposed that

A Functional Core of IncA Is Required for Chlamydia trachomatis Inclusion Fusion.

Science.gov (United States)

Weber, Mary M; Noriea, Nicholas F; Bauler, Laura D; Lam, Jennifer L; Sager, Janet; Wesolowski, Jordan; Paumet, Fabienne; Hackstadt, Ted

2016-04-01

Chlamydia trachomatis is an obligate intracellular pathogen that is the etiological agent of a variety of human diseases, including blinding trachoma and sexually transmitted infections. Chlamydiae replicate within a membrane-bound compartment, termed an inclusion, which they extensively modify by the insertion of type III secreted proteins called Inc proteins. IncA is an inclusion membrane protein that encodes two coiled-coil domains that are homologous to eukaryotic SNARE (soluble N-ethylmaleimide-sensitive factor attachment receptor) motifs. Recent biochemical evidence suggests that a functional core, composed of SNARE-like domain 1 (SLD-1) and part of SNARE-like domain 2 (SLD-2), is required for the characteristic homotypic fusion of C. trachomatis inclusions in multiply infected cells. To verify the importance of IncA in homotypic fusion in Chlamydia, we generated an incA::bla mutant. Insertional inactivation of incA resulted in the formation of nonfusogenic inclusions, a phenotype that was completely rescued by complementation with full-length IncA. Rescue of homotypic inclusion fusion was dependent on the presence of the functional core consisting of SLD-1 and part of SLD-2. Collectively, these results confirm in vitro membrane fusion assays identifying functional domains of IncA and expand the genetic tools available for identification of chlamydia with a method for complementation of site-specific mutants. Chlamydia trachomatis replicates within a parasitophorous vacuole termed an inclusion. The chlamydial inclusions are nonfusogenic with vesicles in the endocytic pathway but, in multiply infected cells, fuse with each other to form a single large inclusion. This homotypic fusion is dependent upon the presence of a chlamydial inclusion membrane-localized protein, IncA. Specificity of membrane fusion in eukaryotic cells is regulated by SNARE (soluble N-ethylmaleimide sensitive factor attachment receptor) proteins on the cytosolic face of vesicles and target

Hard sphere-like glass transition in eye lens α-crystallin solutions.

Science.gov (United States)

Foffi, Giuseppe; Savin, Gabriela; Bucciarelli, Saskia; Dorsaz, Nicolas; Thurston, George M; Stradner, Anna; Schurtenberger, Peter

2014-11-25

We study the equilibrium liquid structure and dynamics of dilute and concentrated bovine eye lens α-crystallin solutions, using small-angle X-ray scattering, static and dynamic light scattering, viscometry, molecular dynamics simulations, and mode-coupling theory. We find that a polydisperse Percus-Yevick hard-sphere liquid-structure model accurately reproduces both static light scattering data and small-angle X-ray scattering liquid structure data from α-crystallin solutions over an extended range of protein concentrations up to 290 mg/mL or 49% vol fraction and up to ca. 330 mg/mL for static light scattering. The measured dynamic light scattering and viscosity properties are also consistent with those of hard-sphere colloids and show power laws characteristic of an approach toward a glass transition at α-crystallin volume fractions near 58%. Dynamic light scattering at a volume fraction beyond the glass transition indicates formation of an arrested state. We further perform event-driven molecular dynamics simulations of polydisperse hard-sphere systems and use mode-coupling theory to compare the measured dynamic power laws with those of hard-sphere models. The static and dynamic data, simulations, and analysis show that aqueous eye lens α-crystallin solutions exhibit a glass transition at high concentrations that is similar to those found in hard-sphere colloidal systems. The α-crystallin glass transition could have implications for the molecular basis of presbyopia and the kinetics of molecular change during cataractogenesis.

Specific racemization and isomerization of the aspartyl residue of αA-crystallin due to UV-B irradiation

International Nuclear Information System (INIS)

Fujii, Noriko; Momose, Yuko; Ishibasi, Yoshihiro; Uemura, Toshimasa; Takita, Masatoshi; Takehana, Makoto

1997-01-01

We have reported that the aspartyl (Asp)-151 residue in αA-crystallin in human eye lens was inverted to the D-isomer and isomerized to β-Asp residue with age. We report here that ultraviolet (UV)-B irradiation induces the racemization and isomerization of the Asp-151 residue of αA-crystallin from lenses of 6-week-old rats to form D-isomer and β-Asp residue. Simultaneous racemization and isomerization of the specific Asp residue indicate that the reaction proceeds via formation of a succinimide intermediate. This modification was not observed in UV-A irradiated and normal lenses. UV-B irradiation induced the racemization of only the Asp-151 residue and did not affect the other Asp residues in αA-crystallin. On the other hand, the high molecular weight fraction of the lens protein increased upon UV-B irradiation. Modification of the Asp residue would affect the three-dimensional packing array of the lens protein. (author)

Vacuolar processing enzyme plays an essential role in the crystalline structure of glutelin in rice seed.

Science.gov (United States)

Kumamaru, Toshihiro; Uemura, Yuji; Inoue, Yoshimi; Takemoto, Yoko; Siddiqui, Sadar Uddin; Ogawa, Masahiro; Hara-Nishimura, Ikuko; Satoh, Hikaru

2010-01-01

To identify the function of genes that regulate the processing of proglutelin, we performed an analysis of glup3 mutants, which accumulates excess amounts of proglutelin and lack the vacuolar processing enzyme (VPE). VPE activity in developing seeds from glup3 lines was reduced remarkably compared with the wild type. DNA sequencing of the VPE gene in glup3 mutants revealed either amino acid substitutions or the appearance of a stop codon within the coding region. Microscopic observations showed that alpha-globulin and proglutelin were distributed homogeneously within glup3 protein storage vacuoles (PSVs), and that glup3 PSVs lacked the crystalline lattice structure typical of wild-type PSVs. This suggests that the processing of proglutelin by VPE in rice is essential for proper PSV structure and compartmentalization of storage proteins. Growth retardation in glup3 seedlings was also observed, indicating that the processing of proglutelin influences early seedling development. These findings indicate that storage of glutelin in its mature form as a crystalline structure in PSVs is required for the rapid use of glutelin as a source of amino acids during early seedling development. In conclusion, VPE plays an important role in the formation of protein crystalline structures in PSVs.

Analysis of the 3’ untranslated regions of α-tubulin and S-crystallin mRNA and the identification of CPEB in dark- and light-adapted octopus retinas

Science.gov (United States)

Kelly, Shannan; Yamamoto, Hideki

2008-01-01

Purpose We previously reported the differential expression and translation of mRNA and protein in dark- and light-adapted octopus retinas, which may result from cytoplasmic polyadenylation element (CPE)–dependent mRNA masking and unmasking. Here we investigate the presence of CPEs in α-tubulin and S-crystallin mRNA and report the identification of cytoplasmic polyadenylation element binding protein (CPEB) in light- and dark-adapted octopus retinas. Methods 3’-RACE and sequencing were used to isolate and analyze the 3’-UTRs of α-tubulin and S-crystallin mRNA. Total retinal protein isolated from light- and dark-adapted octopus retinas was subjected to western blot analysis followed by CPEB antibody detection, PEP-171 inhibition of CPEB, and dephosphorylation of CPEB. Results The following CPE-like sequence was detected in the 3’-UTR of isolated long S-crystallin mRNA variants: UUUAACA. No CPE or CPE-like sequences were detected in the 3’-UTRs of α-tubulin mRNA or of the short S-crystallin mRNA variants. Western blot analysis detected CPEB as two putative bands migrating between 60-80 kDa, while a third band migrated below 30 kDa in dark- and light-adapted retinas. Conclusions The detection of CPEB and the identification of the putative CPE-like sequences in the S-crystallin 3’-UTR suggest that CPEB may be involved in the activation of masked S-crystallin mRNA, but not in the regulation of α-tubulin mRNA, resulting in increased S-crystallin protein synthesis in dark-adapted octopus retinas. PMID:18682811

Dipeptide repeat protein inclusions are rare in the spinal cord and almost absent from motor neurons in C9ORF72 mutant amyotrophic lateral sclerosis and are unlikely to cause their degeneration.

Science.gov (United States)

Gomez-Deza, Jorge; Lee, Youn-Bok; Troakes, Claire; Nolan, Matthew; Al-Sarraj, Safa; Gallo, Jean-Marc; Shaw, Christopher E

2015-06-25

Cytoplasmic TDP-43 inclusions are the pathological hallmark of amyotrophic lateral sclerosis (ALS) and tau-negative frontotemporal lobar dementia (FTLD). The G4C2 repeat mutation in C9ORF72 is the most common cause of ALS and FTLD in which, in addition to TDP-43 inclusions, five different di-peptide repeat (DPR) proteins have been identified. Di-peptide repeat proteins are translated in a non-canonical fashion from sense and antisense transcripts of the G4C2 repeat (GP, GA, GR, PA, PR). DPR inclusions are abundant in the cerebellum, as well as in the frontal and temporal lobes of ALS and FTLD patients and some are neurotoxic in a range of cellular and animal models, implying that DPR aggregation directly contributes to disease pathogenesis. Here we sought to quantify inclusions for each DPR and TDP-43 in ALS cases with and without the C9ORF72 mutation. We characterised the abundance of DPRs and their cellular location and compared this to cytoplasmic TDP-43 inclusions in order to explore the role of each inclusion in lower motor neuron degeneration. Spinal cord sections from ten cases positive for the C9ORF72 repeat expansion (ALS-C9+ve) and five cases that were not were probed by double immunofluorescence staining for individual DPRs and TDP-43. Inclusions immunoreactive for each of the DPRs were present in the spinal cord but they were rare or very rare in abundance (in descending order of frequency: GA, GP, GR, PA and PR). TDP-43 cytoplasmic inclusions were 45- to 750-fold more frequent than any DPR, and fewer than 4 % of DPR inclusions colocalized with TDP-43 inclusions. In motor neurons, a single cytoplasmic DPR inclusion was detected (0.1 %) in contrast to the 34 % of motor neurons that contained cytoplasmic TDP-43 inclusions. Furthermore, the number of TDP-43 inclusions in ALS cases with and without the C9ORF72 mutation was nearly identical. For all other neurodegenerative diseases, the neurotoxic protein aggregates are detected in the affected

Vibron Solitons and Soliton-Induced Infrared Spectra of Crystalline Acetanilide

Science.gov (United States)

Takeno, S.

1986-01-01

Red-shifted infrared spectra at low temperatures of amide I (C=O stretching) vibrations of crystalline acetanilide measured by Careri et al. are shown to be due to vibron solitons, which are nonlinearity-induced localized modes of vibrons arising from their nonlinear interactions with optic-type phonons. A nonlinear eigenvalue equation giving the eigenfrequency of stationary solitons is solved approximately by introducing lattice Green's functions, and the obtained result is in good agreement with the experimental result. Inclusion of interactions with acoustic phonons yields the Debye-Waller factor in the zero-phonon line spectrum of vibron solitons, in a manner analogous to the case of impurity-induced localized harmonic phonon modes in alkali halides.

Liquid crystalline order in polymers

CERN Document Server

Blumstein, Alexandre

1978-01-01

Liquid Crystalline Order in Polymers examines the topic of liquid crystalline order in systems containing rigid synthetic macromolecular chains. Each chapter of the book provides a review of one important area of the field. Chapter 1 discusses scattering in polymer systems with liquid crystalline order. It also introduces the field of liquid crystals. Chapter 2 treats the origin of liquid crystalline order in macromolecules by describing the in-depth study of conformation of such macromolecules in their unassociated state. The chapters that follow describe successively the liquid crystalli

Dynamic recruitment of active proteasomes into polyglutamine initiated inclusion bodies.

Science.gov (United States)

Schipper-Krom, Sabine; Juenemann, Katrin; Jansen, Anne H; Wiemhoefer, Anne; van den Nieuwendijk, Rianne; Smith, Donna L; Hink, Mark A; Bates, Gillian P; Overkleeft, Hermen; Ovaa, Huib; Reits, Eric

2014-01-03

Neurodegenerative disorders such as Huntington's disease are hallmarked by neuronal intracellular inclusion body formation. Whether proteasomes are irreversibly recruited into inclusion bodies in these protein misfolding disorders is a controversial subject. In addition, it has been proposed that the proteasomes may become clogged by the aggregated protein fragments, leading to impairment of the ubiquitin-proteasome system. Here, we show by fluorescence pulse-chase experiments in living cells that proteasomes are dynamically and reversibly recruited into inclusion bodies. As these recruited proteasomes remain catalytically active and accessible to substrates, our results challenge the concept of proteasome sequestration and impairment in Huntington's disease, and support the reported absence of proteasome impairment in mouse models of Huntington's disease. Copyright © 2013 Federation of European Biochemical Societies. All rights reserved.

Expression of nattokinase in Escherichia coli and renaturation of its inclusion body.

Science.gov (United States)

Ni, He; Guo, Peng-Cheng; Jiang, Wei-Ling; Fan, Xiao-Min; Luo, Xiang-Yu; Li, Hai-Hang

2016-08-10

Nattokinase is an important fibrinolytic enzyme with therapeutic applications for cardiovascular diseases. The full-length and mature nattokinase genes were cloned from Bacillus subtilis var. natto and expressed in pQE30 vector in Escherichia coli. The full-length gene expressed low nattokinase activity in the intracellular soluble and the medium fractions. The mature gene expressed low soluble nattokinase activity and large amount insoluble protein in inclusion bodies without enzyme activity. Large amount of refolding solutions (RSs) at different pH values were screening and RS-10 and RS-11 at pH 9 were selected to refold nattokinase inclusion bodies. The recombinant cells were lysed with 0.1mg/mL lysozyme and ultrasonic treatment. After centrifugation, the pellete was washed twice with 20mM Tris-HCl buffer (pH 7.5) containing 1% Triton X-100 to purify the inclusion bodies. The inclusion bodies were dissolved in water at pH 12.0 and refolded with RS-10. The refolded proteins showed 42.8IU/mg and 79.3IU/mg fibrinolytic activity by the traditional dilution method (20-fold dilution into RS-10) and the directly mixing the protein solution with equal volume RS-10, respectively, compared to the 52.0IU/mg of total water-soluble proteins from B. subtilis var. natto. This work demonstrated that the inclusion body of recombinant nattokinase expressed in E. coli could be simply refolded to the natural enzyme activity level by directly mixing the protein solution with equal volume refolding solution. Copyright © 2016 Elsevier B.V. All rights reserved.

[Inclusion Bodies are Formed in SFTSV-infected Human Macrophages].

Science.gov (United States)

Jin, Cong; Song, Jingdong; Han, Ying; Li, Chuan; Qiu, Peihong; Liang, Mifang

2016-01-01

The severe fever with thrombocytopenia syndrome virus (SFTSV) is a new member in the genus Phlebovirus of the family Bunyaviridae identified in China. The SFTSV is also the causative pathogen of an emerging infectious disease: severe fever with thrombocytopenia syndrome. Using immunofluorescent staining and confocal microscopy, the intracellular distribution of nucleocapsid protein (NP) in SFTSV-infected THP-1 cells was investigated with serial doses of SFTSV at different times after infection. Transmission electron microscopy was used to observe the ultrafine intracellular structure of SFTSV-infected THP-1 cells at different times after infection. SFTSV NP could form intracellular inclusion bodies in infected THP-1 cells. The association between NP-formed inclusion bodies and virus production was analyzed: the size of the inclusion body formed 3 days after infection was correlated with the viral load in supernatants collected 7 days after infection. These findings suggest that the inclusion bodies formed in SFTSV-infected THP-1 cells could be where the SFTSV uses host-cell proteins and intracellular organelles to produce new viral particles.

Pathogenesis of axonal dystrophy and demyelination in alphaA-crystallin-expressing transgenic mice.

NARCIS (Netherlands)

Rijk, A. van; Sweers, M.A.; Merkx, G.F.M.; Lammens, M.M.Y.; Bloemendal, H.

2003-01-01

We recently described a transgenic mouse strain overexpressing hamster alphaA-crystallin, a small heat shock protein, under direction of the hamster vimentin promoter. As a result myelin was degraded and axonal dystrophy in both central nervous system (especially spinal cord) and peripheral nervous

Spatial patterns of FUS-immunoreactive neuronal cytoplasmic inclusions (NCI) in neuronal intermediate filament inclusion disease (NIFID).

Science.gov (United States)

Armstrong, Richard A; Gearing, Marla; Bigio, Eileen H; Cruz-Sanchez, Felix F; Duyckaerts, Charles; Mackenzie, Ian R A; Perry, Robert H; Skullerud, Kari; Yokoo, Hideaki; Cairns, Nigel J

2011-11-01

Neuronal intermediate filament inclusion disease (NIFID), a rare form of frontotemporal lobar degeneration (FTLD), is characterized neuropathologically by focal atrophy of the frontal and temporal lobes, neuronal loss, gliosis, and neuronal cytoplasmic inclusions (NCI) containing epitopes of ubiquitin and neuronal intermediate filament (IF) proteins. Recently, the 'fused in sarcoma' (FUS) protein (encoded by the FUS gene) has been shown to be a component of the inclusions of NIFID. To further characterize FUS proteinopathy in NIFID, we studied the spatial patterns of the FUS-immunoreactive NCI in frontal and temporal cortex of 10 cases. In the cerebral cortex, sectors CA1/2 of the hippocampus, and the dentate gyrus (DG), the FUS-immunoreactive NCI were frequently clustered and the clusters were regularly distributed parallel to the tissue boundary. In a proportion of cortical gyri, cluster size of the NCI approximated to those of the columns of cells was associated with the cortico-cortical projections. There were no significant differences in the frequency of different types of spatial patterns with disease duration or disease stage. Clusters of NCI in the upper and lower cortex were significantly larger using FUS compared with phosphorylated, neurofilament heavy polypeptide (NEFH) or α-internexin (INA) immunohistochemistry (IHC). We concluded: (1) FUS-immunoreactive NCI exhibit similar spatial patterns to analogous inclusions in the tauopathies and synucleinopathies, (2) clusters of FUS-immunoreactive NCI are larger than those revealed by NEFH or ΙΝΑ, and (3) the spatial patterns of the FUS-immunoreactive NCI suggest the degeneration of the cortico-cortical projections in NIFID.

Preparation, physicochemical characterization and release behavior of the inclusion complex of trans-anethole and β-cyclodextrin.

Science.gov (United States)

Zhang, Wenwen; Li, Xinying; Yu, Taocheng; Yuan, Lun; Rao, Gang; Li, Defu; Mu, Changdao

2015-08-01

Trans-anethole (AT) has a variety of antimicrobial properties and is widely used as food functional ingredient. However, the applications of AT are limited due to its low water solubility, strong odor and low physicochemical stability. Therefore, the aim of this work was to encapsulate AT with β-cyclodextrin (β-CD) for obtaining inclusion complex by co-precipitation method. The measurements effectively confirmed the formation of inclusion complex between AT and β-CD. The results showed that the inclusion complex presented new solid crystalline phases and was more thermally stable than the physical mixture and β-CD. The phase solubility study showed that the aqueous solubility of AT was increased by being included in β-CD. The calculated stability constant of inclusion complex was 1195M -1 , indicating the strong interaction between AT and β-CD. Furthermore, the release study suggested that β-CD provided the protection for AT against evaporation. The release behavior of AT from the inclusion complex was controlled. Copyright © 2015 Elsevier Ltd. All rights reserved.

Probing the inhibitory potency of epigallocatechin gallate against human γB-crystallin aggregation: Spectroscopic, microscopic and simulation studies

Science.gov (United States)

Chaudhury, Susmitnarayan; Dutta, Anirudha; Bag, Sudipta; Biswas, Pranandita; Das, Amit Kumar; Dasgupta, Swagata

2018-03-01

Aggregation of human ocular lens proteins, the crystallins is believed to be one of the key reasons for age-onset cataract. Previous studies have shown that human γD-crystallin forms amyloid like fibres under conditions of low pH and elevated temperature. In this article, we have investigated the aggregation propensity of human γB-crystallin in absence and presence of epigallocatechin gallate (EGCG), in vitro, when exposed to stressful conditions. We have used different spectroscopic and microscopic techniques to elucidate the inhibitory effect of EGCG towards aggregation. The experimental results have been substantiated by molecular dynamics simulation studies. We have shown that EGCG possesses inhibitory potency against the aggregation of human γB-crystallin at low pH and elevated temperature.

Refolding in high hydrostatic pressure of recombinant proteins from inclusion bodies in Escherichia Coli; Renaturacao em altas pressoes hidrostaticas de proteinas recombinantes agregadas em corpos de inclusao produzidos em Escherichia Coli

Energy Technology Data Exchange (ETDEWEB)

Balduino, Keli Nunes

2009-07-01

The expression of proteins as inclusion bodies in bacteria is a widely used alternative for production of recombinant protein. However, the aggregation is a problem often encountered during refolding of these proteins. High hydrostatic pressure are able to solubilise the inclusion bodies in the presence of low concentrations of denaturant reagents, encouraging refolding protein with high efficiency and reduce costs. This work aims to refolding of recombinant proteins expressed in Escherichia coli from inclusion bodies using high hydrostatic pressure. Three toxins, all featuring five or more disulfide bonds were studied: NXH8, Natterin 2 and Bothropstoxin 1. Suspensions of inclusion bodies of the three proteins were pressurized to 2000 bars for 16 hours. The buffers were optimized for refolding of the three proteins. The buffer used in the refolding of NXH8 was 50 mM Tris HCl, pH 9.0 with proportion of 1GSH: 4GSSG at a concentration of 6 mM and 2 M GdnHCl. Inclusion bodies were used in O.D. (A600nm) of 0.5. After refolding process, dialysis was performed at pH 7.0. The final yield of obtaining soluble NXH8 was 40% (28,6 mg of soluble NXH8/L of culture medium). The refolding of Bothropstoxin 1 was obtained in refolding buffer of Tris HCl 50 mM, pH 7,5 with proportion of 2 GSH: GSSG 3 and concentration of 3 mM and 1 M GdnHCl. Use with a suspension of O.D. (A600nm) of 0.5. The final yield of recovery of Bothropstoxin 1 refolded was 32% (9,2 mg of refolded Bothropstoxin 1/L of culture medium). The refolding of Natterin 2 was performed in the refolding buffer: 20 mM Tris HCl pH 9.0 at a ratio of 2 GSH: 3GSSG and concentration of 10 mM and 1 M GdnHCl and inclusion bodies O.D. (A600nm) of 6.0. The yield of Natterin 2 refolded was 20% (3,7 mg/L of culture medium). Physico-chemical and biological analysis were performed by SDS-PAGE, western blot, scanning electron microscopy, biological tests in vivo and in vitro and structural. The analysis conducted in NXH8 did not show

Sporulation of Bacillus sphaericus 2297: an electron microscope study of crystal-like inclusion biogenesis and toxicity to mosquito larvae.

Science.gov (United States)

Kalfon, A; Charles, J F; Bourgouin, C; de Barjac, H

1984-04-01

Sporulation of Bacillus sphaericus strain 2297 in a synchronous liquid culture was studied by electron microscopy. The t0 of sporulation occurred 7 h after the beginning of the lag phase. Crystal-like inclusions first appeared at t2 and reached their final size between t5 and t6. The release of the spore/inclusion complex occurred at about t15 (22 h after inoculation). Toxicity against Culex pipiens larvae was related to sporulation and appeared during the early stages of sporulation. The LC50 (24 h) decreased about 10(5)-fold between t0-2 and t7, in correlation with the formation of crystalline inclusions. Heat resistance of spores appeared later than toxicity.

Influence of protein source on amino acid uptake patterns and protein utilization in rainbow trout Oncorhynchus mykiss

DEFF Research Database (Denmark)

Rolland, Marine; Holm, Jørgen; Dalsgaard, Anne Johanne Tang

induces reduced growth performances that remain partly unexplained. The aim of the current study was to investigate the effect of exchanging the protein source on protein utilization. Marine (fish meal) and vegetable (pea protein) sources were used with or without supplementation of crystalline amino......Matrixes of different protein sources (fish and plant products) combined with the use of crystalline amino acids allow for formulation of diets that meet fish requirements with little or no effect on protein digestibility and/or feed intake. Despite this, a total or partial replacement of fish meal...... acids to the fishmeal diet level (see Table 1). Amino acid uptake patterns were assessed by the appearance of amino acids in the blood stream following the ingestion of a meal, while dietary protein utilization was evaluated by examining the metabolic response to digestion and ammonium and urea...

Caveolin-2 associates with intracellular chlamydial inclusions independently of caveolin-1

Directory of Open Access Journals (Sweden)

Norkin Leonard C

2004-07-01

Full Text Available Abstract Background Lipid raft domains form in plasma membranes of eukaryotic cells by the tight packing of glycosphingolipids and cholesterol. Caveolae are invaginated structures that form in lipid raft domains when the protein caveolin-1 is expressed. The Chlamydiaceae are obligate intracellular bacterial pathogens that replicate entirely within inclusions that develop from the phagocytic vacuoles in which they enter. We recently found that host cell caveolin-1 is associated with the intracellular vacuoles and inclusions of some chlamydial strains and species, and that entry of those strains depends on intact lipid raft domains. Caveolin-2 is another member of the caveolin family of proteins that is present in caveolae, but of unknown function. Methods We utilized a caveolin-1 negative/caveolin-2 positive FRT cell line and laser confocal immunofluorescence techniques to visualize the colocalization of caveolin-2 with the chlamydial inclusions. Results We show here that in infected HeLa cells, caveolin-2, as well as caveolin-1, colocalizes with inclusions of C. pneumoniae (Cp, C. caviae (GPIC, and C. trachomatis serovars E, F and K. In addition, caveolin-2 also associates with C. trachomatis serovars A, B and C, although caveolin-1 did not colocalize with these organisms. Moreover, caveolin-2 appears to be specifically, or indirectly, associated with the pathogens at the inclusion membranes. Using caveolin-1 deficient FRT cells, we show that although caveolin-2 normally is not transported out of the Golgi in the absence of caveolin-1, it nevertheless colocalizes with chlamydial inclusions in these cells. However, our results also show that caveolin-2 did not colocalize with UV-irradiated Chlamydia in FRT cells, suggesting that in these caveolin-1 negative cells, pathogen viability and very likely pathogen gene expression are necessary for the acquisition of caveolin-2 from the Golgi. Conclusion Caveolin-2 associates with the chlamydial

The spectrum and severity of FUS-immunoreactive inclusions in the frontal and temporal lobes of ten cases of neuronal intermediate filament inclusion disease.

Science.gov (United States)

Armstrong, Richard A; Gearing, Marla; Bigio, Eileen H; Cruz-Sanchez, Felix F; Duyckaerts, Charles; Mackenzie, Ian R A; Perry, Robert H; Skullerud, Kari; Yokoo, Hedeaki; Cairns, Nigel J

2011-02-01

Neuronal intermediate filament inclusion disease (NIFID), a rare form of frontotemporal lobar degeneration (FTLD), is characterized neuropathologically by focal atrophy of the frontal and temporal lobes, neuronal loss, gliosis, and neuronal cytoplasmic inclusions (NCI) containing epitopes of ubiquitin and neuronal intermediate filament proteins. Recently, the 'fused in sarcoma' (FUS) protein (encoded by the FUS gene) has been shown to be a component of the inclusions of familial amyotrophic lateral sclerosis with FUS mutation, NIFID, basophilic inclusion body disease, and atypical FTLD with ubiquitin-immunoreactive inclusions (aFTLD-U). To further characterize FUS proteinopathy in NIFID, and to determine whether the pathology revealed by FUS immunohistochemistry (IHC) is more extensive than α-internexin, we have undertaken a quantitative assessment of ten clinically and neuropathologically well-characterized cases using FUS IHC. The densities of NCI were greatest in the dentate gyrus (DG) and in sectors CA1/2 of the hippocampus. Anti-FUS antibodies also labeled glial inclusions (GI), neuronal intranuclear inclusions (NII), and dystrophic neurites (DN). Vacuolation was extensive across upper and lower cortical layers. Significantly greater densities of abnormally enlarged neurons and glial cell nuclei were present in the lower compared with the upper cortical laminae. FUS IHC revealed significantly greater numbers of NCI in all brain regions especially the DG. Our data suggest: (1) significant densities of FUS-immunoreactive NCI in NIFID especially in the DG and CA1/2; (2) infrequent FUS-immunoreactive GI, NII, and DN; (3) widely distributed vacuolation across the cortex, and (4) significantly more NCI revealed by FUS than α-internexin IHC.

Renal pathophysiologic role of cortical tubular inclusion bodies.

Science.gov (United States)

Radi, Zaher A; Stewart, Zachary S; Grzemski, Felicity A; Bobrowski, Walter F

2013-01-01

Renal tubular inclusion bodies are rarely associated with drug administration. The authors describe the finding of renal cortical tubular intranuclear and intracytoplasmic inclusion bodies associated with the oral administration of a norepinephrine/serotonin reuptake inhibitor (NSRI) test article in Sprague-Dawley (SD) rats. Rats were given an NSRI daily for 4 weeks, and kidney histopathologic, ultrastructural pathology, and immunohistochemical examinations were performed. Round eosinophilic intranuclear inclusion bodies were observed histologically in the tubular epithelial cells of the renal cortex in male and female SD rats given the NSRI compound. No evidence of degeneration or necrosis was noted in the inclusion-containing renal cells. By ultrastructural pathology, inclusion bodies consisted of finely granular, amorphous, and uniformly stained nonmembrane-bound material. By immunohistochemistry, inclusion bodies stained positive for d-amino acid oxidase (DAO) protein. In addition, similar inclusion bodies were noted in the cytoplasmic tubular epithelial compartment by ultrastructural and immunohistochemical examination.  This is the first description of these renal inclusion bodies after an NSRI test article administration in SD rats. Such drug-induced renal inclusion bodies are rat-specific, do not represent an expression of nephrotoxicity, represent altered metabolism of d-amino acids, and are not relevant to human safety risk assessment.

ir overtone spectrum of the vibrational soliton in crystalline acetanilide

International Nuclear Information System (INIS)

Scott, A.C.; Gratton, E.; Shyamsunder, E.; Careri, G.

1985-01-01

The self-trapping (soliton) theory which was recently developed to account for the anomalous amide-I band at 1650 cm -1 in crystalline acetanilide (a model system for protein) has been extended to predict the anharmonicity constant of the overtone spectrum. These infrared-active overtones which have been detected at 3250, 4803, and 6304 cm -1 yield an anharmonicity constant that is in good agreement with the theory

ir overtone spectrum of the vibrational soliton in crystalline acetanilide

Science.gov (United States)

Scott, A. C.; Gratton, E.; Shyamsunder, E.; Careri, G.

1985-10-01

The self-trapping (soliton) theory which was recently developed to account for the anomalous amide-I band at 1650 cm-1 in crystalline acetanilide (a model system for protein) has been extended to predict the anharmonicity constant of the overtone spectrum. These infrared-active overtones which have been detected at 3250, 4803, and 6304 cm-1 yield an anharmonicity constant that is in good agreement with the theory.

Inclusões intracelulares associadas à "espiga branca" do trigo Intracellular inclusions associated with white spike disease of wheat

Directory of Open Access Journals (Sweden)

E. W. Kitajima

1971-05-01

section profiles as loose aggregates of threadlike elements, 7-10 m¼ in diameter and of indeterminable length, within the cytoplasm. They seem to represent filamentous particles found in leaf dip preparations from affected plants, in situ. Nucleolus hypertrophy, chloroplast degeneration and changes in endoplasmic reticulum were frequently noticed in affected tissues. Occasionally small crystalline inclusions were found nearby or within the nucleus. The presence of these fibrous, whorled inclusions associated to WS condition, reinforces the suggested viral etiology, due to the similarity of these inclusions with virus aggregates found in tissues infected with several other viruses of the beet yellows virus group. Besides symptomatology, the particle and inclusion morphology of rice "hoja blanca" virus are very similar to WS of wheat, suggesting that both diseases are induced by viruses of the same complex. The ease with which the inclusions are detected in epidermal strips, constitutes a quick and secure method to diagnose the WS disease.

Improvements in numerical modelling of highly injected crystalline silicon solar cells

Energy Technology Data Exchange (ETDEWEB)

Altermatt, P.P. [University of New South Wales, Centre for Photovoltaic Engineering, 2052 Sydney (Australia); Sinton, R.A. [Sinton Consulting, 1132 Green Circle, 80303 Boulder, CO (United States); Heiser, G. [University of NSW, School of Computer Science and Engineering, 2052 Sydney (Australia)

2001-01-01

We numerically model crystalline silicon concentrator cells with the inclusion of band gap narrowing (BGN) caused by injected free carriers. In previous studies, the revised room-temperature value of the intrinsic carrier density, n{sub i}=1.00x10{sup 10}cm{sup -3}, was inconsistent with the other material parameters of highly injected silicon. In this paper, we show that high-injection experiments can be described consistently with the revised value of n{sub i} if free-carrier induced BGN is included, and that such BGN is an important effect in silicon concentrator cells. The new model presented here significantly improves the ability to model highly injected silicon cells with a high level of precision.

Stoichiometric Assembly of the Cellulosome Generates Maximum Synergy for the Degradation of Crystalline Cellulose, as Revealed by In Vitro Reconstitution of the Clostridium thermocellum Cellulosome.

Science.gov (United States)

Hirano, Katsuaki; Nihei, Satoshi; Hasegawa, Hiroki; Haruki, Mitsuru; Hirano, Nobutaka

2015-07-01

The cellulosome is a supramolecular multienzyme complex formed by species-specific interactions between the cohesin modules of scaffoldin proteins and the dockerin modules of a wide variety of polysaccharide-degrading enzymes. Cellulosomal enzymes bound to the scaffoldin protein act synergistically to degrade crystalline cellulose. However, there have been few attempts to reconstitute intact cellulosomes due to the difficulty of heterologously expressing full-length scaffoldin proteins. We describe the synthesis of a full-length scaffoldin protein containing nine cohesin modules, CipA; its deletion derivative containing two cohesin modules, ΔCipA; and three major cellulosomal cellulases, Cel48S, Cel8A, and Cel9K, of the Clostridium thermocellum cellulosome. The proteins were synthesized using a wheat germ cell-free protein synthesis system, and the purified proteins were used to reconstitute cellulosomes. Analysis of the cellulosome assembly using size exclusion chromatography suggested that the dockerin module of the enzymes stoichiometrically bound to the cohesin modules of the scaffoldin protein. The activity profile of the reconstituted cellulosomes indicated that cellulosomes assembled at a CipA/enzyme molar ratio of 1/9 (cohesin/dockerin = 1/1) and showed maximum synergy (4-fold synergy) for the degradation of crystalline substrate and ∼2.4-fold-higher synergy for its degradation than minicellulosomes assembled at a ΔCipA/enzyme molar ratio of 1/2 (cohesin/dockerin = 1/1). These results suggest that the binding of more enzyme molecules on a single scaffoldin protein results in higher synergy for the degradation of crystalline cellulose and that the stoichiometric assembly of the cellulosome, without excess or insufficient enzyme, is crucial for generating maximum synergy for the degradation of crystalline cellulose. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Blending crystalline/liquid crystalline small molecule semiconductors: A strategy towards high performance organic thin film transistors

Science.gov (United States)

He, Chao; He, Yaowu; Li, Aiyuan; Zhang, Dongwei; Meng, Hong

2016-10-01

Solution processed small molecule polycrystalline thin films often suffer from the problems of inhomogeneity and discontinuity. Here, we describe a strategy to solve these problems through deposition of the active layer from a blended solution of crystalline (2-phenyl[1]benzothieno[3,2-b][1]benzothiophene, Ph-BTBT) and liquid crystalline (2-(4-dodecylphenyl) [1]benzothieno[3,2-b]benzothiophene, C12-Ph-BTBT) small molecule semiconductors with the hot spin-coating method. Organic thin film transistors with average hole mobility approaching 1 cm2/V s, much higher than that of single component devices, have been demonstrated, mainly due to the improved uniformity, continuity, crystallinity, and stronger intermolecular π-π stacking in blend thin films. Our results indicate that the crystalline/liquid crystalline semiconductor blend method is an effective way to enhance the performance of organic transistors.

Effect of Ultrasonic Vibration on Mechanical Properties of 3D Printing Non-Crystalline and Semi-Crystalline Polymers.

Science.gov (United States)

Li, Guiwei; Zhao, Ji; Wu, Wenzheng; Jiang, Jili; Wang, Bofan; Jiang, Hao; Fuh, Jerry Ying Hsi

2018-05-17

Fused deposition modeling 3D printing has become the most widely used additive manufacturing technology because of its low manufacturing cost and simple manufacturing process. However, the mechanical properties of the 3D printing parts are not satisfactory. Certain pressure and ultrasonic vibration were applied to 3D printed samples to study the effect on the mechanical properties of 3D printed non-crystalline and semi-crystalline polymers. The tensile strength of the semi-crystalline polymer polylactic acid was increased by 22.83% and the bending strength was increased by 49.05%, which were almost twice the percentage increase in the tensile strength and five times the percentage increase in the bending strength of the non-crystalline polymer acrylonitrile butadiene styrene with ultrasonic strengthening. The dynamic mechanical properties of the non-crystalline and semi-crystalline polymers were both improved after ultrasonic enhancement. Employing ultrasonic energy can significantly improve the mechanical properties of samples without modifying the 3D printed material or adjusting the forming process parameters.

Fox-2 Splicing Factor Binds to a Conserved Intron Motif to PromoteInclusion of Protein 4.1R Alternative Exon 16

Energy Technology Data Exchange (ETDEWEB)

Ponthier, Julie L.; Schluepen, Christina; Chen, Weiguo; Lersch,Robert A.; Gee, Sherry L.; Hou, Victor C.; Lo, Annie J.; Short, Sarah A.; Chasis, Joel A.; Winkelmann, John C.; Conboy, John G.

2006-03-01

Activation of protein 4.1R exon 16 (E16) inclusion during erythropoiesis represents a physiologically important splicing switch that increases 4.1R affinity for spectrin and actin. Previous studies showed that negative regulation of E16 splicing is mediated by the binding of hnRNP A/B proteins to silencer elements in the exon and that downregulation of hnRNP A/B proteins in erythroblasts leads to activation of E16 inclusion. This paper demonstrates that positive regulation of E16 splicing can be mediated by Fox-2 or Fox-1, two closely related splicing factors that possess identical RNA recognition motifs. SELEX experiments with human Fox-1 revealed highly selective binding to the hexamer UGCAUG. Both Fox-1 and Fox-2 were able to bind the conserved UGCAUG elements in the proximal intron downstream of E16, and both could activate E16 splicing in HeLa cell co-transfection assays in a UGCAUG-dependent manner. Conversely, knockdown of Fox-2 expression, achieved with two different siRNA sequences resulted in decreased E16 splicing. Moreover, immunoblot experiments demonstrate mouse erythroblasts express Fox-2, but not Fox-1. These findings suggest that Fox-2 is a physiological activator of E16 splicing in differentiating erythroid cells in vivo. Recent experiments show that UGCAUG is present in the proximal intron sequence of many tissue-specific alternative exons, and we propose that the Fox family of splicing enhancers plays an important role in alternative splicing switches during differentiation in metazoan organisms.

Tunable geometry of bacterial inclusion bodies as substrate materials for tissue engineering

Energy Technology Data Exchange (ETDEWEB)

GarcIa-Fruitos, Elena; Seras-Franzoso, JoaquIn; Vazquez, Esther; Villaverde, Antonio [CIBER en BioingenierIa, Biomateriales y Nanomedicina, Bellaterra, 08193 Barcelona (Spain); Institut de Biotecnologia i de Biomedicina and Departament de Genetica i de Microbiologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Cerdanyola del Valles), Barcelona (Spain)

2010-05-21

A spectrum of materials for biomedical applications is produced in bacteria, and some of them, such as metals or polyhydroxyalkanoates, are straightforwardly obtained as particulate entities. We have explored the biofabrication process of bacterial inclusion bodies, particulate proteinaceous materials (ranging from 50 to 500 nm in diameter) recently recognized as suitable for surface topographical modification and tissue engineering. Inclusion bodies have been widely described as spherical or pseudo-spherical particles with only minor morphological variability, mostly restricted to their size. Here we have identified a cellular gene in Escherichia coli (clpP) that controls the in vivo fabrication process of inclusion bodies. In the absence of the encoded protease, the dynamics of protein deposition is perturbed, resulting in unusual tear-shaped particles with enhanced surface-volume ratios. This fact modifies the ability of inclusion bodies to promote mammalian cell attachment and differentiation upon surface decoration. The implications of the genetic control of inclusion body geometry are discussed in the context of their biological fabrication and regarding the biomedical potential of these protein clusters in regenerative medicine.

Tunable geometry of bacterial inclusion bodies as substrate materials for tissue engineering

International Nuclear Information System (INIS)

GarcIa-Fruitos, Elena; Seras-Franzoso, JoaquIn; Vazquez, Esther; Villaverde, Antonio

2010-01-01

A spectrum of materials for biomedical applications is produced in bacteria, and some of them, such as metals or polyhydroxyalkanoates, are straightforwardly obtained as particulate entities. We have explored the biofabrication process of bacterial inclusion bodies, particulate proteinaceous materials (ranging from 50 to 500 nm in diameter) recently recognized as suitable for surface topographical modification and tissue engineering. Inclusion bodies have been widely described as spherical or pseudo-spherical particles with only minor morphological variability, mostly restricted to their size. Here we have identified a cellular gene in Escherichia coli (clpP) that controls the in vivo fabrication process of inclusion bodies. In the absence of the encoded protease, the dynamics of protein deposition is perturbed, resulting in unusual tear-shaped particles with enhanced surface-volume ratios. This fact modifies the ability of inclusion bodies to promote mammalian cell attachment and differentiation upon surface decoration. The implications of the genetic control of inclusion body geometry are discussed in the context of their biological fabrication and regarding the biomedical potential of these protein clusters in regenerative medicine.

Nuclear Glycogen Inclusions in Canine Parietal Cells.

Science.gov (United States)

Silvestri, S; Lepri, E; Dall'Aglio, C; Marchesi, M C; Vitellozzi, G

2017-05-01

Nuclear glycogen inclusions occur infrequently in pathologic conditions but also in normal human and animal tissues. Their function or significance is unclear. To the best of the authors' knowledge, no reports of nuclear glycogen inclusions in canine parietal cells exist. After initial observations of nuclear inclusions/pseudoinclusions during routine histopathology, the authors retrospectively examined samples of gastric mucosa from dogs presenting with gastrointestinal signs for the presence of intranuclear inclusions/pseudoinclusions and determined their composition using histologic and electron-microscopic methods. In 24 of 108 cases (22%), the authors observed various numbers of intranuclear inclusions/pseudoinclusions within scattered parietal cells. Nuclei were characterized by marked karyomegaly and chromatin margination around a central optically empty or slightly eosinophilic area. The intranuclear inclusions/pseudoinclusions stained positive with periodic acid-Schiff (PAS) and were diastase sensitive, consistent with glycogen. Several PAS-positive/diastase-sensitive sections were further examined by transmission electron microscopy, also using periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) staining to identify polysaccharides. Ultrastructurally, the nuclear inclusions were composed of electron-dense particles that were not membrane bound, without evidence of nuclear membrane invaginations or cytoplasmic organelles in the nuclei, and positive staining with PA-TCH-SP, confirming a glycogen composition. No cytoplasmic glycogen deposits were observed, suggesting that the intranuclear glycogen inclusions were probably synthesized in loco. Nuclear glycogen inclusions were not associated with gastritis or colonization by Helicobacter-like organisms ( P > .05). Our findings suggest that nuclear glycogen inclusions in canine parietal cells could be an incidental finding. Nevertheless, since nuclear glycogen is present in several pathologic

Dissolution of crystalline ceramics

International Nuclear Information System (INIS)

White, W.B.

1982-01-01

The present program objectives are to lay out the fundamentals of crystalline waste form dissolution. Nuclear waste ceramics are polycrystalline. An assumption of the work is that to the first order, the release rate of a particular radionuclide is the surface-weighted sum of the release rates of the radionuclide from each crystalline form that contains it. In the second order, of course, there will be synergistic effects. There will be also grain boundary and other microstructural influences. As a first approximation, we have selected crystalline phases one at a time. The sequence of investigations and measurements is: (i) Identification of the actual chemical reactions of dissolution including identification of the solid reaction products if such occur. (ii) The rates of these reactions are then determined empirically to give what may be called macroscopic kinetics. (iii) Determination of the rate-controlling mechanisms. (iv) If the rate is controlled by surface reactions, the final step would be to determine the atomic kinetics, that is the specific atomic reactions that occur at the dissolving interface. Our concern with the crystalline forms are in two areas: The crystalline components of the reference ceramic waste form and related ceramics and the alumino-silicate phases that appear in some experimental waste forms and as waste-rock interaction products. Specific compounds are: (1) Reference Ceramic Phases (zirconolite, magnetoplumbite, spinel, Tc-bearing spinel and perovskite); (2) Aluminosilicate phases (nepheline, pollucite, CsAlSi 5 O 12 , Sr-feldspar). 5 figures, 1 table

Social inclusion and inclusive education

Directory of Open Access Journals (Sweden)

Marsela Robo

2014-07-01

In line with global debate on social inclusion and exclusion, the author brings the way this debate has now pervaded both the official and development policy discourse in Albania.Social inclusion is considered as one of the priorities of the current government, with poverty reduction as its main focus, which will be ensured not only through economic development. In the end, the article focuses on the role of education as a very important and useful tool for ensuring social inclusion.Social inclusion through education, in particular through vocational education, considered by the author as the only way towards sustainable development of Albanian society.

Diverse topics in crystalline beams

International Nuclear Information System (INIS)

Wei, Jie; Draeseke, A.; Sessler, A.M.; Li, Xiao-Ping

1995-01-01

Equations of motion are presented, appropriate to interacting charged particles of diverse charge and mass, subject to the external forces produced by various kinds of magnetic fields and radio-frequency (rf) electric fields in storage rings. These equations are employed in the molecular dynamics simulations to study the properties of crystalline beams. The two necessary conditions for the formation and maintenance of crystalline beams are summarized. The transition from ID to 2D, and from 2D to 3D is explored, and the scaling behavior of the heating rates is discussed especially in the high temperature limit. The effectiveness of various cooling techniques in achieving crystalline states has been investigated. Crystalline beams made of two different species of ions via sympathetic cooling are presented, as well as circulating ''crystal balls'' bunched in all directions by magnetic focusing and rf field. By numerically reconstructing the original experimental conditions of the NAP-M ring, it is found that only at extremely low beam intensities, outside of the range of the original measurement, proton particles can form occasionally-passing disks. The proposed New ASTRID ring is shown to be suitable for the formation and maintenance of crystalline beams of all dimensions

Dynamic JUNQ inclusion bodies are asymmetrically inherited in mammalian cell lines through the asymmetric partitioning of vimentin.

Science.gov (United States)

Ogrodnik, Mikołaj; Salmonowicz, Hanna; Brown, Rachel; Turkowska, Joanna; Średniawa, Władysław; Pattabiraman, Sundararaghavan; Amen, Triana; Abraham, Ayelet-chen; Eichler, Noam; Lyakhovetsky, Roman; Kaganovich, Daniel

2014-06-03

Aging is associated with the accumulation of several types of damage: in particular, damage to the proteome. Recent work points to a conserved replicative rejuvenation mechanism that works by preventing the inheritance of damaged and misfolded proteins by specific cells during division. Asymmetric inheritance of misfolded and aggregated proteins has been shown in bacteria and yeast, but relatively little evidence exists for a similar mechanism in mammalian cells. Here, we demonstrate, using long-term 4D imaging, that the vimentin intermediate filament establishes mitotic polarity in mammalian cell lines and mediates the asymmetric partitioning of damaged proteins. We show that mammalian JUNQ inclusion bodies containing soluble misfolded proteins are inherited asymmetrically, similarly to JUNQ quality-control inclusions observed in yeast. Mammalian IPOD-like inclusion bodies, meanwhile, are not always inherited by the same cell as the JUNQ. Our study suggests that the mammalian cytoskeleton and intermediate filaments provide the physical scaffold for asymmetric inheritance of dynamic quality-control JUNQ inclusions. Mammalian IPOD inclusions containing amyloidogenic proteins are not partitioned as effectively during mitosis as their counterparts in yeast. These findings provide a valuable mechanistic basis for studying the process of asymmetric inheritance in mammalian cells, including cells potentially undergoing polar divisions, such as differentiating stem cells and cancer cells.

Molecular reorientations in a substance with liquid-crystalline and plastic-crystalline phases

International Nuclear Information System (INIS)

Nguyen, Xuan Phuc.

1986-05-01

Results of dielectric relaxation (DR), quasielastic neutron scattering (QNS), far infrared absorption (FIR), proton magnetic resonance (PMR), differential scanning calorimetry (DSC) and preliminary X-ray diffraction measurements on the di-n-pentyloxyazoxybenzene (5.OAOB) are presented. The measurements carried out by all these methods showed that 5.OAOB exhibits a nontypical for liquid-crystalline materials phase diagram. It has two mesophases: a nematic (N) and an ''intermediate'' crystalline phase just below it. A complex interpretation of results obtained is given. All suggestions concerning the character of reorientational motions of the molecule as a whole as well as of its segments in mesomorphic phases are analyzed. From comparison of the DR and QNS studies one can conclude that in the N phase the molecule as a whole performs rotational diffusion around the long axis (τ DR ∼ 100 ps) and at the same time the two moieties perform faster independent reorientations around N - benzene rings bonds withτ QNS ∼ 5 ps. On the basis of various experimental data it is shown that the CrI phase is a plastic-crystalline phase for which the molecule and its segments perform fast stochastic unaxial reorientations. This is the first case where the existence of such a phase in liquid-crystalline materials has been experimentally confirmed. (author)

Neutron transmission through crystalline Fe

International Nuclear Information System (INIS)

Adib, M.; Habib, N.; Kilany, M.; El-Mesiry, M.S.

2004-01-01

The neutron transmission through crystalline Fe has been calculated for neutron energies in the range 10 4 < E<10 eV using an additive formula. The formula permits calculation of the nuclear capture, thermal diffuse and Bragg scattering cross-section as a function of temperature and crystalline form. The obtained agreement between the calculated values and available experimental ones justifies the applicability of the used formula. A feasibility study on using poly-crystalline Fe as a cold neutron filter and a large Fe single crystal as a thermal one is given

Paleohydrogeological events recorded by stable isotopes, fluid inclusions and trace elements in fracture minerals in crystalline rock, Simpevarp area, SE Sweden

International Nuclear Information System (INIS)

Drake, Henrik; Tullborg, Eva-Lena

2009-01-01

Fracture minerals calcite, pyrite, gypsum, barite and quartz, formed during several events have been analysed for δ 13 C, δ 18 O, δ 34 S, 87 Sr/ 86 Sr, trace element chemistry and fluid inclusions in order to gain knowledge of the paleohydrogeological evolution of the Simpevarp area, south-eastern Sweden. This area is dominated by Proterozoic crystalline rocks and is currently being investigated by the Swedish Nuclear Fuel and Waste Management Co. (SKB) in order to find a suitable location for a deep-seated repository for spent nuclear fuel. Knowledge of the paleohydrogeological evolution is essential to understand the stability or evolution of the groundwater system over a time scale relevant to the performance assessment for a spent nuclear fuel repository. The ages of the minerals analysed range from the Proterozoic to possibly the Quaternary. The Proterozoic calcite and pyrite show inorganic and hydrothermal-magmatic stable isotope signatures and were probably formed during a long time period as indicated by the large span in temperatures (c. 200-360 deg. C) and salinities (0-24 wt.% eq. CaCl 2 ), obtained from fluid inclusion analyses. The Paleozoic minerals were formed from organically influenced brine-type fluids at temperatures of 80-145 deg. C. The isotopic results indicate that low temperature calcite and pyrite may have formed during different events ranging in time possibly from the end of the Paleozoic until the Quaternary. Formation conditions ranging from fresh to brackish and saline waters have been distinguished based on calcite crystal morphologies. The combination of δ 18 O and crystal morphologies show that the fresh-saline water interface has changed considerably over time, and water similar to the present meteoric water and brackish seawater at the site, have most probably earlier been residing in the bedrock. Organic influence and closed system in situ microbial activity causing disequilibrium are indicated by extremely low δ 13 C (down

αA-crystallin R49Cneo mutation influences the architecture of lens fiber cell membranes and causes posterior and nuclear cataracts in mice

Directory of Open Access Journals (Sweden)

Andley Usha P

2009-07-01

Full Text Available Abstract Background αA-crystallin (CRYAA/HSPB4, a major component of all vertebrate eye lenses, is a small heat shock protein responsible for maintaining lens transparency. The R49C mutation in the αA-crystallin protein is linked with non-syndromic, hereditary human cataracts in a four-generation Caucasian family. Methods This study describes a mouse cataract model generated by insertion of a neomycin-resistant (neor gene into an intron of the gene encoding mutant R49C αA-crystallin. Mice carrying the neor gene and wild-type Cryaa were also generated as controls. Heterozygous knock-in mice containing one wild type gene and one mutated gene for αA-crystallin (WT/R49Cneo and homozygous knock-in mice containing two mutated genes (R49Cneo/R49Cneo were compared. Results By 3 weeks, WT/R49Cneo mice exhibited large vacuoles in the cortical region 100 μm from the lens surface, and by 3 months posterior and nuclear cataracts had developed. WT/R49Cneo mice demonstrated severe posterior cataracts at 9 months of age, with considerable posterior nuclear migration evident in histological sections. R49Cneo/R49Cneo mice demonstrated nearly complete lens opacities by 5 months of age. In contrast, R49C mice in which the neor gene was deleted by breeding with CreEIIa mice developed lens abnormalities at birth, suggesting that the neor gene may suppress expression of mutant R49C αA-crystallin protein. Conclusion It is apparent that modification of membrane and cell-cell interactions occurs in the presence of the αA-crystallin mutation and rapidly leads to lens cell pathology in vivo.

Enhancement of crystallinity of cellulose produced by Escherichia coli through heterologous expression of bcsD gene from Gluconacetobacter xylinus.

Science.gov (United States)

Sajadi, Elaheh; Babaipour, Valiollah; Deldar, Ali Asghar; Yakhchali, Bagher; Fatemi, Seyed Safa-Ali

2017-09-01

To evaluate the crystallinity index of the cellulose produced by Escherichia coli Nissle 1917 after heterologous expression of the cellulose synthase subunit D (bcsD) gene of Gluconacetobacter xylinus BPR2001. The bcsD gene of G. xylinus BPR2001 was expressed in E. coli and its protein product was visualized using SDS-PAGE. FTIR analysis showed that the crystallinity index of the cellulose produced by the recombinants was 0.84, which is 17% more than that of the wild type strain. The increased crystallinity index was also confirmed by X-ray diffraction analysis. The cellulose content was not changed significantly after over-expressing the bcsD. The bcsD gene can improve the crystalline structure of the bacterial cellulose but there is not any significant difference between the amounts of cellulose produced by the recombinant and wild type E. coli Nissle 1917.

BAG3 Directly Interacts with Mutated alphaB-Crystallin to Suppress Its Aggregation and Toxicity

NARCIS (Netherlands)

Hishiya, Akinori; Salman, Mortada Najem; Carra, Serena; Kampinga, Harm H.; Takayama, Shinichi

2011-01-01

A homozygous disruption or genetic mutation of the bag3 gene causes progressive myofibrillar myopathy in mouse and human skeletal and cardiac muscle disorder while mutations in the small heat shock protein aB-crystallin gene ( CRYAB) are reported to be responsible for myofibrillar myopathy. Here, we

The make up of crystalline bedrock - crystalline body and blocks

International Nuclear Information System (INIS)

Huber, M.; Huber, A.

1986-01-01

Statements of a geological nature can be made on the basis of investigations of the bedrock exposed in southern Black Forest and these can, in the form of prognoses, be applied to the crystalline Basement of northern Switzerland. Such statements relate to the average proportions of the main lithological groups at the bedrock surface and the surface area of the granite body. Some of the prognoses can be compared and checked with the results from the deep drilling programme in northern Switzerland. Further, analogical interferences from the situation in the southern Black Forest allow predictions to be made on the anticipated block structure of the crystalline Basement. (author)

Pengaruh Kecepatan Pendinginan Terhadap Perubahan Volume Leburan Polymer Crystalline dan Non-Crystalline

OpenAIRE

Fahrurrozi, Mohammad; Moristanto, Bagus Senowulung dan

2003-01-01

AbstractThe study was directed to develop a method to predict the influence of the rate of cooling to the degree of crystallittitv (DOC) and volume change of crystalline polymers. Crystalline polymer melts exhibit volume shrinkage on cooling below melting point due to crystallization. Crystallization and volunrc shrinkage will proceed with varies rate as long as the temperature is above the glass tansition temperatrre. DOC achieved by polymer is not only determined by the inherent crystallini...

Soni-removal of nucleic acids from inclusion bodies.

Science.gov (United States)

Neerathilingam, Muniasamy; Mysore, Sumukh; Gandham, Sai Hari A

2014-05-23

Inclusion bodies (IBs) are commonly formed in Escherichia coli due to over expression of recombinant proteins in non-native state. Isolation, denaturation and refolding of these IBs is generally performed to obtain functional protein. However, during this process IBs tend to form non-specific interactions with sheared nucleic acids from the genome, thus getting carried over into downstream processes. This may hinder the refolding of IBs into their native state. To circumvent this, we demonstrate a methodology termed soni-removal which involves disruption of nucleic acid-inclusion body interaction using sonication; followed by solvent based separation. As opposed to conventional techniques that use enzymes and column-based separations, soni-removal is a cost effective alternative for complete elimination of buried and/or strongly bound short nucleic acid contaminants from IBs. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

The quest for crystalline ion beams

CERN Document Server

Schramm, U; Bussmann, M; Habs, D

2002-01-01

The phase transition of an ion beam into its crystalline state has long been expected to dramatically influence beam dynamics beyond the limitations of standard accelerator physics. Yet, although considerable improvement in beam cooling techniques has been made, strong heating mechanisms inherent to existing high-energy storage rings have prohibited the formation of the crystalline state in these machines up to now. Only recently, laser cooling of low-energy beams in the table-top rf quadrupole storage ring PAaul Laser cooLing Acceleration System (PALLAS) has lead to the experimental realization of crystalline beams. In this article, the quest for crystalline beams as well as their unique properties as experienced in PALLAS will be reviewed.

Biomimetic processing of oriented crystalline ceramic layers

Energy Technology Data Exchange (ETDEWEB)

Cesarano, J.; Shelnutt, J.A.

1997-10-01

The aim of this project was to develop the capabilities for Sandia to fabricate self assembled Langmuir-Blodgett (LB) films of various materials and to exploit their two-dimensional crystalline structure to promote the growth of oriented thin films of inorganic materials at room temperature. This includes the design and synthesis of Langmuir-active (amphiphilic) organic molecules with end groups offering high nucleation potential for various ceramics. A longer range goal is that of understanding the underlying principles, making it feasible to use the techniques presented in this report to fabricate unique oriented films of various materials for electronic, sensor, and membrane applications. Therefore, whenever possible, work completed in this report was completed with the intention of addressing the fundamental phenomena underlying the growth of crystalline, inorganic films on template layers of highly organized organic molecules. This problem was inspired by biological processes, which often produce exquisitely engineered structures via templated growth on polymeric layers. Seashells, for example, exhibit great toughness owing to their fine brick-and-mortar structure that results from templated growth of calcium carbonate on top of layers of ordered organic proteins. A key goal in this work, therefore, is to demonstrate a positive correlation between the order and orientation of the template layer and that of the crystalline ceramic material grown upon it. The work completed was comprised of several parallel efforts that encompassed the entire spectrum of biomimetic growth from solution. Studies were completed on seashells and the mechanisms of growth for calcium carbonate. Studies were completed on the characterization of LB films and the capability developed for the in-house fabrication of these films. Standard films of fatty acids were studied as well as novel polypeptides and porphyrins that were synthesized.

Liquid crystalline dihydroazulene photoswitches

DEFF Research Database (Denmark)

Petersen, Anne Ugleholdt; Jevric, Martyn; Mandle, Richard J.

2015-01-01

A large selection of photochromic dihydroazulene (DHA) molecules incorporating various substituents at position 2 of the DHA core was prepared and investigated for their ability to form liquid crystalline phases. Incorporation of an octyloxy-substituted biphenyl substituent resulted in nematic...... phase behavior and it was possible to convert one such compound partly into its vinylheptafulvene (VHF) isomer upon irradiation with light when in the liquid crystalline phase. This conversion resulted in an increase in the molecular alignment of the phase. In time, the meta-stable VHF returns...... to the DHA where the alignment is maintained. The systematic structural variation has revealed that a biaryl spacer between the DHA and the alkyl chain is needed for liquid crystallinity and that the one aromatic ring in the spacer cannot be substituted by a triazole. This work presents an important step...

Blue Emission in Proteins

OpenAIRE

Sarkar, Sohini; Sengupta, Abhigyan; Hazra, Partha; Mandal, Pankaj

2014-01-01

Recent literatures reported blue-green emission from amyloid fibril as exclusive signature of fibril formation. This unusual visible luminescence is regularly used to monitor fibril growth. Blue-green emission has also been observed in crystalline protein and in solution. However, the origin of this emission is not known exactly. Our spectroscopic study of serum proteins reveals that the blue-green emission is a property of protein monomer. Evidences suggest that semiconductor-like band struc...

Inclusion body myositis.

Science.gov (United States)

Dimachkie, Mazen M; Barohn, Richard J

2014-08-01

The idiopathic inflammatory myopathies (IIMs) are a heterogeneous group of rare disorders that share many similarities. In addition to sporadic inclusion body myositis (IBM), these include dermatomyositis, polymyositis, and autoimmune necrotizing myopathy. IBM is the most common IIM after age 50 years. Muscle histopathology shows endomysial inflammatory exudates surrounding and invading nonnecrotic muscle fibers often accompanied by rimmed vacuoles and protein deposits. It is likely that IBM is has a prominent degenerative component. This article reviews the evolution of knowledge in IBM, with emphasis on recent developments in the field, and discusses ongoing clinical trials. Copyright © 2014 Elsevier Inc. All rights reserved.

Extracellular requirements for the endocytosis of carcinogenic crystalline nickel sulfide particles by facultative phagocytes

International Nuclear Information System (INIS)

Heck, J.D.; Costa, M.

1982-01-01

Various culture medium components were examined for their effect upon the phagocytosis of carcinogenic crystalline and non-carcinogenic amorphous NiS by cultured fibroblastic cells using both a visual and radioactive assay for phagocytosis. Crystalline 63 NiS was phagocytosed by cells in a simple salts/glucose maintenance medium to an extent similar to that observed in complex culture medium fortified with 10% fetal bovine serum (FBS), suggesting that serum proteins and other components in complex culture medium exert little influence upon the uptake of these heavy metal particles. Phagocytosis of crystalline NiS was shown to be highly dependent upon Ca 2+ since omission of Ca 2+ from the salts/glucose medium substantially reduced phagocytosis, while readdition of Ca 2+ stimulated uptake in a concentration-dependent manner. The uptake of the NiS particles was inhibited by trifluoperazine, a calmodulin antagonist, implicating intracellular Ca 2+ in this phagocytosis process. Since the opposite surface charge of crystalline and amorphous NiS has been related to their different phagocytic uptake by cells whose primary function is not phagocytosis (facultative phagocytes), these results show that the culture medium components do not modify the surface charge of these particles in a way that significantly influences their uptake. (Auth.)

Differential metabolism and leakage of protein in an inherited cataract and a normal lens cultured with ouabain

International Nuclear Information System (INIS)

Piatigorsky, J.; Fukui, H.N.; Kinoshita, J.H.

1978-01-01

Ocular lenses in Nakano mice showed marked changes in synthesis, degradation and leakage of protein during cataractogenesis. The cataract-associated changes included the differential lowering of crystalline synthesis, the cleavage of crystallin polypeptides to lower molecular weight forms and the leakage of crystallins from cultured lenses. Ouabain treatment of normal lenses induced these alterations, suggesting that changes in the intracellular levels of Na + and K + affect the anabolism and catabolism of protein during cataract formation. 35 S-methionine was used during the course of the experiments as a method of protein identification. (author)

Amyloid-linked cellular toxicity triggered by bacterial inclusion bodies

International Nuclear Information System (INIS)

Gonzalez-Montalban, Nuria; Villaverde, Antonio; Aris, Anna

2007-01-01

The aggregation of proteins in the form of amyloid fibrils and plaques is the characteristic feature of some pathological conditions ranging from neurodegenerative disorders to systemic amyloidoses. The mechanisms by which the aggregation processes result in cell damage are under intense investigation but recent data indicate that prefibrillar aggregates are the most proximate mediators of toxicity rather than mature fibrils. Since it has been shown that prefibrillar forms of the nondisease-related misfolded proteins are highly toxic to cultured mammalian cells we have studied the cytoxicity associated to bacterial inclusion bodies that have been recently described as protein deposits presenting amyloid-like structures. We have proved that bacterial inclusion bodies composed by a misfolding-prone β-galactosidase fusion protein are clearly toxic for mammalian cells but the β-galactosidase wild type enzyme forming more structured thermal aggregates does not impair cell viability, despite it also binds and enter into the cells. These results are in the line that the most cytotoxic aggregates are early prefibrilar assemblies but discard the hypothesis that the membrane destabilization is Key event to subsequent disruption of cellular processes, such as ion balance, oxidative state and the eventually cell death

Effects of stoichiometry on the transport properties of crystalline phase-change materials.

Science.gov (United States)

Zhang, Wei; Wuttig, Matthias; Mazzarello, Riccardo

2015-09-03

It has recently been shown that a metal-insulator transition due to disorder occurs in the crystalline state of the GeSb2Te4 phase-change compound. The transition is triggered by the ordering of the vacancies upon thermal annealing. In this work, we investigate the localization properties of the electronic states in selected crystalline (GeTe)x-(Sb2Te3)y compounds with varying GeTe content by large-scale density functional theory simulations. In our models, we also include excess vacancies, which are needed to account for the large carrier concentrations determined experimentally. We show that the models containing a high concentration of stoichiometric vacancies possess states at the Fermi energy localized inside vacancy clusters, as occurs for GeSb2Te4. On the other hand, the GeTe-rich models display metallic behavior, which stems from two facts: a) the tail of localized states shrinks due to the low probability of having sizable vacancy clusters, b) the excess vacancies shift the Fermi energy to the region of extended states. Hence, a stoichiometry-controlled metal-insulator transition occurs. In addition, we show that the localization properties obtained by scalar-relativistic calculations with gradient-corrected functionals are unaffected by the inclusion of spin-orbit coupling or the use of hybrid functionals.

Thermodynamics of Crystalline States

CERN Document Server

Fujimoto, Minoru

2010-01-01

Thermodynamics is a well-established discipline of physics for properties of matter in thermal equilibrium surroundings. Applying to crystals, however, the laws encounter undefined properties of crystal lattices, which therefore need to be determined for a clear and well-defined description of crystalline states. Thermodynamics of Crystalline States explores the roles played by order variables and dynamic lattices in crystals in a wholly new way. This book is divided into three parts. The book begins by clarifying basic concepts for stable crystals. Next, binary phase transitions are discussed to study collective motion of order variables, as described mostly as classical phenomena. In the third part, the multi-electron system is discussed theoretically, as a quantum-mechanical example, for the superconducting state in metallic crystals. Throughout the book, the role played by the lattice is emphasized and examined in-depth. Thermodynamics of Crystalline States is an introductory treatise and textbook on meso...

Studies on the site of protein and RNA syntheses in poxvirus-infected cells

Energy Technology Data Exchange (ETDEWEB)

Sakaue, Y [Osaka Univ. (Japan). Research Inst. for Microbial Diseases

1974-04-01

Pulse labelling of short time and the chase of it were conducted to Poxvirus-infected cells using /sup 3/H-uridine and /sup 3/H-leucine with high concentration, and autoradiography (AR) was taken. As the result, protein synthesis, which was in accordance with ''B''-type inclusion, was markedly observed in one-minute labelling at the site of protein synthesis of infected cells. Although the protein synthesis was observed at the peripheral site of ''A''-type inclusion, it was not found within inclusions. However, it was found from the experiment of chase that protein collected markedly within ''B''-type inclusion. They were found that ''B''-type inclusion is the site of Virus DNA synthesis as well as the site of Virus mRNA synthesis, and that it is also absolutely possible for ''B''-type inclusion to synthesize Virus protein. In addition, it was found that ''A''-type inclusion is not the site of synthesis, but newly-synthesized protein.

Studies on the site of protein and RNA syntheses in poxvirus-infected cells

International Nuclear Information System (INIS)

Sakaue, Yoshihiro

1974-01-01

Pulse labelling of short time and the chase of it were conducted to Poxvirus-infected cells using 3 H-uridine and 3 H-leucine with high concentration, and autoradiography (AR) was taken. As the result, protein synthesis, which was in accordance with ''B''-type inclusion, was markedly observed in one-minute labelling at the site of protein synthesis of infected cells. Although the protein synthesis was observed at the peripheral site of ''A''-type inclusion, it was not found within inclusions. However, it was found from the experiment of chase that protein collected markedly within ''B''-type inclusion. They were found that ''B''-type inclusion is the site of Virus DNA synthesis as well as the site of Virus mRNA synthesis, and that it is also absolutely possible for ''B''-type inclusion to synthesize Virus protein. In addition, it was found that ''A''-type inclusion is not the site of synthesis, but newly-synthesized protein. (Ichikawa, K.)

Chemical Assistance in Refolding of Bacterial Inclusion Bodies

Directory of Open Access Journals (Sweden)

Mona Alibolandi

2011-01-01

Full Text Available Escherichia coli is one of the most widely used hosts for the production of recombinant proteins but insoluble expression of heterologous proteins is a major bottleneck in production of recombinant proteins in E. coli. In vitro refolding of inclusion body into proteins with native conformations is a solution for this problem but there is a need for optimization of condition for each protein specifically. Several approaches have been described for in vitro refolding; most of them involve the use of additives for assisting correct folding. Cosolutes play a major role in refolding process and can be classified according to their function as aggregation suppressors and folding enhancers. This paper presents a review of additives that are used in refolding process of insoluble recombinant proteins in small scale and industrial processes.

neutron transmission through crystalline materials

International Nuclear Information System (INIS)

El Mesiry, M.S.

2011-01-01

The aim of the present work is to study the neutron transmission through crystalline materials. Therefore a study of pyrolytic graphite (PG) as a highly efficient selective thermal neutron filter and Iron single crystal as a whole one, as well as the applicability of using their polycrystalline powders as a selective cold neutron filters is given. Moreover, the use of PG and iron single crystal as an efficient neutron monochromator is also investigated. An additive formula is given which allows calculating the contribution of the total neutron cross-section including the Bragg scattering from different )(hkl planes to the neutron transmission through crystalline iron and graphite. The formula takes into account their crystalline form. A computer CFe program was developed in order to provide the required calculations for both poly- and single-crystalline iron. The validity of the CFe program was approved from the comparison of the calculated iron cross-section data with the available experimental ones. The CFe program was also adapted to calculate the reflectivity from iron single crystal when it used as a neutron monochromator The computer package GRAPHITE, developed in Neutron Physics laboratory, Nuclear Research Center, has been used in order to provide the required calculations for crystalline graphite in the neutron energy range from 0.1 meV to 10 eV. A Mono-PG code was added to the computer package GRAPHITE in order to calculate the reflectivity from PG crystal when it used as a neutron monochromator.

Crystalline polymorphism induced by charge regulation in ionic membranes.

Science.gov (United States)

Leung, Cheuk-Yui; Palmer, Liam C; Kewalramani, Sumit; Qiao, Baofu; Stupp, Samuel I; Olvera de la Cruz, Monica; Bedzyk, Michael J

2013-10-08

The crystallization of molecules with polar and hydrophobic groups, such as ionic amphiphiles and proteins, is of paramount importance in biology and biotechnology. By coassembling dilysine (+2) and carboxylate (-1) amphiphiles of various tail lengths into bilayer membranes at different pH values, we show that the 2D crystallization process in amphiphile membranes can be controlled by modifying the competition of long-range and short-range interactions among the polar and the hydrophobic groups. The pH and the hydrophobic tail length modify the intermolecular packing and the symmetry of their crystalline phase. For hydrophobic tail lengths of 14 carbons (C14), we observe the coassembly into crystalline bilayers with hexagonal molecular ordering via in situ small- and wide-angle X-ray scattering. As the tail length increases, the hexagonal lattice spacing decreases due to an increase in van der Waals interactions, as demonstrated by atomistic molecular dynamics simulations. For C16 and C18 we observe a reentrant crystalline phase transition sequence, hexagonal-rectangular-C-rectangular-P-rectangular-C-hexagonal, as the solution pH is increased from 3 to 10.5. The stability of the rectangular phases, which maximize tail packing, increases with increasing tail length. As a result, for very long tails (C22), the possibility of observing packing symmetries other than rectangular-C phases diminishes. Our work demonstrates that it is possible to systematically exchange chemical and mechanical energy by changing the solution pH value within a range of physiological conditions at room temperature in bilayers of molecules with ionizable groups.

Adverse effects of cyclosporine A on HSP25, alpha B-crystallin and myofibrillar cytoskeleton in rat heart

International Nuclear Information System (INIS)

Stacchiotti, Alessandra; Bonomini, Francesca; Lavazza, Antonio; Rodella, Luigi Fabrizio; Rezzani, Rita

2009-01-01

Cyclosporine (CsA) is a universally used immunosuppressive drug which induces adverse side effects in several organs, but its impact on the heart is still controversial. Small heat shock proteins (sHSPs), such as HSP25 and alpha B-crystallin, are cytoprotective stress proteins exceptionally represented in the heart. They act as myofibrillar chaperones that help actin and desmin to maintain their optimum configuration and stability, thereby antagonizing oxidative damage. The present study examined: (1) the cardiac distribution and abundance of HSP25 and alpha B-crystallin in rats receiving CsA at a therapeutic dosage (15 mg/kg/day) for 42 days and 63 days; (2) the presence of myofibrillar proteins, such as actin, alpha-actinin and desmin following the CsA treatments; (3) the subcellular effects of prolonged CsA exposure on the cardiomyocytes by histopathology and transmission electron microscopy. After 63 days CsA intake, sHSPs translocated from a regular sarcomeric pattern to peripheral sarcolemma and intercalated discs, together with actin and desmin. In contrast, the sarcomeric alpha-actinin pattern did not change in all experimental groups. The abundance of actin and HSP25 was unchanged in every time point of treatment while after 63 days CsA, alpha B-crystallin and desmin levels significantly decreased. Furthermore CsA induced fibrosis, irregular sarcomeric alignment and damaged desmosomes. These findings indicate that following prolonged CsA exposure, the cardiac muscle network was affected. In particular, the translocation of sHSPs to intercalated discs merits special consideration as a direct compensatory mechanism to limit CsA cardiotoxicity.

Sodium 4-phenylbutyrate ameliorates the effects of cataract-causing mutant gammaD-crystallin in cultured cells.

Science.gov (United States)

Gong, Bo; Zhang, Li-Yun; Lam, Dennis Shun-Chiu; Pang, Chi-Pui; Yam, Gary Hin-Fai

2010-06-04

gammaD-Crystallin (CRYGD) is a major structural lens crystallin and its mutations result in congenital cataract formation. In this study, we attempted to correct the altered protein features of G165fsX8 CRYGD protein with small chemical molecules. Recombinant FLAG-tagged mutants (R15C, R15S, P24T, R61C, and G165fsX8) of CRYGD were expressed in COS-7 cells and treated with small chemical molecules with reported protein chaperoning properties (sodium 4-phenylbutyrate [4-PBA], trimethylamine N-oxide [TMAO], and glycerol and DMSO [DMSO]). Protein solubility in 0.5% Triton X-100 and subcellular distribution was examined by western blotting and immunofluorescence, respectively. Apoptosis was assayed as the percentage of fragmented nuclei in transfected cells. Expression of heat-shock proteins (Hsp70 and Hsp90) was examined by reverse transcription-polymerase chain reaction analysis. Unlike WT and most mutants (R15C, R15S, P24T, and R61C) of CRYGD, G165fsX8 CRYGD was significantly insoluble in 0.5% Triton X-100. This insolubility was alleviated by dose-dependent 4-PBA treatment. The treatment relieved the mislocalization of G165fsX8 CRYGD from the nuclear envelope. Also, 4-PBA treatment reduced cell apoptosis and caused an upregulation of Hsp70. 4-PBA treatment reduced the defective phenotype of mutant G165fsX8 CRYGD and rescued the affected cells from apoptosis. This could be a potential treatment for lens structural protein and prevent lens opacity in cataract formation.

TDP-43 in Familial and Sporadic Frontotemporal Lobar Degeneration with Ubiquitin Inclusions

NARCIS (Netherlands)

Cairns, Nigel J.; Neumann, Manuela; Bigio, Eileen H.; Holm, Ida E.; Troost, Dirk; Hatanpaa, Kimmo J.; Foong, Chan; White, Charles L.; Schneider, Julie A.; Kretzschmar, Hans A.; Carter, Deborah; Taylor-Reinwald, Lisa; Paulsmeyer, Katherine; Strider, Jeffrey; Gitcho, Michael; Goate, Alison M.; Morris, John C.; Mishrall, Manjari; Kwong, Linda K.; Stieber, Anna; Xu, Yan; Forman, Mark S.; Trojanowski, John Q.; Lee, Virginia M.-Y.; Mackenzie, Ian R. A.

2007-01-01

TAR DNA-binding protein 43 (TDP-43) is a major pathological protein of sporadic and familial frontotemporal lobar degeneration with ubiquitin-positive, tau-negative inclusions (FTLD-U) with or without motor neuron disease (MND). Thus, TDP-43 defines a novel class of neurodegenerative diseases called

Bio-based liquid crystalline polyesters

Science.gov (United States)

Wilsens, Carolus; Rastogi, Sanjay; Dutch Collaboration

2013-03-01

The reported thin-film polymerization has been used as a screening method in order to find bio-based liquid crystalline polyesters with convenient melting temperatures for melt-processing purposes. An in depth study of the structural, morphological and chemical changes occurring during the ongoing polycondensation reactions of these polymers have been performed. Structural and conformational changes during polymerization for different compositions have been followed by time resolved X-ray and Infrared spectroscopy. In this study, bio-based monomers such as vanillic acid and 2,5-furandicarboxylic acid are successfully incorporated in liquid crystalline polyesters and it is shown that bio-based liquid crystalline polymers with high aromatic content and convenient processing temperatures can be synthesized. Special thanks to the Dutch Polymer Institute for financial support

Crystallinity in starch plastics: consequences for material properties

NARCIS (Netherlands)

Soest, van J.J.G.; Vliegenthart, J.F.G.

1997-01-01

The processing of starches with biodegradable additives has made biodegradable plastics suitable for a number of applications. Starch plastics are partially crystalline as a result of residual crystallinity and the recrystallization of amylose and amylopectin. Such crystallinity is a key determinant

COMPOSITION OF FOWLPOX VIRUS AND INCLUSION MATRIX.

Science.gov (United States)

RANDALL, C C; GAFFORD, L G; DARLINGTON, R W; HYDE, J

1964-04-01

Randall, Charles C. (University of Mississippi School of Medicine, Jackson), Lanelle G. Gafford, Robert W. Darlington, and James M. Hyde. Composition of fowlpox virus and inclusion matrix. J. Bacteriol. 87:939-944. 1964.-Inclusion bodies of fowlpox virus infection are especially favorable starting material for the isolation of virus and inclusion matrix. Electron micrographs of viral particles and matrix indicated a high degree of purification. Density-gradient centrifugation of virus in cesium chloride and potassium tartrate was unsatisfactory because of inactivation, and clumping or disintegration. Chemical analyses of virus and matrix revealed significant amounts of lipid, protein, and deoxyribonucleic acid, but no ribonucleic acid or carbohydrate. Approximately 47% of the weight of the virus and 83% of the matrix were extractable in chloroform-methanol. The lipid partitions of the petroleum ether extracts were similar, except that the phospholipid content of the matrix was 2.2 times that of the virus. Viral particles were sensitive to diethyl ether and chloroform.

Co-solute assistance in refolding of recombinant proteins | Gerami ...

African Journals Online (AJOL)

Prokaryotic expression system is the most widely used host for the production of recombinant proteins but inclusion body formation is a major bottleneck in the production of recombinant proteins in prokaryotic cells, especially in Escherichia coli. In vitro refolding of inclusion body into the the proteins with native ...

Nanoscale crystallinity modulates cell proliferation on plasma sprayed surfaces

Energy Technology Data Exchange (ETDEWEB)

Smith, Alan M. [School of Applied Sciences, University of Huddersfield, Huddersfield HD1 3DH (United Kingdom); Paxton, Jennifer Z.; Hung, Yi-Pei; Hadley, Martin J.; Bowen, James; Williams, Richard L. [School of Chemical Engineering, University of Birmingham, Edgbaston, B15 2TT (United Kingdom); Grover, Liam M., E-mail: l.m.grover@bham.ac.uk [School of Chemical Engineering, University of Birmingham, Edgbaston, B15 2TT (United Kingdom)

2015-03-01

Calcium phosphate coatings have been applied to the surface of metallic prostheses to mediate hard and soft tissue attachment for more than 40 years. Most coatings are formed of high purity hydroxyapatite, and coating methods are often designed to produce highly crystalline surfaces. It is likely however, that coatings of lower crystallinity can facilitate more rapid tissue attachment since the surface will exhibit a higher specific surface area and will be considerably more reactive than a comparable highly crystalline surface. Here we test this hypothesis by growing a population of MC3T3 osteoblast-like cells on the surface of two types of hip prosthesis with similar composition, but with differing crystallinity. The surfaces with lower crystallinity facilitated more rapid cell attachment and increased proliferation rate, despite having a less heterogeneous surface topography. This work highlights that the influence of the crystallinity of HA at the nano-scale is dominant over macro-scale topography for cell adhesion and growth. Furthermore, crystallinity could be easily adjusted by without compromising coating purity. These findings could facilitate designing novel coated calcium phosphate surfaces that more rapidly bond tissue following implantation. - Highlights: • Crystallinity of HA at the nano-scale was dominant over macro-scale topography. • Lower crystallinity caused rapid cell attachment and proliferation rate. • Crystallinity could be easily adjusted by without compromising coating purity.

Nanoscale crystallinity modulates cell proliferation on plasma sprayed surfaces

International Nuclear Information System (INIS)

Smith, Alan M.; Paxton, Jennifer Z.; Hung, Yi-Pei; Hadley, Martin J.; Bowen, James; Williams, Richard L.; Grover, Liam M.

2015-01-01

Calcium phosphate coatings have been applied to the surface of metallic prostheses to mediate hard and soft tissue attachment for more than 40 years. Most coatings are formed of high purity hydroxyapatite, and coating methods are often designed to produce highly crystalline surfaces. It is likely however, that coatings of lower crystallinity can facilitate more rapid tissue attachment since the surface will exhibit a higher specific surface area and will be considerably more reactive than a comparable highly crystalline surface. Here we test this hypothesis by growing a population of MC3T3 osteoblast-like cells on the surface of two types of hip prosthesis with similar composition, but with differing crystallinity. The surfaces with lower crystallinity facilitated more rapid cell attachment and increased proliferation rate, despite having a less heterogeneous surface topography. This work highlights that the influence of the crystallinity of HA at the nano-scale is dominant over macro-scale topography for cell adhesion and growth. Furthermore, crystallinity could be easily adjusted by without compromising coating purity. These findings could facilitate designing novel coated calcium phosphate surfaces that more rapidly bond tissue following implantation. - Highlights: • Crystallinity of HA at the nano-scale was dominant over macro-scale topography. • Lower crystallinity caused rapid cell attachment and proliferation rate. • Crystallinity could be easily adjusted by without compromising coating purity

alpha-Crystallin A sequences of Alligator mississippiensis and the lizard Tupinambis teguixin: molecular evolution and reptilian phylogeny.

Science.gov (United States)

de Jong, W W; Zweers, A; Versteeg, M; Dessauer, H C; Goodman, M

1985-11-01

The amino acid sequences of the eye lens protein alpha-crystallin A from many mammalian and avian species, two frog species, and a dogfish have provided detailed information about the molecular evolution of this protein and allowed some useful inferences about phylogenetic relationships among these species. We now have isolated and sequenced the alpha-crystallins of the American alligator and the common tegu lizard. The reptilian alpha A chains appear to have evolved as slowly as those of other vertebrates, i.e., at two to three amino acid replacements per 100 residues in 100 Myr. The lack of charged replacements and the general types and distribution of replacements also are similar to those in other vertebrate alpha A chains. Maximum-parsimony analyses of the total data set of 67 vertebrate alpha A sequences support the monophyletic origin of alligator, tegu, and birds and favor the grouping of crocodilians and birds as surviving sister groups in the subclass Archosauria.

Protein Crystals as Novel Catalytic Materials.

Science.gov (United States)

Margolin, Alexey L.; Navia, Manuel A.

2001-06-18

In this era of molecular biology, protein crystallization is often considered to be a necessary first step in obtaining structural information through X-ray diffraction analysis. In a different light, protein crystals can also be thought of as materials, whose chemical and physical properties make them broadly attractive and useful across a larger spectrum of disciplines. The full potential of these protein crystalline materials has been severely restricted in practice, however, both by their inherent fragility, and by strongly held skepticism concerning their routine and predictable growth, formulation, and practical application. Fortunately, these problems have turned out to be solvable. A systematic exploration of the biophysics and biochemistry of protein crystallization has shown that one can dependably create new protein crystalline materials more or less at will. In turn, these crystals can be readily strengthened, both chemically and mechanically, to make them suitable for practical commercialization. Today, these novel materials are used as industrial catalysts on a commercial scale, in bioremediation and "green chemistry" applications, and in enantioselective chromatography of pharmaceuticals and fine chemicals. In the near future, their utility will expand, to include the purification of protein drugs, formulation of direct protein therapeutics, and development of adjuvant-less vaccines.

Mechanical dispersion in fractured crystalline rock systems

International Nuclear Information System (INIS)

Lafleur, D.W.; Raven, K.G.

1986-12-01

This report compiles and evaluates the hydrogeologic parameters describing the flow of groundwater and transport of solutes in fractured crystalline rocks. This report describes the processes of mechanical dispersion in fractured crystalline rocks, and compiles and evaluates the dispersion parameters determined from both laboratory and field tracer experiments. The compiled data show that extrapolation of the reliable test results performed over intermediate scales (10's of m and 10's to 100's of hours) to larger spatial and temporal scales required for performance assessment of a nuclear waste repository in crystalline rock is not justified. The reliable measures of longitudinal dispersivity of fractured crystalline rock are found to range between 0.4 and 7.8 m

Accumulation of glycation products in. cap alpha. -H pig lens crystallin and its bearing to diabetic cataract genesis

Energy Technology Data Exchange (ETDEWEB)

Vidal, P; Cabezas-Cerrato, J

1988-01-01

The incorporation of /sup 11/C-glucose in native pig crystalline by in vitro incubation was found, after subsequent dialysis, to affect all 5 classes of crystallin separated by Sepharose CL-6B column chromatography. Though the radioactivity of the ..cap alpha..-H fraction was three times greater than that of any of the others, autoradiographs of SDS-PAGE gels showed /sup 11/C-glucose adducts to be present in all soluble protein subunits, without there being any evidence of preferential glycation of the ..cap alpha..-H subunits. The concentration of stable glycation products in the ..cap alpha..-H chromatographic fraction of soluble crystallins is suggested to be due the addition of glycated material to this fraction as result of glycation-induced hyperaggregation, and not because the ..cap alpha..-H subunits were especially susceptible to glycation.

Crystallinity and mechanical effects from annealing Parylene thin films

Energy Technology Data Exchange (ETDEWEB)

Jackson, Nathan, E-mail: Nathan.Jackson@tyndall.ie [Tyndall National Institute, University College Cork, Cork (Ireland); Stam, Frank; O' Brien, Joe [Tyndall National Institute, University College Cork, Cork (Ireland); Kailas, Lekshmi [University of Limerick, Limerick (Ireland); Mathewson, Alan; O' Murchu, Cian [Tyndall National Institute, University College Cork, Cork (Ireland)

2016-03-31

Parylene is commonly used as thin film polymer for MEMS devices and smart materials. This paper investigates the impact on bulk properties due to annealing various types of Parylene films. A thin film of Parylene N, C and a hybrid material consisting of Parylene N and C were deposited using a standard Gorham process. The thin film samples were annealed at varying temperatures from room temperature up to 300 °C. The films were analyzed to determine the mechanical and crystallinity effects due to different annealing temperatures. The results demonstrate that the percentage of crystallinity and the full-width-half-maximum value on the 2θ X-ray diffraction scan increases as the annealing temperature increases until the melting temperature of the Parylene films was achieved. Highly crystalline films of 85% and 92% crystallinity were achieved for Parylene C and N respectively. Investigation of the hybrid film showed that the individual Parylene films behave independently to each other, and the crystallinity of one film had no significant impact to the other film. Mechanical testing showed that the elastic modulus and yield strength increase as a function of annealing, whereas the elongation-to-break parameter decreases. The change in elastic modulus was more significant for Parylene C than Parylene N and this is attributed to the larger change in crystallinity that was observed. Parylene C had a 112% increase in crystallinity compared to a 61% increase for Parylene N, because the original Parylene N material was more crystalline than Parylene C so the change of crystallinity was greater for Parylene C. - Highlights: • A hybrid material consisting of Parylene N and C was developed. • Parylene N has greater crystallinity than Parylene C. • Phase transition of Parylene N due to annealing results in increased crystallinity. • Annealing caused increased crystallinity and elastic modulus in Parylene films. • Annealed hybrid Parylene films crystallinity behave

Frequency of nuclear mutant huntingtin inclusion formation in neurons and glia is cell-type-specific

NARCIS (Netherlands)

Jansen, Anne H. P.; van Hal, Maurik; Op den Kelder, Ilse C.; Meier, Romy T.; de Ruiter, Anna-Aster; Schut, Menno H.; Smith, Donna L.; Grit, Corien; Brouwer, Nieske; Kamphuis, Willem; Boddeke, H. W. G. M.; den Dunnen, Wilfred F. A.; van Roon, Willeke M. C.; Bates, Gillian P.; Hol, Elly M.; Reits, Eric A.

2017-01-01

Huntington's disease (HD) is an autosomal dominant inherited neurodegenerative disorder that is caused by a CAG expansion in the Huntingtin (HTT) gene, leading to HTT inclusion formation in the brain. The mutant huntingtin protein (mHTT) is ubiquitously expressed and therefore nuclear inclusions

Frequency of Nuclear Mutant Huntingtin Inclusion Formation in Neurons and Glia is Cell-Type-Specific

NARCIS (Netherlands)

Jansen, Anne H P; van Hal, Maurik; op den Kelder, Ilse C.; Meier, Romy T.; de Ruiter, Anna-Aster; Schut, Menno H.; Smith, Donna L.; Grit, Corien; Brouwer, Nieske; Kamphuis, Willem; Boddeke, H. W. G. M.; den Dunnen, Wilfred F. A.; van Roon, Willeke M. C.; Bates, Gillian P.; Hol, Elly M.; Reits, Eric A.

Huntington's disease (HD) is an autosomal dominant inherited neurodegenerative disorder that is caused by a CAG expansion in the Huntingtin (HTT) gene, leading to HTT inclusion formation in the brain. The mutant huntingtin protein (mHTT) is ubiquitously expressed and therefore nuclear inclusions

Identification of Protein-Protein Interactions with Glutathione-S-Transferase (GST) Fusion Proteins.

Science.gov (United States)

Einarson, Margret B; Pugacheva, Elena N; Orlinick, Jason R

2007-08-01

INTRODUCTIONGlutathione-S-transferase (GST) fusion proteins have had a wide range of applications since their introduction as tools for synthesis of recombinant proteins in bacteria. GST was originally selected as a fusion moiety because of several desirable properties. First and foremost, when expressed in bacteria alone, or as a fusion, GST is not sequestered in inclusion bodies (in contrast to previous fusion protein systems). Second, GST can be affinity-purified without denaturation because it binds to immobilized glutathione, which provides the basis for simple purification. Consequently, GST fusion proteins are routinely used for antibody generation and purification, protein-protein interaction studies, and biochemical analysis. This article describes the use of GST fusion proteins as probes for the identification of protein-protein interactions.

Photodamaging mechanisms of the eye structure: the aggregates appearance at UV-illumination of lens proteins

International Nuclear Information System (INIS)

El'chaninov, V.V.; Fedorovich, I.B.

1989-01-01

UV-light injury of individual crystallines (water soluble proteins of the cattle eye crystalline lens) were studied by SDS PSSG technique. Photodamage resulted in olygomer formation. The appearance of high molecular aggregates with the molecular mass as large as 10 5 D were seen in all fractions of the crystalline

Differential Precipitation and Solubilization of Proteins.

Science.gov (United States)

Ryan, Barry J; Kinsella, Gemma K

2017-01-01

Differential protein precipitation is a rapid and economical step in protein purification and is based on exploiting the inherent physicochemical properties of the polypeptide. Precipitation of recombinant proteins, lysed from the host cell, is commonly used to concentrate the protein of choice before further polishing steps with more selective purification columns (e.g., His-Tag, Size Exclusion, etc.). Recombinant proteins can also precipitate naturally as inclusion bodies due to various influences during overexpression in the host cell. Although this phenomenon permits easier initial separation from native proteins, these inclusion bodies must carefully be differentially solubilized so as to reform functional, correctly folded proteins. Here, appropriate bioinformatics tools to aid in understanding a protein's propensity to aggregate and solubilize are explored as a backdrop for a typical protein extraction, precipitation, and selective resolubilization procedure, based on a recombinantly expressed protein.

Determination of crystallinity of ceramic materials from the Ruland Method

International Nuclear Information System (INIS)

Kniess, C.T.; Prates, P.B.; Gomes Junior, J.C.; Lima, J.C. de; Riella, H.G.; Kuhnen, N.C.

2011-01-01

Some methods found in literature approach the different characteristics between crystalline and amorphous phases by X ray diffraction technique. These methods use the relation between the intensities of the crystalline peaks and background amorphous or the absolute intensity of one of these to determine the relative amount of crystalline and amorphous material. However, a crystalline substance presents shows coherent diffuse scattering and a loss in the intensity of the peaks of diffraction in function of thermal vibrations of atoms and imperfections in the crystalline structure. A correct method for the determination of the crystallinity must take in account these effects. This work has as objective to determine the crystallinity of ceramic materials obtained with the addition of mineral coal bottom ashes, using the X ray diffraction technique and the Ruland Method, that considers the diminution of the intensity of the crystalline peak because of the disorder affects. The Ruland Method shows adequate for the determination of the crystallinity of the ceramic materials. (author)

Protein surface shielding agents in protein crystallization

International Nuclear Information System (INIS)

Hašek, J.

2011-01-01

The crystallization process can be controlled by protein surface shielding agents blocking undesirable competitive adhesion modes during non-equilibrium processes of deposition of protein molecules on the surface of growing crystalline blocks. The hypothesis is based on a number of experimental proofs from diffraction experiments and also retrieved from the Protein Data Bank. The molecules adhering temporarily on the surface of protein molecules change the propensity of protein molecules to deposit on the crystal surface in a definite position and orientation. The concepts of competitive adhesion modes and protein surface shielding agents acting on the surface of molecules in a non-equilibrium process of protein crystallization provide a useful platform for the control of crystallization. The desirable goal, i.e. a transient preference of a single dominating adhesion mode between protein molecules during crystallization, leads to uniform deposition of proteins in a crystal. This condition is the most important factor for diffraction quality and thus also for the accuracy of protein structure determination. The presented hypothesis is a generalization of the experimentally well proven behaviour of hydrophilic polymers on the surface of protein molecules of other compounds

Hydroimidazolone modification of the conserved Arg12 in small heat shock proteins: studies on the structure and chaperone function using mutant mimics.

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Ram H Nagaraj

Full Text Available Methylglyoxal (MGO is an α-dicarbonyl compound present ubiquitously in the human body. MGO reacts with arginine residues in proteins and forms adducts such as hydroimidazolone and argpyrimidine in vivo. Previously, we showed that MGO-mediated modification of αA-crystallin increased its chaperone function. We identified MGO-modified arginine residues in αA-crystallin and found that replacing such arginine residues with alanine residues mimicked the effects of MGO on the chaperone function. Arginine 12 (R12 is a conserved amino acid residue in Hsp27 as well as αA- and αB-crystallin. When treated with MGO at or near physiological concentrations (2-10 µM, R12 was modified to hydroimidazolone in all three small heat shock proteins. In this study, we determined the effect of arginine substitution with alanine at position 12 (R12A to mimic MGO modification on the structure and chaperone function of these proteins. Among the three proteins, the R12A mutation improved the chaperone function of only αA-crystallin. This enhancement in the chaperone function was accompanied by subtle changes in the tertiary structure, which increased the thermodynamic stability of αA-crystallin. This mutation induced the exposure of additional client protein binding sites on αA-crystallin. Altogether, our data suggest that MGO-modification of the conserved R12 in αA-crystallin to hydroimidazolone may play an important role in reducing protein aggregation in the lens during aging and cataract formation.

Generic Crystalline Disposal Reference Case

Energy Technology Data Exchange (ETDEWEB)

Painter, Scott Leroy [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Chu, Shaoping [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Harp, Dylan Robert [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Perry, Frank Vinton [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Wang, Yifeng [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

2015-02-20

A generic reference case for disposal of spent nuclear fuel and high-level radioactive waste in crystalline rock is outlined. The generic cases are intended to support development of disposal system modeling capability by establishing relevant baseline conditions and parameters. Establishment of a generic reference case requires that the emplacement concept, waste inventory, waste form, waste package, backfill/buffer properties, EBS failure scenarios, host rock properties, and biosphere be specified. The focus in this report is on those elements that are unique to crystalline disposal, especially the geosphere representation. Three emplacement concepts are suggested for further analyses: a waste packages containing 4 PWR assemblies emplaced in boreholes in the floors of tunnels (KBS-3 concept), a 12-assembly waste package emplaced in tunnels, and a 32-assembly dual purpose canister emplaced in tunnels. In addition, three failure scenarios were suggested for future use: a nominal scenario involving corrosion of the waste package in the tunnel emplacement concepts, a manufacturing defect scenario applicable to the KBS-3 concept, and a disruptive glaciation scenario applicable to both emplacement concepts. The computational approaches required to analyze EBS failure and transport processes in a crystalline rock repository are similar to those of argillite/shale, with the most significant difference being that the EBS in a crystalline rock repository will likely experience highly heterogeneous flow rates, which should be represented in the model. The computational approaches required to analyze radionuclide transport in the natural system are very different because of the highly channelized nature of fracture flow. Computational workflows tailored to crystalline rock based on discrete transport pathways extracted from discrete fracture network models are recommended.

Improvement of broiler meat quality due to dietary inclusion of soybean oligosaccharide derived from soybean meal extract

Science.gov (United States)

Suthama, N.; Pramono, Y. B.; Sukamto, B.

2018-01-01

Dietary inclusion of antibiotics as growth promoters (AGPs) in poultry production has been applied for decades worldwide, but recently AGPs have been banned due to the negative consequences for health and food safety. Soybean oligosccharide (SOS) derived from soybean meal extract is one of natural compound without carrying-over the residue to product and is consumer’s health friendly. The purpose of the present study was to evaluate dietary inclusion of SOS on broiler meat quality. A total of 120 broilers of 7-day-old were allocated into 3 treatments with 4 replications (10 birds each) in completely randomized design. Treatments applied were D1: diet without SOS, D2: D1 plus 0.15% SOS, and D3: D1 plus 0.30% SOS. Intestinal lactic acid bacteria (LAB), protein digestibility, meat protein and fat depositions, and meat cholesterol were the parameters observed. Data were statistically tested using analysis of variance and Duncan test. Dietary SOS inclusion at 0.30% (D3) significantly (P<0.05) increased LAB population (7.21x104 cfu/g), protein digestibility (72.80%), and meat protein deposition (90.83 g/bird), but it decreased meat fat (8.27 g/bird) and meat cholesterol (37.28 mg/100 g). In conclusion, dietary SOS inclusion at 0.30% improves meat quality of broiler based on the increase in meat protein deposition with lower fat and cholesterol.

Semiclassical and quantum polarons in crystalline acetanilide

Science.gov (United States)

Hamm, P.; Tsironis, G. P.

2007-08-01

Crystalline acetanilide is a an organic solid with peptide bond structure similar to that of proteins. Two states appear in the amide I spectral region having drastically different properties: one is strongly temperature dependent and disappears at high temperatures while the other is stable at all temperatures. Experimental and theoretical work over the past twenty five years has assigned the former to a selftrapped state while the latter to an extended free exciton state. In this article we review the experimental and theoretical developments on acetanilide paying particular attention to issues that are still pending. Although the interpretation of the states is experimentally sound, we find that specific theoretical comprehension is still lacking. Among the issues that that appear not well understood is the effective dimensionality of the selftrapped polaron and free exciton states.

Effects of alkaline or liquid-ammonia treatment on crystalline cellulose: changes in crystalline structure and effects on enzymatic digestibility

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Himmel Michael E

2011-10-01

Full Text Available Abstract Background In converting biomass to bioethanol, pretreatment is a key step intended to render cellulose more amenable and accessible to cellulase enzymes and thus increase glucose yields. In this study, four cellulose samples with different degrees of polymerization and crystallinity indexes were subjected to aqueous sodium hydroxide and anhydrous liquid ammonia treatments. The effects of the treatments on cellulose crystalline structure were studied, in addition to the effects on the digestibility of the celluloses by a cellulase complex. Results From X-ray diffractograms and nuclear magnetic resonance spectra, it was revealed that treatment with liquid ammonia produced the cellulose IIII allomorph; however, crystallinity depended on treatment conditions. Treatment at a low temperature (25°C resulted in a less crystalline product, whereas treatment at elevated temperatures (130°C or 140°C gave a more crystalline product. Treatment of cellulose I with aqueous sodium hydroxide (16.5 percent by weight resulted in formation of cellulose II, but also produced a much less crystalline cellulose. The relative digestibilities of the different cellulose allomorphs were tested by exposing the treated and untreated cellulose samples to a commercial enzyme mixture (Genencor-Danisco; GC 220. The digestibility results showed that the starting cellulose I samples were the least digestible (except for corn stover cellulose, which had a high amorphous content. Treatment with sodium hydroxide produced the most digestible cellulose, followed by treatment with liquid ammonia at a low temperature. Factor analysis indicated that initial rates of digestion (up to 24 hours were most strongly correlated with amorphous content. Correlation of allomorph type with digestibility was weak, but was strongest with cellulose conversion at later times. The cellulose IIII samples produced at higher temperatures had comparable crystallinities to the initial cellulose I

Mineralization of alpha-1-antitrypsin inclusion bodies in Mmalton alpha-1-antitrypsin deficiency.

Science.gov (United States)

Callea, Francesco; Giovannoni, Isabella; Francalanci, Paola; Boldrini, Renata; Faa, Gavino; Medicina, Daniela; Nobili, Valerio; Desmet, Valeer J; Ishak, Kamal; Seyama, Kuniaki; Bellacchio, Emanuele

2018-05-16

Alpha-1-antitrypsin (AAT) deficiency (AATD) of Z, Mmalton, Siiyama type is associated with liver storage of the mutant proteins and liver disease. The Z variant can be diagnosed on isoelectric focusing (IEF) while Mmalton and Siiyama may be missed or misdiagnosed with this technique. Therefore, molecular analysis is mandatory for their characterization. In particular, that holds true for the Mmalton variant as on IEF profile it resembles the wild M2 subtype. This is a retrospective analysis involving review of medical records and of liver biopsy specimens from a series of Mmalton, Z and Siiyama Alpha-1-antitrypsin deficiency patients. The review has been implemented by additional histological stains, electron microscopic observations and 3-D modeling studies of the sites of the mutations. Z, Mmalton and Siiyama liver specimen contained characteristic intrahepatocytic PAS-D globules. The globules differed in the three variants as only Mmalton cases showed dark basophilic precipitates within the AAT inclusions. The precipitates were visualized in haematoxylin-eosin (H.E.) stained preparations and corresponded to calcium precipitates as demonstrated by von Kossa staining. On immunohistochemistry, ZAAT inclusions were stained by polyclonal as well as monoclonal noncommercial anti-AAT antibody (AZT11), whilst Mmalton and Siiyama inclusion bodies remained negative with the monoclonal anti-Z antibody. 3-D protein analysis allowed to predict more severe misfolding of the Mmalton molecule as compared to Z and Siiyama that could trigger anomalous interaction with endoplasmic reticulum chaperon proteins, namely calcium binding proteins. Mmalton AAT inclusion bodies contain calcium precipitates inside them that allow the differential diagnosis with Siiyama and ZAAT inclusions in routine histological sections. The study has confirmed the specificity of the monoclonal AZT11 for the Z mutant. Thus, the combination of these two features is crucial for the distinction between the

A Histone-Like Protein Induces Plasmid DNA to Form Liquid Crystals in Vitro and Gene Compaction in Vivo

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Shiyong Sun

2013-12-01

Full Text Available The liquid crystalline state is a universal phenomenon involving the formation of an ordered structure via a self-assembly process that has attracted attention from numerous scientists. In this study, the dinoflagellate histone-like protein HCcp3 is shown to induce super-coiled pUC18 plasmid DNA to enter a liquid crystalline state in vitro, and the role of HCcp3 in gene condensation in vivo is also presented. The plasmid DNA (pDNA-HCcp3 complex formed birefringent spherical particles with a semi-crystalline selected area electronic diffraction (SAED pattern. Circular dichroism (CD titrations of pDNA and HCcp3 were performed. Without HCcp3, pUC18 showed the characteristic B conformation. As the HCcp3 concentration increased, the 273 nm band sharply shifted to 282 nm. When the HCcp3 concentration became high, the base pair (bp/dimer ratio fell below 42/1, and the CD spectra of the pDNA-HCcp3 complexes became similar to that of dehydrated A-form DNA. Microscopy results showed that HCcp3 compacted the super-coiled gene into a condensed state and that inclusion bodies were formed. Our results indicated that HCcp3 has significant roles in gene condensation both in vitro and in histone-less eukaryotes in vivo. The present study indicates that HCcp3 has great potential for applications in non-viral gene delivery systems, where HCcp3 may compact genetic material to form liquid crystals.

Confronting New Demands : Inclusive Growth, Inclusive Trade ...

International Development Research Centre (IDRC) Digital Library (Canada)

Confronting New Demands : Inclusive Growth, Inclusive Trade. Policymakers, businesspeople and civil society advocates need evidence-based research to react ... understood implications, such as labour standards and intellectual property; ...

Interaction of amyloid inhibitor proteins with amyloid beta peptides: insight from molecular dynamics simulations.

Directory of Open Access Journals (Sweden)

Payel Das

Full Text Available Knowledge of the detailed mechanism by which proteins such as human αB- crystallin and human lysozyme inhibit amyloid beta (Aβ peptide aggregation is crucial for designing treatment for Alzheimer's disease. Thus, unconstrained, atomistic molecular dynamics simulations in explicit solvent have been performed to characterize the Aβ17-42 assembly in presence of the αB-crystallin core domain and of lysozyme. Simulations reveal that both inhibitor proteins compete with inter-peptide interaction by binding to the peptides during the early stage of aggregation, which is consistent with their inhibitory action reported in experiments. However, the Aβ binding dynamics appear different for each inhibitor. The binding between crystallin and the peptide monomer, dominated by electrostatics, is relatively weak and transient due to the heterogeneous amino acid distribution of the inhibitor surface. The crystallin-bound Aβ oligomers are relatively long-lived, as they form more extensive contact surface with the inhibitor protein. In contrast, a high local density of arginines from lysozyme allows strong binding with Aβ peptide monomers, resulting in stable complexes. Our findings not only illustrate, in atomic detail, how the amyloid inhibitory mechanism of human αB-crystallin, a natural chaperone, is different from that of human lysozyme, but also may aid de novo design of amyloid inhibitors.

Developing a financial inclusion index and inclusive growth in India

OpenAIRE

Susanta Kumar SETHY

2016-01-01

Financial inclusion is one of the systems through which Inclusive Growth can be achieved in developing countries like India where large sections are unable or hopeless to contribute in the financial system. An inclusive financial system mobilizes more resources for productive purposes leading to higher economic growth, better opportunities and reduction of poverty. This study, proposed an Index of financial inclusion – a multidimensional measure. The Financial Inclusion Index c...

Autokinase activity of alpha-crystallin inhibits its specific interaction with the DOTIS element in the murine gamma D/E/F-crystallin promoter in vitro.

Science.gov (United States)

Pietrowski, D; Graw, J

1997-10-01

In a previous report we demonstrated the in vitro interaction of alpha-crystallin with an element downstream of the transcriptional initiation site (DOTIS) of the murine gamma E-crystallin promoter (Pietrowski et al., 1994, Gene 144, 171-178). The aim of the present study was to investigate the influence of phosphorylation on this particular interaction. We could demonstrate that the autophosphorylation of alpha-crystallin leads to a complete loss of interaction with the DOTIS element, however, PKA-dependent phosphorylation of alpha-crystallin is without effect on the interaction. It is hypothesized that the autophosphorylation of alpha-crystallin might be involved in regulatory mechanisms of the murine gamma D/E/F-crystallin gene expression.

Protein folding and protein metallocluster studies using synchrotron small angler X-ray scattering

International Nuclear Information System (INIS)

Eliezer, D.

1994-06-01

Proteins, biological macromolecules composed of amino-acid building blocks, possess unique three dimensional shapes or conformations which are intimately related to their biological function. All of the information necessary to determine this conformation is stored in a protein's amino acid sequence. The problem of understanding the process by which nature maps protein amino-acid sequences to three-dimensional conformations is known as the protein folding problem, and is one of the central unsolved problems in biophysics today. The possible applications of a solution are broad, ranging from the elucidation of thousands of protein structures to the rational modification and design of protein-based drugs. The scattering of X-rays by matter has long been useful as a tool for the characterization of physical properties of materials, including biological samples. The high photon flux available at synchrotron X-ray sources allows for the measurement of scattering cross-sections of dilute and/or disordered samples. Such measurements do not yield the detailed geometrical information available from crystalline samples, but do allow for lower resolution studies of dynamical processes not observable in the crystalline state. The main focus of the work described here has been the study of the protein folding process using time-resolved small-angle x-ray scattering measurements. The original intention was to observe the decrease in overall size which must accompany the folding of a protein from an extended conformation to its compact native state. Although this process proved too fast for the current time-resolution of the technique, upper bounds were set on the probable compaction times of several small proteins. In addition, an interesting and unexpected process was detected, in which the folding protein passes through an intermediate state which shows a tendency to associate. This state is proposed to be a kinetic molten globule folding intermediate

Inhibition of Fusion of Chlamydia trachomatis Inclusions at 32°C Correlates with Restricted Export of IncA

OpenAIRE

Fields, K. A.; Fischer, E.; Hackstadt, T.

2002-01-01

Chlamydia trachomatis is an obligate intracellular bacterium that develops within a parasitophorous vacuole termed an inclusion. The inclusion is nonfusogenic with lysosomes but intercepts lipids from a host cell exocytic pathway. Initiation of chlamydial development is concurrent with modification of the inclusion membrane by a set of C. trachomatis-encoded proteins collectively designated Incs. One of these Incs, IncA, is functionally associated with the homotypic fusion of inclusions. Incl...

Access to gram scale amounts of functional globular adiponectin from E. coli inclusion bodies by alkaline-shock solubilization.

Science.gov (United States)

Heiker, John T; Klöting, Nora; Blüher, Matthias; Beck-Sickinger, Annette G

2010-07-16

The adipose tissue derived protein adiponectin exerts anti-diabetic, anti-inflammatory and anti-atherosclerotic effects. Adiponectin serum concentrations are in the microgram per milliliter range in healthy humans and inversely correlate with obesity and metabolic disorders. Accordingly, raising circulating adiponectin levels by direct administration may be an intriguing strategy in the treatment of obesity-related metabolic disorders. However production of large amounts of recombinant adiponectin protein is a primary obstacle so far. Here, we report a novel method for large amount production of globular adiponectin from E. coli inclusion bodies utilizing an alkaline-shock solubilization method without chaotropic agents followed by precipitation of the readily renaturing protein. Precipitation of the mildly solubilized protein capitalizes on advantages of inclusion body formation. This approach of inclusion body protein recovery provides access to gram scale amounts of globular adiponectin with standard laboratory equipment avoiding vast dilution or dialysis steps to neutralize the pH and renature the protein, thus saving chemicals and time. The precipitated protein is readily renaturing in buffer, is of adequate purity without a chromatography step and shows biological activity in cultured MCF7 cells and significantly lowered blood glucose levels in mice with streptozotocin induced type 1 diabetes. Copyright 2010 Elsevier Inc. All rights reserved.

What Is Crystalline Silica?

Science.gov (United States)

... and ceramic manufacturing and the tool and die, steel and foundry industries. Crystalline silica is used in manufacturing, household abrasives, adhesives, paints, soaps, and glass. Additionally, ...

A γA-Crystallin Mouse Mutant Secc with Small Eye, Cataract and Closed Eyelid.

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Man Hei Cheng

Full Text Available Cataract is the most common cause of visual loss in humans. A spontaneously occurred, autosomal dominant mouse mutant Secc, which displayed combined features of small eye, cataract and closed eyelid was discovered in our laboratory. In this study, we identified the mutation and characterized the cataract phenotype of this novel Secc mutant. The Secc mutant mice have eyelids that remain half-closed throughout their life. The mutant lens has a significant reduction in size and with opaque spots clustered in the centre. Histological analysis showed that in the core region of the mutant lens, the fiber cells were disorganized and clefts and vacuoles were observed. The cataract phenotype was evident from new born stage. We identified the Secc mutation by linkage analysis using whole genome microsatellite markers and SNP markers. The Secc locus was mapped at chromosome 1 flanked by SNPs rs3158129 and rs13475900. Based on the chromosomal position, the candidate cataract locus γ-crystallin gene cluster (Cryg was investigated by sequencing. A single base deletion (299delG in exon 3 of Cryga which led to a frame-shift of amino acid sequence from position 91 was identified. As a result of this mutation, the sequences of the 3rd and 4th Greek-key motifs of the γA-crystallin are replaced with an unrelated C-terminal peptide of 75 residues long. Coincidentally, the point mutation generated a HindIII restriction site, allowing the identification of the CrygaSecc mutant allele by RFLP. Western blot analysis of 3-week old lenses showed that the expression of γ-crystallins was reduced in the CrygaSecc mutant. Furthermore, in cell transfection assays using CrygaSecc mutant cDNA expression constructs in 293T, COS-7 and human lens epithelial B3 cell lines, the mutant γA-crystallins were enriched in the insoluble fractions and appeared as insoluble aggregates in the transfected cells. In conclusion, we have demonstrated that the Secc mutation leads to the

Characterization of crystalline structures in Opuntia ficus-indica

OpenAIRE

Contreras-Padilla, Margarita; Rivera-Muñoz, Eric M.; Gutiérrez-Cortez, Elsa; del López, Alicia Real; Rodríguez-García, Mario Enrique

2014-01-01

This research studies the crystalline compounds present in nopal (Opuntia ficus-indica) cladodes. The identification of the crystalline structures was performed using X-ray diffraction, scanning electron microscopy, mass spectrometry, and Fourier transform infrared spectroscopy. The crystalline structures identified were calcium carbonate (calcite) [CaCO3], calcium-magnesium bicarbonate [CaMg(CO3)2], magnesium oxide [MgO], calcium oxalate monohydrate [Ca(C2O4)•(H2O)], potassium peroxydiphosph...

United States Crystalline Repository Project - key research areas

International Nuclear Information System (INIS)

Patera, E.S.

1986-01-01

The Crystalline Repository Project is responsible for siting the second high-level nuclear waste repository in crystalline rock for the US Department of Energy. A methodology is being developed to define data and information needs and a way to evaluate that information. The areas of research the Crystalline Repository Project is involved in include fluid flow in a fractured network, coupled thermal, chemical and flow processes and cooperation in other nations and OECD research programs

Oligomerization and chaperone-like activity of Drosophila melanogaster small heat shock protein DmHsp27 and three arginine mutants in the alpha-crystallin domain.

Science.gov (United States)

Moutaoufik, Mohamed Taha; Morrow, Geneviève; Maaroufi, Halim; Férard, Céline; Finet, Stéphanie; Tanguay, Robert M

2017-07-01

The small Hsp DmHsp27 from Drosophila melanogaster is one of the few small heat shock proteins (sHsps) found within the nucleus. We report that its dimerization is independent of disulfide bond formation and seems to rely on salt bridges. Unlike metazoan sHsps, DmHsp27 forms two populations of oligomers not in equilibrium. Mutations at highly conserved arginine residues in mammalian sHsps have been reported to be associated with protein conformational defects and intracellular aggregation. Independent mutation of three highly conserved arginines (R122, R131, and R135) to glycine in DmHsp27 results in only one population of higher molecular weight form. In vitro, the chaperone-like activity of wild-type DmHsp27 was comparable with that of its two isolated populations and to the single population of the R122G, R131G, and R135G using luciferase as substrate. However, using insulin, the chaperone-like activity of wild-type DmHsp27 was lower than that of R122G and R131G mutants. Altogether, the results characterize wild-type DmHsp27 and its alpha-crystallin domain (ACD) arginine mutants and may give insight into protection mechanism of sHsps.

Irreducible tensor operators and crystalline potentials

International Nuclear Information System (INIS)

Boutron, F.; Saint-James, D.

1961-01-01

It is often accepted that the effects of its neighbourhood on the quantum state of an ion A may be obtained by the model of the crystalline effective field approximation. Within this assumption Stevens has developed a method which provides equivalent operators that facilitate the calculation of the matrix elements of the crystalline field in a given multiplicity. This method has been extended here. We demonstrate that in the expansion of the crystalline field in powers of the electrons coordinates of the ion A - for electrons of the same sub-shell of A - only even terms can contribute. Equivalent operators and matrix elements, in a given multiplicity, are given for these development terms - up to order 6 - and for potential invariant by the operations of one of the thirty-two point-groups. (author) [fr

Ultrastructure of inclusion bodies in annulus cells in the degenerating human intervertebral disc.

Science.gov (United States)

Gruber, H E; Hanley, E N

2009-06-01

The rough endoplasmic reticulum (rER) of the cell has an architectural editing function that checks whether protein structure and three-dimensional assembly have occurred properly prior to export of newly synthesized material out of the cell. If these have been faulty, the material is retained within the rER as an inclusion body. Inclusion bodies have been identified previously in chondrocytes and osteoblasts in chondrodysplasias and osteogenesis imperfecta. Inclusion bodies in intervertebral disc cells, however, have only recently been recognized. Our objectives were to use transmission electron microscopy to analyze more fully inclusion bodies in the annulus pulposus and to study the extracellular matrix (ECM) surrounding cells containing inclusion bodies. ECM frequently encapsulated cells with inclusion bodies, and commonly contained prominent banded aggregates of Type VI collagen. Inclusion body material had several morphologies, including relatively smooth, homogeneous material, or a rougher, less homogeneous feature. Such findings expand our knowledge of the fine structure of the human disc cell and ECM during disc degeneration, and indicate the potential utility of ultrastructural identification of discs with intracellular inclusion bodies as a screening method for molecular studies directed toward identification of defective gene products in degenerating discs.

Effect of the essential amino acids upon inclusion in vitro of 14C-phenylalanine and 14C-leucine in the protein of mammary gland

International Nuclear Information System (INIS)

Alexandrov, S.; Ivanov, N.; Sirakov, L.

1983-01-01

It is admitted that the essential amino acids could be divided into two groups, depending on the need of them for synthesis of milk protein: group i - amino acids, which are absorbed in quantities precisely corresponding to their content in milk protein (methionine, phenyl-alanine, histidine, thyrosine and triptophane), and group ii - amino acids, which are absorbed in quantities greater than their content in milk protein and which, because of this, could fullfil other metabolic functions in the mammary gland (threonine, valine, isoleucine, lysine and arginine). According to this concept, tissue slices of lactating mammary gland of guinea-pigs were incubated in the presence of grour i or group ii essential amino acids. Slices were incubated for 60 min at 37+-0.5 deg C, In a Crebs-Ringer phosphate buffer plus 0.2 glucose and 3.7 KBq/ml incubation medium DL-(I- 14 C)-phenylalanine or L-(U- 14 C)-leucine and their incorporation in the tissue proteins of mammary gland was measured in vitro. Group ii essential amino acids provoked significantly more intensive (P<0.001) inclusion in protein synthesis of these labelled amino acids in the tissue of mammary gland, as compared with group i essential amino acids

Crystalline morphology of the matrix of PEEK-carbon fiber aromatic polymer composites. I. Assessment of crystallinity

International Nuclear Information System (INIS)

Blundell, D.J.; Chalmers, J.M.; Mackenzie, M.W.; Gaskin, W.F.

1985-01-01

The crystallinity of the polyetheretherketone (PEEK) matrix polymer in the Aromatic Polymer Composite APC-2 has been estimated using a combination of techniques based on wide angle x-ray diffraction and infrared reflection spectroscopy. Crystallinity varies systematically with cooling rate and annealing time over the range 20 to 40%. The occurrence of oriented crystal growth of the PEEK relative to the carbon fiber can be monitored by x-ray diffraction. 8 references, 10 figures, 1 table

Irradiation sterilization of semi-crystalline polymers

International Nuclear Information System (INIS)

Williams, J.; Dunn, T.; Stannett, V.

1978-01-01

A semi-crystalline polymer such as polypropylene, is sterilized by high energy irradiation, with the polymer containing a non-crystalline mobilizing additive which increases the free volume of the polymer, to prevent embrittlement of the polymer during and subsequent to the irradiation. The additive has a density of from 0.6 to 1.9 g/cm 3 and a molecular weight from 100 to 10,000 g/mole

Crystalline structure of metals

International Nuclear Information System (INIS)

Holas, A.

1972-01-01

An attempt is made to find the crystalline structure of metals on the basis of the existing theory of metals. The considerations are limited to the case of free crystals, that is, not subjected to any stresses and with T=0. The energy of the crystal lattice has been defined and the dependence of each term on structures and other properties of metals has been described. The energy has been used to find the values of crystalline structure parameters as the values at which the energy has an absolute minimum. The stability of the structure has been considered in cases of volume changes and shearing deformations. A semiqualitative description has been obtained which explains characteristic properties of one-electron metals. (S.B.)

Approaching Inclusion as Social Practice: Processes of Inclusion and Exclusion

DEFF Research Database (Denmark)

Molbæk, Mette; Hansen, Janne Hedegaard; Lassen, Mikkel

2018-01-01

The article presents the results of a review of international research investigating mechanisms and processes of inclusion and exclusion as an ongoing part of social practice in a school context. The review forms part of a research project investigating the social practices of inclusive education...... in primary and lower-secondary education (age 6–16) in public schools as constituted by processes of inclusion and exclusion. The project aims to shift the scientific focus of research in inclusive education from the development of pedagogical and didactic practice to the importance of community construction...... through inclusion and exclusion processes. The project arises in context of Danish education policy, while the review looked for international research findings on the limits between inclusion and exclusion: how they are drawn, by whom, for what reasons, and for whose benefit? On the background...

Quantum vibrational polarons: Crystalline acetanilide revisited

Science.gov (United States)

Hamm, Peter; Edler, Julian

2006-03-01

We discuss a refined theoretical description of the peculiar spectroscopy of crystalline acetanilide (ACN). Acetanilide is a molecular crystal with quasi-one-dimensional chains of hydrogen-bonded units, which is often regarded as a model system for the vibrational spectroscopy of proteins. In linear spectroscopy, the CO stretching (amide I) band of ACN features a double-peak structure, the lower of which shows a pronounced temperature dependence which has been discussed in the context of polaron theory. In nonlinear spectroscopy, both of these peaks respond distinctly differently. The lower-frequency band exhibits the anharmonicity expected from polaron theory, while the higher-frequency band responds as if it were quasiharmonic. We have recently related the response of the higher-frequency band to that of a free exciton [J. Edler and P. Hamm, J. Chem. Phys. 117, 2415 (2002)]. However, as discussed in the present paper, the free exciton is not an eigenstate of the full quantum version of the Holstein polaron Hamiltonian, which is commonly used to describe these phenomena. In order to resolve this issue, we present a numerically exact solution of the Holstein polaron Hamiltonian in one dimension (1D) and 3D. In 1D, we find that the commonly used displaced oscillator picture remains qualitatively correct, even for relatively large exciton coupling. However, the result is not in agreement with the experiment, as it fails to explain the free-exciton band. In contrast, when taking into account the 3D nature of crystalline acetanilide, certain parameter regimes exist where the displaced oscillator picture breaks down and states appear in the spectrum that indeed exhibit the characteristics of a free exciton. The appearance of these states is a speciality of vibrational polarons, whose source of exciton coupling is transition dipole coupling which is expected to have opposite signs of interchain and intrachain coupling.

Limits to Inclusion

Science.gov (United States)

Hansen, Janne Hedegaard

2012-01-01

In this article, I will argue that a theoretical identification of the limit to inclusion is needed in the conceptual identification of inclusion. On the one hand, inclusion is formulated as a vision that is, in principle, limitless. On the other hand, there seems to be an agreement that inclusion has a limit in the pedagogical practice. However,…

Inclusive Business - What It Is All About? Managing Inclusive Companies

Directory of Open Access Journals (Sweden)

Tea Golja

2012-01-01

Full Text Available Following the challenges we face today, the inclusive business models are future business models through which the Millennium Development Goals can be fostered and strengthen. These are the models which, through their strategic orientation on inclusivity, include low income communities in their value chain. This can be done through combining variety of strategies which all have two common points – recognition of stakeholders and adjustment of the product to the target market. The paper presents the analysis of inclusive markets. Hence, the research results show the dispersion of inclusive businesses worldwide, type of the organization, sector coverage, and contribution to MDGs as well as the particular way of inclusion of low income communities in their value chain. The aim is to present how inclusive business benefits not only the low income societies, but the companies that operate in this way as well.

Effect of graded inclusion of dietary soybean meal on nutrient digestibility, health, and metabolic indices of adult dogs.

Science.gov (United States)

Menniti, M F; Davenport, G M; Shoveller, A K; Cant, J P; Osborne, V R

2014-05-01

Two studies were conducted using adult dogs to evaluate the effect of increasing the inclusion of soybean meal (SBM) in an adult dog food on body composition, hematological and biochemical blood analyses, and total tract nutrient digestibility. Nutritionally complete and balanced diets were formulated with commercial-grade SBM (48% CP) to replace 0, 10, 20, or 30% of the protein provided by dried chicken protein resulting in final SBM inclusion of 0, 6.0, 11.5, and 17.0% (as-fed basis), respectively. In study 1, diets were fed during a 24-wk feeding trial using 36 female (spayed), adult hounds to evaluate food intake, BW, body composition, and blood measurements. There were no diet-related differences in food intake or BW. Body composition responded in a quadratic manner to increased dietary SBM inclusion with the percentage (%) of lean mass responding positively (P dogs. Serum concentrations of C-reactive protein and IGF-1 were similar among diets. In study 2, diets were evaluated in a digestibility study using 12 adult dogs in a 4 × 4 Latin square design. Increased SBM inclusion was associated with linear increases in the digestibility of CP (P dogs.

Designing Inclusive Systems Designing Inclusion for Real-world Applications

CERN Document Server

Clarkson, John; Robinson, Peter; Lazar, Jonathan; Heylighen, Ann

2012-01-01

The Cambridge Workshops on Universal Access and Assistive Technology (CWUAAT) are a series of workshops held at a Cambridge University College every two years. The workshop theme: “Designing inclusion for real-world applications” refers to the emerging potential and relevance of the latest generations of inclusive design thinking, tools, techniques, and data, to mainstream project applications such as healthcare and the design of working environments. Inclusive Design Research involves developing tools and guidance enabling product designers to design for the widest possible population, for a given range of capabilities. There are five main themes: •Designing for the Real-World •Measuring Demand And Capabilities •Designing Cognitive Interaction with Emerging Technologies •Design for Inclusion •Designing Inclusive Architecture In the tradition of CWUAAT, we have solicited and accepted contributions over a wide range of topics, both within individual themes and also across the workshop’s scope. ...

Trypanosoma cruzi Epimastigotes Are Able to Store and Mobilize High Amounts of Cholesterol in Reservosome Lipid Inclusions

Science.gov (United States)

Pereira, Miria G.; Nakayasu, Ernesto S.; Sant'Anna, Celso; De Cicco, Nuccia N. T.; Atella, Georgia C.; de Souza, Wanderley; Almeida, Igor C.; Cunha-e-Silva, Narcisa

2011-01-01

Background Reservosomes are lysosome-related organelles found in Trypanosoma cruzi epimastigotes. They represent the last step in epimastigote endocytic route, accumulating a set of proteins and enzymes related to protein digestion and lipid metabolism. The reservosome matrix contains planar membranes, vesicles and lipid inclusions. Some of the latter may assume rectangular or sword-shaped crystalloid forms surrounded by a phospholipid monolayer, resembling the cholesterol crystals in foam cells. Methodology/Principal Findings Using Nile Red fluorimetry and fluorescence microscopy, as well as electron microscopy, we have established a direct correlation between serum concentration in culture medium and the presence of crystalloid lipid inclusions. Starting from a reservosome purified fraction, we have developed a fractionation protocol to isolate lipid inclusions. Gas-chromatography mass-spectrometry (GC-MS) analysis revealed that lipid inclusions are composed mainly by cholesterol and cholesterol esters. Moreover, when the parasites with crystalloid lipid-loaded reservosomes were maintained in serum free medium for 48 hours the inclusions disappeared almost completely, including the sword shaped ones. Conclusions/Significance Taken together, our results suggest that epimastigote forms of T. cruzi store high amounts of neutral lipids from extracellular medium, mostly cholesterol or cholesterol esters inside reservosomes. Interestingly, the parasites are able to disassemble the reservosome cholesterol crystalloid inclusions when submitted to serum starvation. PMID:21818313

Multi-crystalline II-VI based multijunction solar cells and modules

Science.gov (United States)

Hardin, Brian E.; Connor, Stephen T.; Groves, James R.; Peters, Craig H.

2015-06-30

Multi-crystalline group II-VI solar cells and methods for fabrication of same are disclosed herein. A multi-crystalline group II-VI solar cell includes a first photovoltaic sub-cell comprising silicon, a tunnel junction, and a multi-crystalline second photovoltaic sub-cell. A plurality of the multi-crystalline group II-VI solar cells can be interconnected to form low cost, high throughput flat panel, low light concentration, and/or medium light concentration photovoltaic modules or devices.

Controlling the morphology of side chain liquid crystalline block copolymer thin films through variations in liquid crystalline content.

Science.gov (United States)

Verploegen, Eric; Zhang, Tejia; Jung, Yeon Sik; Ross, Caroline; Hammond, Paula T

2008-10-01

In this paper, we describe methods for manipulating the morphology of side-chain liquid crystalline block copolymers through variations in the liquid crystalline content. By systematically controlling the covalent attachment of side chain liquid crystals to a block copolymer (BCP) backbone, the morphology of both the liquid crystalline (LC) mesophase and the phase-segregated BCP microstructures can be precisely manipulated. Increases in LC functionalization lead to stronger preferences for the anchoring of the LC mesophase relative to the substrate and the intermaterial dividing surface. By manipulating the strength of these interactions, the arrangement and ordering of the ultrathin film block copolymer nanostructures can be controlled, yielding a range of morphologies that includes perpendicular and parallel cylinders, as well as both perpendicular and parallel lamellae. Additionally, we demonstrate the utilization of selective etching to create a nanoporous liquid crystalline polymer thin film. The unique control over the orientation and order of the self-assembled morphologies with respect to the substrate will allow for the custom design of thin films for specific nanopatterning applications without manipulation of the surface chemistry or the application of external fields.

[Representation and mathematical analysis of human crystalline lens].

Science.gov (United States)

Tălu, Stefan; Giovanzana, Stefano; Tălu, Mihai

2011-01-01

The surface of human crystalline lens can be described and analyzed using mathematical models based on parametric representations, used in biomechanical studies and 3D solid modeling of the lens. The mathematical models used in lens biomechanics allow the study and the behavior of crystalline lens on variables and complex dynamic loads. Also, the lens biomechanics has the potential to improve the results in the development of intraocular lenses and cataract surgery. The paper presents the most representative mathematical models currently used for the modeling of human crystalline lens, both optically and biomechanically.

Crystallinity and flux pinning properties of MgB2 bulks

International Nuclear Information System (INIS)

Yamamoto, A.; Shimoyama, J.; Ueda, S.; Katsura, Y.; Iwayama, I.; Horii, S.; Kishio, K.

2006-01-01

The relationship between flux pinning properties and crystallinity of MgB 2 bulks was systematically studied. Improved flux pinning properties under high fields were observed for samples with low crystallinity. Increased impurity scattering due to strain and defects in lattice corresponding to the degraded crystallinity was considered to enhance flux pinning strength at grain boundaries. Low-temperature synthesis and carbon substitution were confirmed to be effective for degrading crystallinity of MgB 2 bulks, resulting in high critical current properties under high fields

Novel ATPase activity of the polyprotein intermediate, Viral Protein genome-linked-Nuclear Inclusion-a protease, of Pepper vein banding potyvirus

International Nuclear Information System (INIS)

Mathur, Chhavi; Savithri, Handanahal S.

2012-01-01

Highlights: ► Pepper vein banding potyvirus VPg harbors Walker motifs. ► VPg exhibits ATPase activity in the presence of NIa-Pro. ► Plausible structural and functional interplay between VPg and NIa-Pro. ► Functional relevance of prolonged presence of VPg-Pro during infection. -- Abstract: Potyviruses temporally regulate their protein function by polyprotein processing. Previous studies have shown that VPg (Viral Protein genome-linked) of Pepper vein banding virus interacts with the NIa-Pro (Nuclear Inclusion-a protease) domain, and modulates the kinetics of the protease. In the present study, we report for the first time that VPg harbors the Walker motifs A and B, and the presence of NIa-Pro, especially in cis (cleavage site (E191A) VPg-Pro mutant), is essential for manifestation of the ATPase activity. Mutation of Lys47 (Walker motif A) and Asp88:Glu89 (Walker motif B) to alanine in E191A VPg-Pro lead to reduced ATPase activity, confirming that this activity was inherent to VPg. We propose that potyviral VPg, established as an intrinsically disordered domain, undergoes plausible structural alterations upon interaction with globular NIa-Pro which induces the ATPase activity.

Solution processed nanogap organic diodes based on liquid crystalline materials

Science.gov (United States)

Wang, Yi-Fei; Iino, Hiroaki; Hanna, Jun-ichi

2017-09-01

Co-planar nanogap organic diodes were fabricated with smectic liquid crystalline materials of the benzothienobenzothiophene (BTBT) derivative by a spin-coating technique. A high rectification ratio of the order of 106 at ±3 V was achieved when a liquid crystalline material of 2,7-didecyl benzothieno[3,2-b][1]benzothiophene (10-BTBT-10) was used in a device configuration of Al/10-BTBT-10/pentafluorobenzenethiol-treated Au on a glass substrate, which was 4 orders higher than that of the device based on non-liquid crystalline materials of 2,7-dibutyl benzothieno[3,2-b][1]benzothiophene (4-BTBT-4) and BTBT. Similar results were also observed when another liquid crystalline material of ω, ω'-dioctylterthiophene (8-TTP-8) and a non-liquid crystalline material of terthiophene (TTP) were used. These improved rectifications can be ascribed to the self-assembly properties and controllable molecular orientation of liquid crystalline materials, which made uniform perpendicular oriented polycrystalline films favorable for superior charge transport in nano-channels.

Biocompatibility of crystalline opal nanoparticles.

Science.gov (United States)

Hernández-Ortiz, Marlen; Acosta-Torres, Laura S; Hernández-Padrón, Genoveva; Mendieta, Alicia I; Bernal, Rodolfo; Cruz-Vázquez, Catalina; Castaño, Victor M

2012-10-22

Silica nanoparticles are being developed as a host of biomedical and biotechnological applications. For this reason, there are more studies about biocompatibility of silica with amorphous and crystalline structure. Except hydrated silica (opal), despite is presents directly and indirectly in humans. Two sizes of crystalline opal nanoparticles were investigated in this work under criteria of toxicology. In particular, cytotoxic and genotoxic effects caused by opal nanoparticles (80 and 120 nm) were evaluated in cultured mouse cells via a set of bioassays, methylthiazolyldiphenyl-tetrazolium-bromide (MTT) and 5-bromo-2'-deoxyuridine (BrdU). 3T3-NIH cells were incubated for 24 and 72 h in contact with nanocrystalline opal particles, not presented significant statistically difference in the results of cytotoxicity. Genotoxicity tests of crystalline opal nanoparticles were performed by the BrdU assay on the same cultured cells for 24 h incubation. The reduction of BrdU-incorporated cells indicates that nanocrystalline opal exposure did not caused unrepairable damage DNA. There is no relationship between that particles size and MTT reduction, as well as BrdU incorporation, such that the opal particles did not induce cytotoxic effect and genotoxicity in cultured mouse cells.

Graphene on insulating crystalline substrates

International Nuclear Information System (INIS)

Akcoeltekin, S; El Kharrazi, M; Koehler, B; Lorke, A; Schleberger, M

2009-01-01

We show that it is possible to prepare and identify ultra-thin sheets of graphene on crystalline substrates such as SrTiO 3 , TiO 2 , Al 2 O 3 and CaF 2 by standard techniques (mechanical exfoliation, optical and atomic force microscopy). On the substrates under consideration we find a similar distribution of single layer, bilayer and few-layer graphene and graphite flakes as with conventional SiO 2 substrates. The optical contrast C of a single graphene layer on any of those substrates is determined by calculating the optical properties of a two-dimensional metallic sheet on the surface of a dielectric, which yields values between C = -1.5% (G/TiO 2 ) and C = -8.8% (G/CaF 2 ). This contrast is in reasonable agreement with experimental data and is sufficient to make identification by an optical microscope possible. The graphene layers cover the crystalline substrate in a carpet-like mode and the height of single layer graphene on any of the crystalline substrates as determined by atomic force microscopy is d SLG = 0.34 nm and thus much smaller than on SiO 2 .

UVA photolysis using the protein-bound sensitizers present in human lens

International Nuclear Information System (INIS)

Ortwerth, B.J.; Olesen, P.R.

1994-01-01

This research was undertaken to demonstrate that the protein-bound chromophores in aged human lens can act as sensitizers for protein damage by UVA light. The water-insoluble (WI) proteins from pooled human and bovine lenses were solubilized by sonication in water and illuminated with UV light similar in output to that transmitted by the cornea. Analysis of the irradiated proteins showed a linear decrease in sulfhydryl groups with a 30% loss after 2 h. No loss was seen when native α-crystallin was irradiated under the same conditions. A 25% loss of histidine residues was also observed with the human lens WI fraction, and sodium dodecyl sulfate polyacrylamide gels indicated considerable protein cross-linking. Similar photodamage was seen with a WI fraction from old bovine lenses. While the data show the presence of UVA sensitizers, some histidine destruction and protein cross-linking were also obtained with α-crystallin and with lysozyme which argue that part of the histidine loss in the human WISS was likely due to tryptophan acting as a sensitizer. (Author)

Diffractometric method for determining the degree of crystallinity of materials

Energy Technology Data Exchange (ETDEWEB)

Chukhchin, D. G., E-mail: dimatsch@mail.ru; Malkov, A. V.; Tyshkunova, I. V.; Mayer, L. V.; Novozhilov, E. V. [Lomonosov Northen (Arctic) Federal University (Russian Federation)

2016-05-15

A new method for determining the degree of crystallinity of a material from X-ray diffraction data has been developed. The method is based on estimating the rate of change in function I = f(2θ) in the entire range of scattering angles. A calculation is performed using the ratio of the integral modulus of the first derivative of intensity with respect to angle 2θ to the integral area under the diffraction pattern curve. The method was tested on two substances with known amorphous and crystalline components. A linear relationship is revealed between the specified ratio of crystalline and amorphous parts and the calculated crystallinity index. The proposed method allows one to estimate impartially and compare the degree of crystallinity for samples of different nature.

Early hydration of portland cement with crystalline mineral additions

International Nuclear Information System (INIS)

Rahhal, V.; Talero, R.

2005-01-01

This research presents the effects of finely divided crystalline mineral additions (quartz and limestone), commonly known as filler, on the early hydration of portland cements with very different mineralogical composition. The used techniques to study the early hydration of blended cements were conduction calorimeter, hydraulicity (Fratini's test), non-evaporable water and X-ray diffraction. Results showed that the stimulation and the dilution effects increase when the percentage of crystalline mineral additions used is increased. Depending on the replacement proportion, the mineralogical cement composition and the type of crystalline addition, at 2 days, the prevalence of the dilution effect or the stimulation effect shows that crystalline mineral additions could act as sites of heat dissipation or heat stimulation, respectively

MB109 as bioactive human bone morphogenetic protein-9 refolded and purified from E. coli inclusion bodies

Science.gov (United States)

2014-01-01

Background The development of chemical refolding of transforming growth factor-beta (TGF-β) superfamily ligands has been instrumental to produce the recombinant proteins for biochemical studies and exploring the potential of protein therapeutics. The osteogenic human bone morphogenetic protein-2 (hBMP-2) and its Drosophila DPP homolog were the early successful cases of refolding into functional form. Despite the similarity in their three dimensional structure and amino acid sequences, several other TGF-β superfamily ligands could not be refolded readily by the same methods. Results Here, we report a comprehensive study on the variables of a rapid-dilution refolding method, including the concentrations of protein, salt, detergent and redox agents, pH, refolding duration and the presence of aggregation suppressors and host-cell contaminants, in order to identify the optimal condition to refold human BMP-9 (hBMP-9). To produce a recombinant form of hBMP-9 in E. coli cells, a synthetic codon-optimized gene was designed to encode the mature domain of hBMP-9 (Ser320 – Arg429) directly behind the first methionine, which we herein referred to as MB109. An effective purification scheme was also developed to purify the refolded MB109 to homogeneity with a final yield of 7.8 mg from 100 mg of chromatography-purified inclusion bodies as a starting material. The chemically refolded MB109 binds to ALK1, ActRIIb and BMPRII receptors with relatively high affinity as compared to other Type I and Type II receptors based on surface plasmon resonance analysis. Smad1-dependent luciferase assay in C2C12 cells shows that the MB109 has an EC50 of 0.61 ng/mL (25 pM), which is nearly the same as hBMP-9. Conclusion MB109 is prone to be refolded as non-functional dimer and higher order multimers in most of the conditions tested, but bioactive MB109 dimer can be refolded with high efficiency in a narrow window, which is strongly dependent on the pH, refolding duration, the presence of

Thermal gradient brine inclusion migration in salt study: gas-liquid inclusions, preliminary model

International Nuclear Information System (INIS)

Olander, D.R.; Machiels, A.J.

1979-10-01

Natural salt deposits contain small cubical inclusions of brine distributed through the salt. Temperature gradients, resulting from storing heat-generating wastes in the salt, can cause the inclusions to move through the salt. Prediction of the rate and amount of brine-inclusion migration is necessary for the evaluation of bedded or domed salts as possible media for waste repositories. Inclusions filled exclusively with liquid migrate up the temperature gradient towards the heat source. The solubility of salt in the brine inclusion increases with temperature. Consequently, salt dissolves into the inclusion across the hot surface and crystallizes out at the cold surface. Diffusion of salt within the liquid phase from the hot to the cold faces causes the inclusions to move in the opposite direction. In so doing, they change shape and eventually become rectangular parallelipipeds with a width (dimension perpendicular to the thermal gradient) much larger than the thickness (dimension in the direction of the thermal gradient). The inclusions may also contain a gas phase predominantly consisting of water vapor. These entities are termed two-phase or gas-liquid inclusions. The two-phase inclusions usually migrate down the temperature gradient away from the heat source remaining more-or-less cubical. A two-phase inclusion also forms when an all-liquid inclusion reaches the waste package; upon opening up at the salt-package interface, the brine partially evaporates and the inclusion reseals with some insoluble gas trapped inside. These gas-liquid inclusions proceed to move down the temperature gradient, in the opposite sense of the all-liquid inclusions. The gas-liquid inclusions phenomenon provides a pathway by which radionuclides leached from the wasteform by the brine can be transported away from the waste package and thus might have greater access to the biosphere

Oligomeric structure and chaperone-like activity of Drosophila melanogaster mitochondrial small heat shock protein Hsp22 and arginine mutants in the alpha-crystallin domain.

Science.gov (United States)

Dabbaghizadeh, Afrooz; Finet, Stéphanie; Morrow, Genevieve; Moutaoufik, Mohamed Taha; Tanguay, Robert M

2017-07-01

The structure and chaperone function of DmHsp22WT, a small Hsp of Drosophila melanogaster localized within mitochondria were examined. Mutations of conserved arginine mutants within the alpha-crystallin domain (ACD) domain (R105G, R109G, and R110G) were introduced, and their effects on oligomerization and chaperone function were assessed. Arginine to glycine mutations do not induce significant changes in tryptophan fluorescence, and the mutated proteins form oligomers that are of equal or smaller size than the wild-type protein. They all form oligomer with one single peak as determined by size exclusion chromatography. While all mutants demonstrate the same efficiency as the DmHsp22WT in a DTT-induced insulin aggregation assay, all are more efficient chaperones to prevent aggregation of malate dehydrogenase. Arginine mutants of DmHsp22 are efficient chaperones to retard aggregation of CS and Luc. In summary, this study shows that mutations of arginine to glycine in DmHsp22 ACD induce a number of structural changes, some of which differ from those described in mammalian sHsps. Interestingly, only the R110G-DmHsp22 mutant, and not the expected R109G equivalent to human R140-HspB1, R116-HspB4, and R120-HspB5, showed different structural properties compared with the DmHsp22WT.

Isolation and Characterization of Two Cellulose Morphology Mutants of Gluconacetobacter hansenii ATCC23769 Producing Cellulose with Lower Crystallinity

Science.gov (United States)

Deng, Ying; Nagachar, Nivedita; Fang, Lin; Luan, Xin; Catchmark, Jeffrey M.; Tien, Ming; Kao, Teh-hui

2015-01-01

Gluconacetobacter hansenii, a Gram-negative bacterium, produces and secrets highly crystalline cellulose into growth medium, and has long been used as a model system for studying cellulose synthesis in higher plants. Cellulose synthesis involves the formation of β-1,4 glucan chains via the polymerization of glucose units by a multi-enzyme cellulose synthase complex (CSC). These glucan chains assemble into ordered structures including crystalline microfibrils. AcsA is the catalytic subunit of the cellulose synthase enzymes in the CSC, and AcsC is required for the secretion of cellulose. However, little is known about other proteins required for the assembly of crystalline cellulose. To address this question, we visually examined cellulose pellicles formed in growth media of 763 individual colonies of G. hansenii generated via Tn5 transposon insertion mutagenesis, and identified 85 that produced cellulose with altered morphologies. X-ray diffraction analysis of these 85 mutants identified two that produced cellulose with significantly lower crystallinity than wild type. The gene disrupted in one of these two mutants encoded a lysine decarboxylase and that in the other encoded an alanine racemase. Solid-state NMR analysis revealed that cellulose produced by these two mutants contained increased amounts of non-crystalline cellulose and monosaccharides associated with non-cellulosic polysaccharides as compared to the wild type. Monosaccharide analysis detected higher percentages of galactose and mannose in cellulose produced by both mutants. Field emission scanning electron microscopy showed that cellulose produced by the mutants was unevenly distributed, with some regions appearing to contain deposition of non-cellulosic polysaccharides; however, the width of the ribbon was comparable to that of normal cellulose. As both lysine decarboxylase and alanine racemase are required for the integrity of peptidoglycan, we propose a model for the role of peptidoglycan in the

Isolation and characterization of two cellulose morphology mutants of Gluconacetobacter hansenii ATCC23769 producing cellulose with lower crystallinity.

Directory of Open Access Journals (Sweden)

Ying Deng

Full Text Available Gluconacetobacter hansenii, a Gram-negative bacterium, produces and secrets highly crystalline cellulose into growth medium, and has long been used as a model system for studying cellulose synthesis in higher plants. Cellulose synthesis involves the formation of β-1,4 glucan chains via the polymerization of glucose units by a multi-enzyme cellulose synthase complex (CSC. These glucan chains assemble into ordered structures including crystalline microfibrils. AcsA is the catalytic subunit of the cellulose synthase enzymes in the CSC, and AcsC is required for the secretion of cellulose. However, little is known about other proteins required for the assembly of crystalline cellulose. To address this question, we visually examined cellulose pellicles formed in growth media of 763 individual colonies of G. hansenii generated via Tn5 transposon insertion mutagenesis, and identified 85 that produced cellulose with altered morphologies. X-ray diffraction analysis of these 85 mutants identified two that produced cellulose with significantly lower crystallinity than wild type. The gene disrupted in one of these two mutants encoded a lysine decarboxylase and that in the other encoded an alanine racemase. Solid-state NMR analysis revealed that cellulose produced by these two mutants contained increased amounts of non-crystalline cellulose and monosaccharides associated with non-cellulosic polysaccharides as compared to the wild type. Monosaccharide analysis detected higher percentages of galactose and mannose in cellulose produced by both mutants. Field emission scanning electron microscopy showed that cellulose produced by the mutants was unevenly distributed, with some regions appearing to contain deposition of non-cellulosic polysaccharides; however, the width of the ribbon was comparable to that of normal cellulose. As both lysine decarboxylase and alanine racemase are required for the integrity of peptidoglycan, we propose a model for the role of

Crystalline beam ground state

International Nuclear Information System (INIS)

Wei, Jie; Li, Xiao-Ping

1993-01-01

In order to employ molecular dynamics (MD) methods, commonly used in condensed matter physics, we have derived the equations of motion for a beam of charged particles in the rotating rest frame of the reference particle. We include in the formalism that the particles are confined by the guiding and focusing magnetic fields, and that they are confined in a conducting vacuum pipe while interacting with each other via a Coulomb force. Numerical simulations using MD methods has been performed to obtain the equilibrium crystalline beam structure. The effect of the shearing force, centrifugal force, and azimuthal variation of the focusing strength are investigated. It is found that a constant gradient storage ring can not give a crystalline beam, but that an alternating-gradient (AG) structure can. In such a machine the ground state is, except for one-dimensional (1-D) crystals, time dependent. The ground state is a zero entropy state, despite the time-dependent, periodic variation of the focusing force. The nature of the ground state, similar to that found by Schiffer et al. depends upon the density and the relative focusing strengths in the transverse directions. At low density, the crystal is 1-D. As the density increases, it transforms into various kinds of 2-D and 3-D crystals. If the energy of the beam is higher than the transition energy of the machine, the crystalline structure can not be formed for lack of radial focusing

Inclusive Education in Italy: Description and Reflections on Full Inclusion

Science.gov (United States)

Anastasiou, Dimitris; Kauffman, James M.; Di Nuovo, Santo

2015-01-01

Inclusion of students with disabilities when appropriate is an important goal of special education for students with special needs. Full inclusion, meaning no education for any child in a separate setting, is held to be desirable by some, and Italy is likely the nation with an education system most closely approximating full inclusion on the…

Automated solid-state NMR resonance assignment of protein microcrystals and amyloids

Energy Technology Data Exchange (ETDEWEB)

Schmidt, Elena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany); Gath, Julia [ETH Zurich, Physical Chemistry (Switzerland); Habenstein, Birgit [UMR 5086 CNRS/Universite de Lyon 1, Institut de Biologie et Chimie des Proteines (France); Ravotti, Francesco; Szekely, Kathrin; Huber, Matthias [ETH Zurich, Physical Chemistry (Switzerland); Buchner, Lena [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany); Boeckmann, Anja, E-mail: a.bockmann@ibcp.fr [UMR 5086 CNRS/Universite de Lyon 1, Institut de Biologie et Chimie des Proteines (France); Meier, Beat H., E-mail: beme@ethz.ch [ETH Zurich, Physical Chemistry (Switzerland); Guentert, Peter, E-mail: guentert@em.uni-frankfurt.de [Goethe University Frankfurt am Main, Center for Biomolecular Magnetic Resonance, Institute of Biophysical Chemistry (Germany)

2013-07-15

Solid-state NMR is an emerging structure determination technique for crystalline and non-crystalline protein assemblies, e.g., amyloids. Resonance assignment constitutes the first and often very time-consuming step to a structure. We present ssFLYA, a generally applicable algorithm for automatic assignment of protein solid-state NMR spectra. Application to microcrystals of ubiquitin and the Ure2 prion C-terminal domain, as well as amyloids of HET-s(218-289) and {alpha}-synuclein yielded 88-97 % correctness for the backbone and side-chain assignments that are classified as self-consistent by the algorithm, and 77-90 % correctness if also assignments classified as tentative by the algorithm are included.

Technical refolding of proteins: Do we have freedom to operate?

Science.gov (United States)

Eiberle, Maria K; Jungbauer, Alois

2010-06-01

Expression as inclusion bodies in Escherichia coli is a widely used method for the large-scale production of therapeutic proteins that do not require post-translational modifications. High expression yields and simple recovery steps of inclusion bodies from the host cells are attractive features industrially. However, the value of an inclusion body-based process is dominated by the solubilization and refolding technologies. Scale-invariant technologies that are economical and applicable for a wide range of proteins are requested by industry. The main challenge is to convert the denatured protein into its native conformation at high yields. Refolding competes with misfolding and aggregation. Thus, the yield of native monomer depends strongly on the initial protein concentrations in the refolding solution. Reasonable yields are attained at low concentrations (freedom to operate.

Quantitative aspects of crystalline lactose in milk products

NARCIS (Netherlands)

Roetman, K.

1982-01-01

The occurrence of crystalline lactose in milk products and its influence on their physical properties are briefly reviewed. The importance of the quantitive determination of crystalline lactose for scientific and industrial purposes is indicated, and a summary is given of our earlier work. This

Attenuation of Thermal Neutrons by Crystalline Silicon

International Nuclear Information System (INIS)

Adib, M.; Habib, N.; Ashry, A.; Fathalla, M.

2002-01-01

A simple formula is given which allows to calculate the contribution of the total neutron cross - section including the Bragg scattering from different (hkt) planes to the neutron * transmission through a solid crystalline silicon. The formula takes into account the silicon form of poly or mono crystals and its parameters. A computer program DSIC was developed to provide the required calculations. The calculated values of the total neutron cross-section of perfect silicon crystal at room and liquid nitrogen temperatures were compared with the experimental ones. The obtained agreement shows that the simple formula fits the experimental data with sufficient accuracy .A good agreement was also obtained between the calculated and measured values of polycrystalline silicon in the energy range from 5 eV to 500μ eV. The feasibility study on using a poly-crystalline silicon as a cold neutron filter and mono-crystalline as a thermal neutron one is given. The optimum crystal thickness, mosaic spread, temperature and cutting plane for efficiently transmitting the thermal reactor neutrons, while rejecting both fast neutrons and gamma rays accompanying the thermal ones for the mono crystalline silicon are also given

Opacity alterations of bovine crystalline proteins irradiated with 10 Co in vitro in the presence of sulfonate compounds

International Nuclear Information System (INIS)

Bernardes, D.M.L.; Mastro, N.L. del

1990-01-01

Sulfhydrilic compounds with a strong basic function separated from the SH group by no more than three C atoms, as amino ethyl iso thiourea (AET) and mercapto ethyl alanine (MEA) are exceptionally effective in competing with free radicals produced by water radiolysis. In a similar way, dimethyl sulfoxide (DMSO) is also effective in the removal of hydroxyl radicals. In the present work, aqueous solutions of crystalline removed surgically from bovine eyes were used. Crystalline were homogenized, the suspension centrifuged and the supernatant dialysed. From the dialysed supernatant a series of solutions was prepared that was 60 Co irradiated with different doses from 5,000 to 25,000 Gy in the presence of 10 mM AET, MEA and DMSO. The degree of opacification was read spectrophotometricaly at 600 nm. The results pointed out a decrease of the increment of opacity produced by the radiation in the presence of those free radical scavengers, showing a radioprotective action of them at the molecular level, that can be measured by this method that mimics the cataract formation in eye lens. (author)

Delimiting Inclusive Design

DEFF Research Database (Denmark)

Herriott, Richard

2014-01-01

This paper was written as an answer to the question raised by my PhD dissertation on accessibility through user-centred and Inclusive Design (ID) methods: can Inclusive Design be delimited? The literature on Inclusive Design deals almost entrirely with consumer product design and assistive...

Analysis of the post-translational modifications of the individual amino acids in lens proteins which were induced by aging and irradiation

International Nuclear Information System (INIS)

Fujii, Noriko; Kim, Ingu; Saito, Takeshi; Takata, Takumi

2017-01-01

The eye lens is a transparent organ that functions to focus light and images on the retina. The transparency and high refraction of the lens are maintained by the function of α-, β- and γ-crystallins. These long-lived proteins are subject to various post-translational modifications, such as oxidation, deamidation, truncation and isomerization, which occur gradually during the aging process. Such modifications, which are generated by UV light and oxidative stress, decrease crystallin solubility and lens transparency, and ultimately lead to the development of age-related cataracts. Here, we irradiated young rat lenses with γ-rays (5-500 Gy) and extracted the water-soluble (WS) and insoluble (WI) protein fractions. The WS and WI lens proteins were digested with trypsin, and the resulting peptides were analyzed by one-shot LC-MS/MS to determine the specific sites of oxidation of methionine and tryptophan, deamidation of asparagine and glutamine, and isomerization of aspartyl in rat α- and β-crystallins in the WS and WI fractions. Oxidation and deamidation occurred in several crystallins after irradiation at more than, respectively, 50 Gy and 5 Gy; however, isomerization did not occur in any crystallin even after exposure to 500 Gy of irradiation. The number of oxidation and deamidation sites was much higher in the WI than in the WS fraction. Furthermore, the oxidation and deamidation sites in rat crystallins resemble those reported in crystallins from human age-related cataracts. Thus, this study on post-translational modifications of crystallins induced by ionizing irradiation may provide useful information relevant to the formation of human age-related cataracts. (author)

Nonlinear optical properties of TeO$_2$ crystalline phases from first principles

OpenAIRE

Berkaine, Nabil; Orhan, Emmanuelle; Masson, Olivier; Thomas, Philippe; Junquera, Javier

2010-01-01

We have computed second and third nonlinear optical susceptibilities of two crystalline bulk tellurium oxide polymorphs: $\\alpha$-TeO$_{2}$ (the most stable crystalline bulk phase) and $\\gamma$-TeO$_{2}$ (the crystalline phase that ressembles the more to the glass phase. Third order nonlinear susceptibilities of the crystalline phases are two orders of magnitude larger than $\\alpha$-SiO$_{2}$ cristoballite, thus extending the experimental observations on glasses to the case of crystalline com...

Tar DNA binding protein-43 (TDP-43 associates with stress granules: analysis of cultured cells and pathological brain tissue.

Directory of Open Access Journals (Sweden)

Liqun Liu-Yesucevitz

2010-10-01

Full Text Available Tar DNA Binding Protein-43 (TDP-43 is a principle component of inclusions in many cases of frontotemporal lobar degeneration (FTLD-U and amyotrophic lateral sclerosis (ALS. TDP-43 resides predominantly in the nucleus, but in affected areas of ALS and FTLD-U central nervous system, TDP-43 is aberrantly processed and forms cytoplasmic inclusions. The mechanisms governing TDP-43 inclusion formation are poorly understood. Increasing evidence indicates that TDP-43 regulates mRNA metabolism by interacting with mRNA binding proteins that are known to associate with RNA granules. Here we show that TDP-43 can be induced to form inclusions in cell culture and that most TDP-43 inclusions co-localize with SGs. SGs are cytoplasmic RNA granules that consist of mixed protein-RNA complexes. Under stressful conditions SGs are generated by the reversible aggregation of prion-like proteins, such as TIA-1, to regulate mRNA metabolism and protein translation. We also show that disease-linked mutations in TDP-43 increased TDP-43 inclusion formation in response to stressful stimuli. Biochemical studies demonstrated that the increased TDP-43 inclusion formation is associated with accumulation of TDP-43 detergent insoluble complexes. TDP-43 associates with SG by interacting with SG proteins, such as TIA-1, via direct protein-protein interactions, as well as RNA-dependent interactions. The signaling pathway that regulates SGs formation also modulates TDP-43 inclusion formation. We observed that inclusion formation mediated by WT or mutant TDP-43 can be suppressed by treatment with translational inhibitors that suppress or reverse SG formation. Finally, using Sudan black to quench endogenous autofluorescence, we also demonstrate that TDP-43 positive-inclusions in pathological CNS tissue co-localize with multiple protein markers of stress granules, including TIA-1 and eIF3. These data provide support for accumulating evidence that TDP-43 participates in the SG pathway.

Social imaginaries and inclusion

DEFF Research Database (Denmark)

Hansen, Janne Hedegaard

2016-01-01

that inclusion on the one hand may be seen to be about human rights, solidarity, and democracy, and on the other hand, it is about ensuring the cohesion of neoliberal society by means of every person’s obligation to realize one’s potential through learning, development, and education regardless of one’s needs......, the development of inclusive schools and inclusive learning environments will involve both inclusion and exclusion processes. With this starting point, international educational research knowledge about inclusive schools and inclusive learning environments in general will be related to the fundamental dilemma...

Novel polypyrrole films with excellent crystallinity and good thermal stability

International Nuclear Information System (INIS)

Jeeju, Pullarkat P.; Varma, Sreekanth J.; Francis Xavier, Puthampadath A.; Sajimol, Augustine M.; Jayalekshmi, Sankaran

2012-01-01

Polypyrrole has drawn a lot of interest due to its high thermal and environmental stability in addition to high electrical conductivity. The present work highlights the enhanced crystallinity of polypyrrole films prepared from the redoped sample solution. Initially hydrochloric acid doped polypyrrole was prepared by chemical oxidative polymerization of pyrrole using ammonium peroxidisulphate as oxidant. The doped polypyrrole was dedoped using ammonia solution and then redoped with camphor sulphonic acid. Films were coated on ultrasonically cleaned glass substrates from the redoped sample solution in meta-cresol. The enhanced crystallinity of the polypyrrole films has been established from X-ray diffraction (XRD) studies. The room temperature electrical conductivity of the redoped polypyrrole film is about 30 times higher than that of the hydrochloric acid doped pellet sample. The results of Raman spectroscopy, Differential scanning calorimetry (DSC) and Thermogravimetric analysis (TGA) of the samples support the enhancement in crystallinity. Percentage crystallinity of the samples is estimated from XRD and DSC data. The present work is significant, since crystallinity of films is an important parameter for selecting polymers for specific applications. - Highlights: ► Polypyrrole films redoped with CSA have been prepared from meta-cresol solution. ► The solution casted films exhibit semi-crystallinity and good thermal stability. ► Percentage crystallinity estimated using XRD and DSC analysis is about 65%. ► Raman studies support the enhancement in crystallinity based on XRD and DSC data. ► The conductivity of the film is 30 times higher than that of HCl doped sample.

Novel polypyrrole films with excellent crystallinity and good thermal stability

Energy Technology Data Exchange (ETDEWEB)

Jeeju, Pullarkat P., E-mail: jeejupp@gmail.com [Division for Research in Advanced Materials, Department of Physics, Cochin University of Science and Technology, Cochin-22, Kerala (India); Varma, Sreekanth J.; Francis Xavier, Puthampadath A.; Sajimol, Augustine M. [Division for Research in Advanced Materials, Department of Physics, Cochin University of Science and Technology, Cochin-22, Kerala (India); Jayalekshmi, Sankaran, E-mail: jayalekshmi@cusat.ac.in [Division for Research in Advanced Materials, Department of Physics, Cochin University of Science and Technology, Cochin-22, Kerala (India)

2012-06-15

Polypyrrole has drawn a lot of interest due to its high thermal and environmental stability in addition to high electrical conductivity. The present work highlights the enhanced crystallinity of polypyrrole films prepared from the redoped sample solution. Initially hydrochloric acid doped polypyrrole was prepared by chemical oxidative polymerization of pyrrole using ammonium peroxidisulphate as oxidant. The doped polypyrrole was dedoped using ammonia solution and then redoped with camphor sulphonic acid. Films were coated on ultrasonically cleaned glass substrates from the redoped sample solution in meta-cresol. The enhanced crystallinity of the polypyrrole films has been established from X-ray diffraction (XRD) studies. The room temperature electrical conductivity of the redoped polypyrrole film is about 30 times higher than that of the hydrochloric acid doped pellet sample. The results of Raman spectroscopy, Differential scanning calorimetry (DSC) and Thermogravimetric analysis (TGA) of the samples support the enhancement in crystallinity. Percentage crystallinity of the samples is estimated from XRD and DSC data. The present work is significant, since crystallinity of films is an important parameter for selecting polymers for specific applications. - Highlights: Black-Right-Pointing-Pointer Polypyrrole films redoped with CSA have been prepared from meta-cresol solution. Black-Right-Pointing-Pointer The solution casted films exhibit semi-crystallinity and good thermal stability. Black-Right-Pointing-Pointer Percentage crystallinity estimated using XRD and DSC analysis is about 65%. Black-Right-Pointing-Pointer Raman studies support the enhancement in crystallinity based on XRD and DSC data. Black-Right-Pointing-Pointer The conductivity of the film is 30 times higher than that of HCl doped sample.

On the formation and functions of high and very high magnesium calcites in the continuously growing teeth of the echinoderm Lytechinus variegatus: development of crystallinity and protein involvement.

Science.gov (United States)

Veis, Arthur; Stock, Stuart R; Alvares, Keith; Lux, Elizabeth

2011-01-01

Sea urchin teeth grow continuously and develop a complex mineralized structure consisting of spatially separate but crystallographically aligned first stage calcitic elements of high Mg content (5-15 mol% mineral). These become cemented together by epitaxially oriented second stage very high Mg calcite (30-40 mol% mineral). In the tooth plumula, ingressing preodontoblasts create layered cellular syncytia. Mineral deposits develop within membrane-bound compartments between cellular syncytial layers. We seek to understand how this complex tooth architecture is developed, how individual crystalline calcitic elements become crystallographically aligned, and how their Mg composition is regulated. Synchrotron microbeam X-ray scattering was performed on live, freshly dissected teeth. We observed that the initial diffracting crystals lie within independent syncytial spaces in the plumula. These diffraction patterns match those of mature tooth calcite. Thus, the spatially separate crystallites grow with the same crystallographic orientation seen in the mature tooth. Mineral-related proteins from regions with differing Mg contents were isolated, sequenced, and characterized. A tooth cDNA library was constructed, and selected matrix-related proteins were cloned. Antibodies were prepared and used for immunolocaliztion. Matrix-related proteins are acidic, phosphorylated, and associated with the syncytial membranes. Time-of-flight secondary ion mass spectroscopy of various crystal elements shows unique amino acid, Mg, and Ca ion distributions. High and very high Mg calcites differ in Asp content. Matrix-related proteins are phosphorylated. Very high Mg calcite is associated with Asp-rich protein, and it is restricted to the second stage mineral. Thus, the composition at each part of the tooth is related to architecture and function. Copyright © 2011 S. Karger AG, Basel.

Enhanced expression and purification of camelid single domain VHH antibodies from classical inclusion bodies.

Science.gov (United States)

Maggi, Maristella; Scotti, Claudia

2017-08-01

Single domain antibodies (sdAbs) are small antigen-binding domains derived from naturally occurring, heavy chain-only immunoglobulins isolated from camelid and sharks. They maintain the same binding capability of full-length IgGs but with improved thermal stability and permeability, which justifies their scientific, medical and industrial interest. Several described recombinant forms of sdAbs have been produced in different hosts and with different strategies. Here we present an optimized method for a time-saving, high yield production and extraction of a poly-histidine-tagged sdAb from Escherichia coli classical inclusion bodies. Protein expression and extraction were attempted using 4 different methods (e.g. autoinducing or IPTG-induced soluble expression, non-classical and classical inclusion bodies). The best method resulted to be expression in classical inclusion bodies and urea-mediated protein extraction which yielded 60-70 mg/l bacterial culture. The method we here describe can be of general interest for an enhanced and efficient heterologous expression of sdAbs for research and industrial purposes. Copyright © 2017 Elsevier Inc. All rights reserved.

Fourier transform infrared spectroscopic estimation of crystallinity in ...

Indian Academy of Sciences (India)

Wintec

The crystallinity parameter is calculated by using a standard procedure which can be used to estimate the distribution of quartz in various rocks for mining purpose. The infrared ... The X-ray diffraction full ... crystallinity and trace mineral components of rocks (Partha- ... infrared techniques (Rice et al 1995). ... The absorption.

Used fuel disposition in crystalline rocks

Energy Technology Data Exchange (ETDEWEB)

Wang, Y. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Hadgu, Teklu [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Kalinina, Elena Arkadievna [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Jerden, James L. [Argonne National Lab. (ANL), Argonne, IL (United States); Copple, Jacqueline M. [Argonne National Lab. (ANL), Argonne, IL (United States); Cruse, T. [Argonne National Lab. (ANL), Argonne, IL (United States); Ebert, W. [Argonne National Lab. (ANL), Argonne, IL (United States); Buck, E. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Eittman, R. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Tinnacher, R. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Tournassat, Christophe. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Davis, J. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Viswanathan, H. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Chu, S. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Dittrich, T. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Hyman, F. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Karra, S. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Makedonska, N. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Reimus, P. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Zavarin, Mavrik [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Joseph, C. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

2016-09-01

The U.S. Department of Energy Office of Nuclear Energy, Office of Fuel Cycle Technology established the Used Fuel Disposition Campaign (UFDC) in fiscal year 2010 (FY10) to conduct the research and development (R&D) activities related to storage, transportation and disposal of used nuclear fuel and high level nuclear waste. The objective of the Crystalline Disposal R&D Work Package is to advance our understanding of long-term disposal of used fuel in crystalline rocks and to develop necessary experimental and computational capabilities to evaluate various disposal concepts in such media.

Irradiation induced crystalline to amorphous transition

International Nuclear Information System (INIS)

Bourgoin, J.

1980-01-01

Irradiation of a crystalline solid with energetic heavy particles results in cascades of defects which, with increasing dose, overlap and form a continuous disordered layer. In semiconductors the physical properties of such disordered layers are found to be similar to those of amorphous layers produced by evaporation. It is shown in the case of silicon, that the transition from a disordered crystalline (X) layer to an amorphous (α) layer occurs when the Gibbs energy of the X phase and of the defects it contains becomes larger than the Gibbs energy of the α phase. (author)

Phenotypic variability within the inclusion body spectrum of basophilic inclusion body disease and neuronal intermediate filament inclusion disease in frontotemporal lobar degenerations with FUS-positive inclusions.

Science.gov (United States)

Gelpi, Ellen; Lladó, Albert; Clarimón, Jordi; Rey, Maria Jesús; Rivera, Rosa Maria; Ezquerra, Mario; Antonell, Anna; Navarro-Otano, Judith; Ribalta, Teresa; Piñol-Ripoll, Gerard; Pérez, Anna; Valldeoriola, Francesc; Ferrer, Isidre

2012-09-01

Basophilic inclusion body disease and neuronal intermediate filament inclusion disease (NIFID) are rare diseases included among frontotemporal lobar degenerations with FUS-positive inclusions (FTLD-FUS). We report clinical and pathologic features of 2 new patients and reevaluate neuropathologic characteristics of 2 previously described cases, including an early-onset case of basophilic inclusion body disease (aged 38 years) with a 5-year disease course and abundant FUS-positive inclusion bodies and 3 NIFID cases. One NIFID case (aged 37 years) presented with early-onset psychiatric disturbances and rapidly progressive cognitive decline. Two NIFID cases had later onset (aged 64 years and 70 years) and complex neurologic deficits. Postmortem neuropathologic studies in late-onset NIFID cases disclosed α-internexin-positive "hyaline conglomerate"-type inclusions that were positive with 1 commercial anti-FUS antibody directed to residues 200 and 250, but these were negative to amino acids 90 and 220 of human FUS. Early-onset NIFID had similar inclusions that were positive with both commercial anti-FUS antibodies. Genetic testing performed on all cases revealed no FUS gene mutations. These findings indicate that phenotypic variability in NIFID, including clinical manifestations and particular neuropathologic findings, may be related to the age at onset and individual differences in the evolution of lesions.

Model for screened, charge-regulated electrostatics of an eye lens protein: Bovine gammaB-crystallin

Science.gov (United States)

Wahle, Christopher W.; Martini, K. Michael; Hollenbeck, Dawn M.; Langner, Andreas; Ross, David S.; Hamilton, John F.; Thurston, George M.

2017-09-01

We model screened, site-specific charge regulation of the eye lens protein bovine gammaB-crystallin (γ B ) and study the probability distributions of its proton occupancy patterns. Using a simplified dielectric model, we solve the linearized Poisson-Boltzmann equation to calculate a 54 ×54 work-of-charging matrix, each entry being the modeled voltage at a given titratable site, due to an elementary charge at another site. The matrix quantifies interactions within patches of sites, including γ B charge pairs. We model intrinsic p K values that would occur hypothetically in the absence of other charges, with use of experimental data on the dependence of p K values on aqueous solution conditions, the dielectric model, and literature values. We use Monte Carlo simulations to calculate a model grand-canonical partition function that incorporates both the work-of-charging and the intrinsic p K values for isolated γ B molecules and we calculate the probabilities of leading proton occupancy configurations, for 4 Debye screening lengths from 6 to 20 Å. We select the interior dielectric value to model γ B titration data. At p H 7.1 and Debye length 6.0 Å, on a given γ B molecule the predicted top occupancy pattern is present nearly 20% of the time, and 90% of the time one or another of the first 100 patterns will be present. Many of these occupancy patterns differ in net charge sign as well as in surface voltage profile. We illustrate how charge pattern probabilities deviate from the multinomial distribution that would result from use of effective p K values alone and estimate the extents to which γ B charge pattern distributions broaden at lower p H and narrow as ionic strength is lowered. These results suggest that for accurate modeling of orientation-dependent γ B -γ B interactions, consideration of numerous pairs of proton occupancy patterns will be needed.

Thermodynamics of Crystalline States

CERN Document Server

Fujimoto, Minoru

2013-01-01

Thermodynamics is a well-established discipline of physics for properties of matter in thermal equilibrium with the surroundings. Applying to crystals, however, the laws encounter undefined properties of crystal lattice, which therefore need to be determined for a clear and well-defined description of crystalline states. Thermodynamics of Crystalline States explores the roles played by order variables and dynamic lattices in crystals in a wholly new way. The book begins by clarifying basic concepts for stable crystals. Next, binary phase transitions are discussed to study collective motion of order variables, as described mostly as classical phenomena. New to this edition is the examination of magnetic crystals, where magnetic symmetry is essential for magnetic phase transitions. The multi-electron system is also discussed  theoretically, as a quantum-mechanical example, for superconductivity in metallic crystals. Throughout the book, the role played by the lattice is emphasized and studied in-depth. Thermod...

Cellulose binding domain proteins

Science.gov (United States)

Shoseyov, Oded; Shpiegl, Itai; Goldstein, Marc; Doi, Roy

1998-01-01

A cellulose binding domain (CBD) having a high affinity for crystalline cellulose and chitin is disclosed, along with methods for the molecular cloning and recombinant production thereof. Fusion products comprising the CBD and a second protein are likewise described. A wide range of applications are contemplated for both the CBD and the fusion products, including drug delivery, affinity separations, and diagnostic techniques.

Enhanced bioavailability of nerve growth factor with phytantriol lipid-based crystalline nanoparticles in cochlea

Directory of Open Access Journals (Sweden)

Bu M

2015-11-01

Full Text Available Meng Bu,1,2 Jingling Tang,3 Yinghui Wei,4 Yanhui Sun,1 Xinyu Wang,1 Linhua Wu,2 Hongzhuo Liu1 1School of Pharmacy, Shenyang Pharmaceutical University, Shenyang, People’s Republic of China; 2Department of Pharmacy, the Second Affiliated Hospital, 3School of Pharmacy, Harbin Medical University, Harbin, People’s Republic of China; 4College of Pharmaceutical Science, Zhejiang Chinese Medical University, Hangzhou, People’s Republic of China Purpose: Supplementation of exogenous nerve growth factor (NGF into the cochlea of deafened animals rescues spiral ganglion cells from degeneration. However, a safe and potent delivery of therapeutic proteins, such as NGF, to spiral ganglion cells remains one of the greatest challenges. This study presents the development of self-assembled cubic lipid-based crystalline nanoparticles to enhance inner ear bioavailability of bioactive NGF via a round window membrane route.Methods: A novel nanocarrier-entrapped NGF was developed based on phytantriol by a liquid precursor dilution, with Pluronic® F127 and propylene glycol as the surfactant and solubilizer, respectively. Upon dilution of the liquid lipid precursors, monodispersed submicron-sized particles with a slight negative charge formed spontaneously.Results: Biological activity of entrapped NGF was assessed using pheochromocytoma cells with NGF-loaded reservoirs to induce significant neuronal outgrowth, similar to that seen in free NGF-treated controls. Finally, a 3.28-fold increase in inner ear bioavailability was observed after administration of phytantriol lipid-based crystalline nanoparticles as compared to free drug, contributing to an enhanced drug permeability of the round window membrane. Conclusion: Data presented here demonstrate the potential of lipid-based crystalline nanoparticles to improve the outcomes of patients bearing cochlear implants. Keywords: nerve growth factor, lipid-based crystalline nanoparticles, PC12 cells, inner ear drug

Morphology and crystallinity of sisal nanocellulose after sonication

Science.gov (United States)

Sosiati, H.; Wijayanti, D. A.; Triyana, K.; Kamiel, B.

2017-09-01

Different preparation methods on the natural fibers resulted in different morphology. However, the relationships between type of natural fibers, preparation methods and the morphology of produced nanocellulose could not be exactly defined. The sisal nanocellulose was presently prepared by alkalization and bleaching followed by sonication to verify changes in the morphology and crystallinity of nanocellulose related to the formation mechanism. The extracted microcellulose was subjected to scanning electron microscopy (SEM) and x-ray diffraction (XRD) analysis. The isolated cellulose nanospheres were examined with respect to morphology by SEM and transmission electron microscopy (TEM) and, to crystallinity by electron diffraction analysis. Bleaching after alkalization made the microfibrils clearly separated from each other to the individual fiber whose width of the single fiber was ranging from 6 to 13 µm. The XRD crystallinity index (CI) of microcellulose gradually increased after the chemical treatments; 83.12% for raw sisal fiber, 88.57% for alkali treated fiber and 94.03% for bleached fibers. The ultrasonic agitation after bleaching that was carried out at 750 Watt, 20 kHz and amplitude of 39% for 2 h produces homogeneous cellulose nanospheres less than 50 nm in diameter with relatively low crystallinity. The electron diffraction analysis confirmed that the low crystallinity of produced nnocellulose is related to the effect of chemical treatment done before sonication.

Lack of Evidence of In-Situ Decay of Aluminum-26 in a FeO-Poor Ferromagnesian Crystalline Silicate Particle, Pyxie, from Comet Wild 2

Science.gov (United States)

Nakashima, D.; Ushikubo, T.; Weisberg, M. K.; Zolensky, M. E.; Ebel, D. S.; Kita, N. T.

2014-01-01

One of the important discoveries from the Stardust mission is the observation of crystalline silicate particles that resemble Ca, Al-rich inclusions (CAIs) and chondrules in carbonaceous chondrites], which suggests radial transport of high temperature solids from the inner to the outer solar nebula regions and capture by accreting cometary objects. The Al-Mg isotope analyses of CAI-like and type II chondrule-like particles revealed no excess of Mg-26 derived from in-situ decay of Al-26 (Tau)(sub 1/2) = 0.705Myr; ), suggesting late formation of these particles. However, the number of Wild 2 particles analyzed for Al-Mg isotopes is still limited (n = 3). In order to better understand the timing of the formation of Wild 2 particles and possible radial transport in the protoplanetary disk, we performed SIMS (Secondary Ion Mass Spectrometer) Al-Mg isotope analyses of plagioclase in a FeO-poor ferromagnesian Wild 2 particle, which is the most abundant type among crystalline Wild 2 particles.

Positive in vitro wound healing effects of functional inclusion bodies of a lipoxygenase from the Mexican axolotl.

Science.gov (United States)

Stamm, Anne; Strauß, Sarah; Vogt, Peter; Scheper, Thomas; Pepelanova, Iliyana

2018-04-07

AmbLOXe is a lipoxygenase, which is up-regulated during limb-redevelopment in the Mexican axolotl, Ambystoma mexicanum, an animal with remarkable regeneration capacity. Previous studies have shown that mammalian cells transformed with the gene of this epidermal lipoxygenase display faster migration and wound closure rate during in vitro wound healing experiments. In this study, the gene of AmbLOXe was codon-optimized for expression in Escherichia coli and was produced in the insoluble fraction as protein aggregates. These inclusion bodies or nanopills were shown to be reservoirs containing functional protein during in vitro wound healing assays. For this purpose, functional inclusion bodies were used to coat cell culture surfaces prior cell seeding or were added directly to the medium after cells reached confluence. In both scenarios, AmbLOXe inclusion bodies led to faster migration rate and wound closure, in comparison to controls containing either no AmbLOXe or GFP inclusion bodies. Our results demonstrate that AmbLOXe inclusion bodies are functional and may serve as stable reservoirs of this enzyme. Nevertheless, further studies with soluble enzyme are also necessary in order to start elucidating the exact molecular substrates of AmbLOXe and the biochemical pathways involved in the wound healing effect.

Mechanism of RPE cell death in α-crystallin deficient mice: a novel and critical role for MRP1-mediated GSH efflux.

Directory of Open Access Journals (Sweden)

Parameswaran G Sreekumar

Full Text Available Absence of α-crystallins (αA and αB in retinal pigment epithelial (RPE cells renders them susceptible to oxidant-induced cell death. We tested the hypothesis that the protective effect of α-crystallin is mediated by changes in cellular glutathione (GSH and elucidated the mechanism of GSH efflux. In α-crystallin overexpressing cells resistant to cell death, cellular GSH was >2 fold higher than vector control cells and this increase was seen particularly in mitochondria. The high GSH levels associated with α-crystallin overexpression were due to increased GSH biosynthesis. On the other hand, cellular GSH was decreased by 50% in murine retina lacking αA or αB crystallin. Multiple multidrug resistance protein (MRP family isoforms were expressed in RPE, among which MRP1 was the most abundant. MRP1 was localized to the plasma membrane and inhibition of MRP1 markedly decreased GSH efflux. MRP1-suppressed cells were resistant to cell death and contained elevated intracellular GSH and GSSG. Increased GSH in MRP1-supressed cells resulted from a higher conversion of GSSG to GSH by glutathione reductase. In contrast, GSH efflux was significantly higher in MRP1 overexpressing RPE cells which also contained lower levels of cellular GSH and GSSG. Oxidative stress further increased GSH efflux with a decrease in cellular GSH and rendered cells apoptosis-prone. In conclusion, our data reveal for the first time that 1 MRP1 mediates GSH and GSSG efflux in RPE cells; 2 MRP1 inhibition renders RPE cells resistant to oxidative stress-induced cell death while MRP1 overexpression makes them susceptible and 3 the antiapoptotic function of α-crystallin in oxidatively stressed cells is mediated in part by GSH and MRP1. Our findings suggest that MRP1 and α crystallin are potential therapeutic targets in pathological retinal degenerative disorders linked to oxidative stress.

Ore Characteristics and Fluid Inclusion of the Base Metal Vein Deposit in Moncong Bincanai Area, Gowa, South Sulawesi, Indonesia

Directory of Open Access Journals (Sweden)

Asmariyadi Asmariyadi

2014-07-01

Full Text Available DOI: 10.17014/ijog.v7i4.146This paper is dealing with ore characteristics and fluid inclusion of the Moncong Bincanai, Biringbulu Subregency of Gowa Regency, South Sulawesi Province, Indonesia. The mineralization is a vein type, with the orientation of N170oE /65oSW, hosted in open-space filling within basalt. The mineralization consists of galena, sphalerite, chalcopyrite, and pyrite. Vein thickness ranges from 5 - 17 cm, showing a crustiform banding texture, with a sequence from outer to centre: quartz, carbonate (siderite, sulphide. The quartz displays primary growth textures such as comb, crystalline, saccharoidal, and colloform. Analytical methods applied include AAS and fluid inclusion microthermometry. Chemical composition of the vein indicates an average of Pb = 47.92%, Cu = 1.27%, Zn = 1.02%, and Fe = 9.46%, which shows a significant concentration of Pb. Fluid inclusion microthermometry results indicate a range of formation temperature of 240 - 250C and salinity of the responsible hydrothermal fluid of 2.1 - 2.5 wt.% NaCl eq. The deposit is categorized into low-sulfidation epithermal deposits, which was formed within a range of 410 - 440 m below paleosurface.

The strength of crystalline color superconductors

International Nuclear Information System (INIS)

Mannarelli, Massimo; Rajagopal, Krishna; Sharma, Rishi

2007-01-01

We present a study of the shear modulus of the crystalline color superconducting phase of quark matter, showing that this phase of dense, but not asymptotically dense, quark matter responds to shear stress as a very rigid solid. This phase is characterized by a gap parameter Δ that is periodically modulated in space and therefore spontaneously breaks translational invariance. We derive the effective action for the phonon fields that describe space- and time-dependent fluctuations of the crystal structure formed by Δ, and obtain the shear modulus from the coefficients of the spatial derivative terms. Within a Ginzburg-Landau approximation, we find shear moduli which are 20 to 1000 times larger than those of neutron star crusts. This phase of matter is thus more rigid than any known material in the universe, but at the same time the crystalline color superconducting phase is also superfluid. These properties raise the possibility that the presence of this phase within neutron stars may have distinct implications for their phenomenology. For example (some) pulsar glitches may originate in crystalline superconducting neutron star cores

76 FR 78313 - Crystalline Silicon Photovoltaic Cells and Modules From China

Science.gov (United States)

2011-12-16

...)] Crystalline Silicon Photovoltaic Cells and Modules From China Determinations On the basis of the record \\1... injured by reason of imports from China of crystalline silicon photovoltaic cells and modules, provided... imports of crystalline silicon photovoltaic cells and modules from China. Accordingly, effective October...

Automated solid-state NMR resonance assignment of protein microcrystals and amyloids

International Nuclear Information System (INIS)

Schmidt, Elena; Gath, Julia; Habenstein, Birgit; Ravotti, Francesco; Székely, Kathrin; Huber, Matthias; Buchner, Lena; Böckmann, Anja; Meier, Beat H.; Güntert, Peter

2013-01-01

Solid-state NMR is an emerging structure determination technique for crystalline and non-crystalline protein assemblies, e.g., amyloids. Resonance assignment constitutes the first and often very time-consuming step to a structure. We present ssFLYA, a generally applicable algorithm for automatic assignment of protein solid-state NMR spectra. Application to microcrystals of ubiquitin and the Ure2 prion C-terminal domain, as well as amyloids of HET-s(218–289) and α-synuclein yielded 88–97 % correctness for the backbone and side-chain assignments that are classified as self-consistent by the algorithm, and 77–90 % correctness if also assignments classified as tentative by the algorithm are included

L-arginine mediated renaturation enhances yield of human, α6 Type IV collagen non-collagenous domain from bacterial inclusion bodies.

Science.gov (United States)

Gunda, Venugopal; Boosani, Chandra Shekhar; Verma, Raj Kumar; Guda, Chittibabu; Sudhakar, Yakkanti Akul

2012-10-01

The anti-angiogenic, carboxy terminal non-collagenous domain (NC1) derived from human Collagen type IV alpha 6 chain, [α6(IV)NC1] or hexastatin, was earlier obtained using different recombinant methods of expression in bacterial systems. However, the effect of L-arginine mediated renaturation in enhancing the relative yields of this protein from bacterial inclusion bodies has not been evaluated. In the present study, direct stirring and on-column renaturation methods using L-arginine and different size exclusion chromatography matrices were applied for enhancing the solubility in purifying the recombinant α6(IV)NC1 from bacterial inclusion bodies. This methodology enabled purification of higher quantities of soluble protein from inclusion bodies, which inhibited endothelial cell proliferation, migration and tube formation. Thus, the scope for L-arginine mediated renaturation in obtaining higher yields of soluble, biologically active NC1 domain from bacterial inclusion bodies was evaluated.

From Cellulosic Based Liquid Crystalline Sheared Solutions to 1D and 2D Soft Materials

Directory of Open Access Journals (Sweden)

Maria Helena Godinho

2014-06-01

Full Text Available Liquid crystalline cellulosic-based solutions described by distinctive properties are at the origin of different kinds of multifunctional materials with unique characteristics. These solutions can form chiral nematic phases at rest, with tuneable photonic behavior, and exhibit a complex behavior associated with the onset of a network of director field defects under shear. Techniques, such as Nuclear Magnetic Resonance (NMR, Rheology coupled with NMR (Rheo-NMR, rheology, optical methods, Magnetic Resonance Imaging (MRI, Wide Angle X-rays Scattering (WAXS, were extensively used to enlighten the liquid crystalline characteristics of these cellulosic solutions. Cellulosic films produced by shear casting and fibers by electrospinning, from these liquid crystalline solutions, have regained wider attention due to recognition of their innovative properties associated to their biocompatibility. Electrospun membranes composed by helical and spiral shape fibers allow the achievement of large surface areas, leading to the improvement of the performance of this kind of systems. The moisture response, light modulated, wettability and the capability of orienting protein and cellulose crystals, opened a wide range of new applications to the shear casted films. Characterization by NMR, X-rays, tensile tests, AFM, and optical methods allowed detailed characterization of those soft cellulosic materials. In this work, special attention will be given to recent developments, including, among others, a moisture driven cellulosic motor and electro-optical devices.

Liquid Crystalline Perylene diimides : Architecture and Charge Carrier Mobilities

NARCIS (Netherlands)

Struijk, C.W.; Sieval, A.B.; Dakhorst, J.E.J.; Dijk, van M.; Kimkes, P.; Koehorst, R.B.M.; Donker, H.

2000-01-01

The phase behavior of three N-alkyl-substituted perylene diimide derivatives is examined by differential scanning calorimetry and polarized optical microscopy. The occurrence of multiple phase transitions indicates several crystalline and several liquid crystalline phases. X-ray diffraction

Diffraction enhanced X-ray imaging of mammals crystalline lens

International Nuclear Information System (INIS)

Antunes, A.; Hoennicke, M.G.; Safatle, A.M.V.; Cusatis, C.; Moraes Barros, P.S.; Morelhao, S.L.

2005-01-01

Crystalline lenses are transparent biological materials where the organization of the lens fibers can also be affected by changes at molecular level, and therefore the structure and morphology of the tissue can be correlated to the loss of transparency of the lens. In this work, internal structure of mammal lenses regarding the long-range ordering of the fibers are investigated by diffraction enhanced X-ray imaging (DEI) radiography. Moreover, DEI and absorption X-ray synchrotron radiographs for healthy and cataractous crystalline lenses are compared. Significant differences in healthy and cataractous crystalline lenses are observed

Electronic processes in non-crystalline materials

CERN Document Server

Mott, Nevill Francis

2012-01-01

Since the first edition of this highly successful book the field saw many great developments both in experimental and theoretical studies of electrical properties of non-crystalline solids. It became necessary to rewrite nearly the whole book, while the aims of the second edition remained the same: to set out the theoretical concepts, to test them by comparison with experiment for a wide variety of phenomena, and to apply them to non-crystalline materials. Sir Nevill Mott shared the1977 Nobel Prize for Physics, awarded for his research work in this field. The reissue of this book as part of th

Solvated protein-DNA docking using HADDOCK

Energy Technology Data Exchange (ETDEWEB)

Dijk, Marc van; Visscher, Koen M.; Kastritis, Panagiotis L.; Bonvin, Alexandre M. J. J., E-mail: a.m.j.j.bonvin@uu.nl [Utrecht University, Bijvoet Center for Biomolecular Research, Faculty of Science-Chemistry (Netherlands)

2013-05-15

Interfacial water molecules play an important role in many aspects of protein-DNA specificity and recognition. Yet they have been mostly neglected in the computational modeling of these complexes. We present here a solvated docking protocol that allows explicit inclusion of water molecules in the docking of protein-DNA complexes and demonstrate its feasibility on a benchmark of 30 high-resolution protein-DNA complexes containing crystallographically-determined water molecules at their interfaces. Our protocol is capable of reproducing the solvation pattern at the interface and recovers hydrogen-bonded water-mediated contacts in many of the benchmark cases. Solvated docking leads to an overall improvement in the quality of the generated protein-DNA models for cases with limited conformational change of the partners upon complex formation. The applicability of this approach is demonstrated on real cases by docking a representative set of 6 complexes using unbound protein coordinates, model-built DNA and knowledge-based restraints. As HADDOCK supports the inclusion of a variety of NMR restraints, solvated docking is also applicable for NMR-based structure calculations of protein-DNA complexes.

Proceedings of the workshop on crystalline ion beams

International Nuclear Information System (INIS)

Hasse, R.W.; Hofmann, I.; Liesen, D.

1989-04-01

The workshop consisted of mainly invited and some contributed papers. More informal discussions took place in three working groups on the following topics: beam cooling techniques; diagnostics of crystalline beams; storage rings for crystalline beams. The present volume collects all papers as well as the summaries of the working groups. See hints under the relevant topics. (orig./HSI)

The etiology of human age-related cataract. Proteins don't last forever.

Science.gov (United States)

Truscott, Roger J W; Friedrich, Michael G

2016-01-01

It is probable that the great majority of human cataract results from the spontaneous decomposition of long-lived macromolecules in the human lens. Breakdown/reaction of long-lived proteins is of primary importance and recent proteomic analysis has enabled the identification of the particular crystallins, and their exact sites of amino acid modification. Analysis of proteins from cataractous lenses revealed that there are sites on some structural proteins that show a consistently greater degree of deterioration than age-matched normal lenses. The most abundant posttranslational modification of aged lens proteins is racemization. Deamidation, truncation and crosslinking, each arising from the spontaneous breakdown of susceptible amino acids within proteins, are also present. Fundamental to an understanding of nuclear cataract etiology, it is proposed that once a certain degree of modification at key sites occurs, that protein-protein interactions are disrupted and lens opacification ensues. Since long-lived proteins are now recognized to be present in many other sites of the body, such as the brain, the information gleaned from detailed analyses of degraded proteins from aged lenses will apply more widely to other age-related human diseases. This article is part of a Special Issue entitled Crystallin Biochemistry in Health and Disease. Copyright © 2015 Elsevier B.V. All rights reserved.

Electrochemical synthesis of highly crystalline copper nanowires

International Nuclear Information System (INIS)

Kaur, Amandeep; Gupta, Tanish; Kumar, Akshay; Kumar, Sanjeev; Singh, Karamjeet; Thakur, Anup

2015-01-01

Copper nanowires were fabricated within the pores of anodic alumina template (AAT) by template synthesis method at pH = 2.9. X-ray diffraction (XRD), scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) were used to investigate the structure, morphology and composition of fabricated nanowires. These characterizations revealed that the deposited copper nanowires were highly crystalline in nature, dense and uniform. The crystalline copper nanowires are promising in application of future nanoelectronic devices and circuits

Mesomorphous versus traces of crystallinity: The itraconazole example

Energy Technology Data Exchange (ETDEWEB)

Atassi, Faraj, E-mail: fatassi@yahoo.com; Behme, Robert J.; Patel, Phenil J.

2013-12-20

Highlights: • Characterizing partially disordered pharmaceuticals is very challenging due to the fact that more than one discrete disordered phase can be present. • Dynamic mechanical analysis and dielectric analysis are extremely helpful in characterizing pharmaceutical mesophases (liquid crystals). • Thermotropic pharmaceutical mesophases, often mistaken as amorphous or partially crystalline, can show different phases of liquid crystallinity at different temperature. • Liquid crystalline pharmaceutical materials often show amorphous behavior along with other characteristics specific to mesomorphous materials. • The thermal and mechanical history of pharmaceutical disordered samples has a significant effect on their phase composition. - Abstract: Characterizing disordered pharmaceutical materials can be challenging, especially materials with partially disordered structures that lose one or two directional order (mesophases) and do not fit the traditional characterization categories of amorphous, crystalline or a combination of the two. Itraconazole, an antifungal agent, was chosen as a model compound that, when quench cooled, exhibits atypical disordered structure. Five different analytical tools were used to map out the molecular structure of this material and how it changes with changing temperature. X-ray diffraction showed some remnant crystallinity while dielectric analysis, dynamic mechanical analysis, DSC and hot stage microscopy gave more detailed molecular structure of the disordered material and explained all temperature related structural changes. The characterization of mesomorphous Itraconazole described here will help characterize a wide range of pharmaceuticals that exhibit thermotropic (temperature induced) mesomorphism at the molecular level.

Mesomorphous versus traces of crystallinity: The itraconazole example

International Nuclear Information System (INIS)

Atassi, Faraj; Behme, Robert J.; Patel, Phenil J.

2013-01-01

Highlights: • Characterizing partially disordered pharmaceuticals is very challenging due to the fact that more than one discrete disordered phase can be present. • Dynamic mechanical analysis and dielectric analysis are extremely helpful in characterizing pharmaceutical mesophases (liquid crystals). • Thermotropic pharmaceutical mesophases, often mistaken as amorphous or partially crystalline, can show different phases of liquid crystallinity at different temperature. • Liquid crystalline pharmaceutical materials often show amorphous behavior along with other characteristics specific to mesomorphous materials. • The thermal and mechanical history of pharmaceutical disordered samples has a significant effect on their phase composition. - Abstract: Characterizing disordered pharmaceutical materials can be challenging, especially materials with partially disordered structures that lose one or two directional order (mesophases) and do not fit the traditional characterization categories of amorphous, crystalline or a combination of the two. Itraconazole, an antifungal agent, was chosen as a model compound that, when quench cooled, exhibits atypical disordered structure. Five different analytical tools were used to map out the molecular structure of this material and how it changes with changing temperature. X-ray diffraction showed some remnant crystallinity while dielectric analysis, dynamic mechanical analysis, DSC and hot stage microscopy gave more detailed molecular structure of the disordered material and explained all temperature related structural changes. The characterization of mesomorphous Itraconazole described here will help characterize a wide range of pharmaceuticals that exhibit thermotropic (temperature induced) mesomorphism at the molecular level

Soil Crystallinity As a Climate Indicator: Field Experiments on Earth and Mars

Science.gov (United States)

Horgan, Briony; Scudder, Noel; Rampe, Elizabeth; Rutledge, Alicia

2016-01-01

Soil crystallinity is largely determined by leaching rates, as high leaching rates favor the rapid precipitation of short order or poorly-crystalline phases like the aluminosilicate allophane. High leaching rates can occur due to high precipitation rates, seasonal monsoons, or weathering of glass, but are also caused by the rapid onset of seasonal melting of snow and ice in cold environments. Thus, cold climate soils are commonly dominated by poorly crystalline phases, which mature into kaolin minerals over time. Thus, we hypothesize that, in some contexts, soils with high abundances of poorly crystalline phases could indicate formation under cold climatic conditions. This model could be helpful in interpreting the poorly-constrained paleoclimate of ancient Mars, as the crystallinity of ancient soils and soil-derived sediments appears to be highly variable in time and space. While strong signatures of crystalline phyllosilicates have been identified in possible ancient paleosols on Mars, Mars Science Laboratory rover investigations of diverse ancient sediments at Gale Crater has shown that they can contain very high abundances (40-50 wt%) of poorly crystalline phases. We hypothesize that these poorly crystalline phases could be the result of weathering by ice/snow melt, perhaps providing support for sustained cold climates on early Mars punctuated by more limited warm climates. Furthermore, such poorly crystalline soils could be highly fertile growth media for future human exploration and colonization on Mars. To test this hypothesis, we are currently using rover-like instrumentation to investigate the mineralogy and chemistry of weathering products generated by snow and ice melt in a Mars analog alpine environment: the glaciated Three Sisters volcanic complex in central Oregon. Alteration in this glacial environment generates high abundances of poorly crystalline phases, many of which have compositions distinct from those identified in previous terrestrial

Gamma-irradiation effects to posttranslational modification and chaperon function of bovine α-crystalline

International Nuclear Information System (INIS)

Hiroki, K; Matsumoto, S.; Awakura, M.; Fujii, N.

2001-01-01

The formation of D-asparate (D-Asp) in αA-crystallin of the aged human eye and the cataract crystalline lens has been reported. Crystalline lens keeps the transparency by forming α-crystallin which consists of a high order association of αA-and αB-crystallin. Bovine α-crystallin for investigating a chaperone function which protects the crystalline lens from getting to opaque or disordered agglutination with heat or light, is irradiated by gamma-ray (Co-60) at 0, 1, 2, 3, and 4 kGy, respectively. The irradiated bovine α-crystallin are analyzed with electrophoresis, gel permeation chromatograph, and UV absorption spectrometer for checking on the agglutination and the isomerization of macromolecules. Oxidation of methionine residues (Met-1) and isomerization of asparagine residues (Asp-151) in the αA-crystallin are ascertained in molecular levels with reversed phase liquid chromatography. The Met-1 oxidation and the Asp-151 isomerization depend on gamma-irradiation doses. It is thought that OH radical and H radical in water generated by the irradiation lead to the oxidation and the isomerization. Stereoinversion in the α-crystallin following to such a chemical change are considered to lead to the agglutination of polymer and the reduction of chaperon function. (M. Suetake)

21 CFR 524.2620 - Liquid crystalline trypsin, Peru balsam, castor oil.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Liquid crystalline trypsin, Peru balsam, castor... NEW ANIMAL DRUGS § 524.2620 Liquid crystalline trypsin, Peru balsam, castor oil. (a)(1) Specifications... delivered to the wound site contains 0.12 milligram of crystalline trypsin, 87.0 milligrams of Peru balsam...

Characterization and use of crystalline bacterial cell surface layers

Science.gov (United States)

Sleytr, Uwe B.; Sára, Margit; Pum, Dietmar; Schuster, Bernhard

2001-10-01

Crystalline bacterial cell surface layers (S-layers) are one of the most common outermost cell envelope components of prokaryotic organisms (archaea and bacteria). S-layers are monomolecular arrays composed of a single protein or glycoprotein species and represent the simplest biological membranes developed during evolution. S-layers as the most abundant of prokaryotic cellular proteins are appealing model systems for studying the structure, synthesis, genetics, assembly and function of proteinaceous supramolecular structures. The wealth of information existing on the general principle of S-layers have revealed a broad application potential. The most relevant features exploited in applied S-layer research are: (i) pores passing through S-layers show identical size and morphology and are in the range of ultrafiltration membranes; (ii) functional groups on the surface and in the pores are aligned in well-defined positions and orientations and accessible for chemical modifications and binding functional molecules in very precise fashion; (iii) isolated S-layer subunits from a variety of organisms are capable of recrystallizing as closed monolayers onto solid supports (e.g., metals, polymers, silicon wafers) at the air-water interface, on lipid films or onto the surface of liposomes; (iv) functional domains can be incorporated in S-layer proteins by genetic engineering. Thus, S-layer technologies particularly provide new approaches for biotechnology, biomimetics, molecular nanotechnology, nanopatterning of surfaces and formation of ordered arrays of metal clusters or nanoparticles as required for nanoelectronics.

Muscle plasticity related to changes in tubulin and αB-crystallin levels induced by eccentric contraction in rat skeletal muscles.

Science.gov (United States)

Jee, H; Ochi, E; Sakurai, T; Lim, J-Y; Nakazato, K; Hatta, H

2016-09-01

We used the model of eccentric contraction of the hindlimb muscle by Ochi et al. to examine the role of eccentric contraction in muscle plasticity. This model aims to focus on stimulated skeletal muscle responses by measuring tissue weights and tracing the quantities of αB-crystallin and tubulin. The medial gastrocnemius muscle (GCM) responded to electrically induced eccentric contraction (EIEC) with significant increases in tissue weight (p muscle weight after EIEC. EIEC in the GCM caused contractile-induced sustenance of the traced proteins, but the soleus muscle exhibited a remarkable decrease in α-tubulin and a 19% decrease in αB-crystallin. EIEC caused fast-to-slow myosin heavy chain (MHC) isoform type-oriented shift within both the GCM and soleus muscle. These results have shown that different MHC isoform type-expressing slow and fast muscles commonly undergo fast-to-slow type MHC isoform transformation. This suggests that different levels of EIEC affected each of the slow and fast muscles to induce different quantitative changes in the expression of αB-crystallin and α-tubulin.

Biosynthesis and characterization of typical fibroin crystalline polypeptides of silkworm Bombyx mori

Energy Technology Data Exchange (ETDEWEB)

Wang Jiannan, E-mail: wangjn@suda.edu.cn [College of Material Engineering, Soochow University, Suzhou 215021 (China); Yan Shuqin [College of Material Engineering, Soochow University, Suzhou 215021 (China); Lu Changde [Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Bai Lun [College of Material Engineering, Soochow University, Suzhou 215021 (China)

2009-05-05

We aimed to investigate the self-organization/self-assembly mechanisms of silkworm fibroin-based material. In the present study, for the first time, we designed and multimerized four DNA 'monomer' sequences from structurally simple fibroin crystalline peptides or analog, [GAGAGX] (X = A, S, Y and V) to encode polypeptides [GAGAGX]{sub 16} (eGA, eGS, eGY and eGV) using a 'head-to-tail' construction strategy. Multimers were cloned into pGEX-KG and fusion proteins GST-[GAGAGX]{sub 16} (KGA, KGS, KGY and KGV) were efficiently expressed in Escherichia coli. These fusion proteins were isolated and purified by GST affinity chromatography and confirmed by SDS-PAGE and Western blot analysis using antibody reactive to GST. The polypeptides were cleavaged from GST fusion proteins by digesting with thrombin enzyme. The composition of the four polypeptides was confirmed by composition analysis of amino acids, and their abilities to form {beta}-sheet structure were determined by ThT fluorescence spectral analysis. The content of {beta}-sheet among the four polypeptides followed the order: eGS > eGV > eGY > eGA.

Effect of inclusion of hydroxycinnamic and chlorogenic acids from green coffee bean in β-cyclodextrin on their interactions with whey, egg white and soy protein isolates.

Science.gov (United States)

Budryn, Grażyna; Pałecz, Bartłomiej; Rachwał-Rosiak, Danuta; Oracz, Joanna; Zaczyńska, Donata; Belica, Sylwia; Navarro-González, Inmaculada; Meseguer, Josefina María Vegara; Pérez-Sánchez, Horacio

2015-02-01

The aim of the study was to characterise the interactions of hydroxycinnamic and chlorogenic acids (CHAs) from green coffee, with isolates of proteins from egg white (EWP), whey (WPC) and soy (SPI), depending on pH and temperature. The binding degree was determined by liquid chromatography coupled to a diode array detector and an ultrahigh resolution hybrid quadruple-time-of-flight mass spectrometer with ESI source (LC-QTOF-MS/MS). As a result of binding, the concentration of CHAs in proteins ranged from 9.44-12.2, 11.8-13.1 and 12.1-14.4g/100g for SPI, WPC and EWP, respectively. Thermodynamic parameters of protein-ligand interactions were determined by isothermal titration calorimetry (ITC) and energetics of interactions at the atomic level by molecular modelling. The amount of CHAs released during proteolytic digestion was in the range 0.33-2.67g/100g. Inclusion of CHAs with β-cyclodextrin strongly limited these interactions to a level of 0.03-0.06g/100g. Copyright © 2014 Elsevier Ltd. All rights reserved.

Inclusive and semi-inclusive analysis from polarised deep-inelastic muon scattering

International Nuclear Information System (INIS)

Kageya, T.

1999-01-01

We present new results for the spin-dependent structure function on the proton and for the polarised quark distributions in the nucleon from semi-inclusive spin asymmetries. With the inclusive asymmetry from deep inelastic scattering of polarised muons on polarised protons, it is found that the Ellis-Jaffe sum rule is violated. Using our results for Γ d 1 , the Bjorken sum rule is confirmed with an accuracy of about 15%. From semi-inclusive spin asymmetries and SMC inclusive spin asymmetries, we determine the polarised quark distributions of valence u and d quarks to be positive and negative, respectively, while the non-strange sea distribution to be consistent with zero

The Chlamydia type III secretion system C-ring engages a chaperone-effector protein complex.

Directory of Open Access Journals (Sweden)

Kris E Spaeth

2009-09-01

Full Text Available In Gram-negative bacterial pathogens, specialized chaperones bind to secreted effector proteins and maintain them in a partially unfolded form competent for translocation by type III secretion systems/injectisomes. How diverse sets of effector-chaperone complexes are recognized by injectisomes is unclear. Here we describe a new mechanism of effector-chaperone recognition by the Chlamydia injectisome, a unique and ancestral line of these evolutionarily conserved secretion systems. By yeast two-hybrid analysis we identified networks of Chlamydia-specific proteins that interacted with the basal structure of the injectisome, including two hubs of protein-protein interactions that linked known secreted effector proteins to CdsQ, the putative cytoplasmic C-ring component of the secretion apparatus. One of these protein-interaction hubs is defined by Ct260/Mcsc (Multiple cargo secretion chaperone. Mcsc binds to and stabilizes at least two secreted hydrophobic proteins, Cap1 and Ct618, that localize to the membrane of the pathogenic vacuole ("inclusion". The resulting complexes bind to CdsQ, suggesting that in Chlamydia, the C-ring of the injectisome mediates the recognition of a subset of inclusion membrane proteins in complex with their chaperone. The selective recognition of inclusion membrane proteins by chaperones may provide a mechanism to co-ordinate the translocation of subsets of inclusion membrane proteins at different stages in infection.

Effect of inclusion of citrus pulp inxaraés grass silage

Directory of Open Access Journals (Sweden)

Júnior Issamu Yasuoka

2016-01-01

Full Text Available The objective of this study was to evaluate the effect of inclusion of different levels of pelleted citrus pulp (PCP on the quality of xaraés grass silage. Xaraés grass was ensiled at 54 days of growth with 0, 10, 20 and 30% PCP and divided into 20 experimental silos (five repetitions/treatment. A completely randomized design was adopted. The silos were opened after 67 days for the determination of dry matter (DM, crude protein (CP, neutral detergent fiber (NDF, acid detergent fiber (ADF, pH, and titratable acidity of the silages. The inclusion of PCP did not influence DM content, but reduced CP content. NDF and ADF content decreased with the addition of citrus pulp. Regarding pH, a significant difference was only observed for the inclusion of 30% PCP. Titratable acidity in the silage was lower in the absence of PCP (0% compared to the treatment with 30% (P<0.05. The inclusion of 30% PCP in xaraés grass silage is indicated since it improves the quality of the fibrous fraction, while maintaining acceptable pH values.

Crystalline to amorphous transformation in silicon

International Nuclear Information System (INIS)

Cheruvu, S.M.

1982-09-01

In the present investigation, an attempt was made to understand the fundamental mechanism of crystalline-to-amorphous transformation in arsenic implanted silicon using high resolution electron microscopy. A comparison of the gradual disappearance of simulated lattice fringes with increasing Frenkel pair concentration with the experimental observation of sharp interfaces between crystalline and amorphous regions was carried out leading to the conclusion that when the defect concentration reaches a critical value, the crystal does relax to an amorphous state. Optical diffraction experiments using atomic models also supported this hypothesis. Both crystalline and amorphous zones were found to co-exist with sharp interfaces at the atomic level. Growth of the amorphous fraction depends on the temperature, dose rate and the mass of the implanted ion. Preliminary results of high energy electron irradiation experiments at 1.2 MeV also suggested that clustering of point defects occurs near room temperature. An observation in a high resolution image of a small amorphous zone centered at the core of a dislocation is presented as evidence that the nucleation of an amorphous phase is heterogeneous in nature involving clustering or segregation of point defects near existing defects

A Protein Aggregation Inhibitor, Leuco-Methylthioninium Bis(Hydromethanesulfonate, Decreases α-Synuclein Inclusions in a Transgenic Mouse Model of Synucleinopathy

Directory of Open Access Journals (Sweden)

Karima Schwab

2018-01-01

Full Text Available α-Synuclein (α-Syn aggregation is a pathological feature of synucleinopathies, neurodegenerative disorders that include Parkinson’s disease (PD. We have tested whether N,N,N′,N′-tetramethyl-10H-phenothiazine-3,7-diaminium bis(hydromethanesulfonate (leuco-methylthioninium bis(hydromethanesulfonate; LMTM, a tau aggregation inhibitor, affects α-Syn aggregation in vitro and in vivo. Both cellular and transgenic models in which the expression of full-length human α-Syn (h-α-Syn fused with a signal sequence peptide to promote α-Syn aggregation were used. Aggregated α-Syn was observed following differentiation of N1E-115 neuroblastoma cells transfected with h-α-Syn. The appearance of aggregated α-Syn was inhibited by LMTM, with an EC50 of 1.1 μM, with minimal effect on h-α-Syn mRNA levels being observed. Two independent lines of mice (L58 and L62 transgenic for the same fusion protein accumulated neuronal h-α-Syn that, with aging, developed into fibrillary inclusions characterized by both resistance to proteinase K (PK-cleavage and their ability to bind thiazin red. There was a significant decrease in α-Syn-positive neurons in multiple brain regions following oral treatment of male and female mice with LMTM administered daily for 6 weeks at 5 and 15 mg MT/kg. The early aggregates of α-Syn and the late-stage fibrillar inclusions were both susceptible to inhibition by LMTM, a treatment that also resulted in the rescue of movement and anxiety-related traits in these mice. The results suggest that LMTM may provide a potential disease modification therapy in PD and other synucleinopathies through the inhibition of α-Syn aggregation.

Used Fuel Disposition in Crystalline Rocks: FY16 Progress Report

Energy Technology Data Exchange (ETDEWEB)

Wang, Y. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Hadgu, Teklu [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Kalinina, Elena Arkadievna [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Jerden, James L. [Argonne National Lab. (ANL), Argonne, IL (United States); Copple, Jacqueline M. [Argonne National Lab. (ANL), Argonne, IL (United States); Cruse, T. [Argonne National Lab. (ANL), Argonne, IL (United States); Ebert, W. [Argonne National Lab. (ANL), Argonne, IL (United States); Buck, E. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Eittman, R. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Tinnacher, R. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Tournassat, Christophe [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Davis, J. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Viswanathan, H. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Chu, S. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Dittrich, T. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Hyman, F. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Karra, S. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Makedonska, N. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Reimus, P. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Zavarin, Mavrik [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Joseph, C. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

2016-09-21

The objective of the Crystalline Disposal R&D Work Package is to advance our understanding of long-term disposal of used fuel in crystalline rocks and to develop necessary experimental and computational capabilities to evaluate various disposal concepts in such media. FY16 continued to be a successful year in both experimental and modeling arenas in evaluation of used fuel disposal in crystalline rocks. The work covers a wide range of research topics identified in the R&D plan.

Inclusive pedagogy

DEFF Research Database (Denmark)

Korsgaard, Morten Timmermann; Skov Mortensen, Stig

This article will present a case for a shift in perspective in inclusive education research towards a continentally inspired approach. Drawing on the age old distinction between continental and Anglo-American educational research the aim is to flesh out what a shift to a continental approach...... will entail, and why it might be beneficial to research in inclusive education...

The effect of crystallinity on cell growth in semi-crystalline microcellular foams by solid-state process: modeling and numerical simulation

Science.gov (United States)

Rezvanpanah, Elham; Ghaffarian Anbaran, S. Reza

2017-11-01

This study establishes a model and simulation scheme to describe the effect of crystallinity as one of the most effective parameters on cell growth phenomena in a solid batch foaming process. The governing model of cell growth dynamics, based on the well-known ‘Cell model’, is attained in details. To include the effect of crystallinity in the model, the properties of the polymer/gas mixtures (i.e. solubility, diffusivity, surface tension and viscosity) are estimated by modifying relations to consider the effect of crystallinity. A finite element-finite difference (FEFD) method is employed to solve the highly nonlinear and coupled equations of cell growth dynamics. The proposed simulation is able to evaluate all properties of the system at the given process condition and uses them to calculate the cell size, pressure and gas concentration gradient with time. A high-density polyethylene/nitrogen (HDPE/N2) system is used herein as a case study. Comparing the simulation results with the others works and experimental results verify the accuracy of the simulation scheme. The cell growth is a complicated combination of several phenomena. This study attempted to reach a better understanding of cell growth trend, driving and retarding forces and the effect of crystallinity on them.

Liquid crystalline epoxy nanocomposite material for dental application.

Science.gov (United States)

Tai, Yun-Yuan; Hsu, Sheng-Hao; Chen, Rung-Shu; Su, Wei-Fang; Chen, Min-Huey

2015-01-01

Novel liquid crystalline epoxy nanocomposites, which exhibit reduced polymerization shrinkage and effectively bond to tooth structures, can be applied in esthetic dentistry, including core and post systems, direct and indirect restorations, and dental brackets. The purposes of this study were to investigate the properties of liquid crystalline epoxy nanocomposites including biocompatibility, microhardness, and frictional forces of bracket-like blocks with different filler contents for further clinical applications. In this study, we evaluated liquid crystalline epoxy nanocomposite materials that exhibited various filler contents, by assessing their cell activity performance using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and their microhardness with or without thermocycling. We also evaluated the frictional force between bracket-like duplicates and commercially available esthetic bracket systems using Instron 5566. The liquid crystalline epoxy nanocomposite materials showed good biocompatibility. The materials having high filler content demonstrated greater microhardness compared with commercially available bracket materials, before and after the thermocycling treatment. Thus, manufacturing processes are important to reduce frictional force experienced by orthodontic brackets. The microhardness of the bracket-like blocks made by our new material is superior to the commercially available brackets, even after thermocycling. Our results indicate that the evaluated liquid crystalline epoxy nanocomposite materials are of an appropriate quality for application in dental core and post systems and in various restorations. By applying technology to refine manufacturing processes, these new materials could also be used to fabricate esthetic brackets for orthodontic treatment. Copyright © 2014. Published by Elsevier B.V.

Nanomembrane structures having mixed crystalline orientations and compositions

Science.gov (United States)

Lagally, Max G.; Scott, Shelley A.; Savage, Donald E.

2014-08-12

The present nanomembrane structures include a multilayer film comprising a single-crystalline layer of semiconductor material disposed between two other single-crystalline layers of semiconductor material. A plurality of holes extending through the nanomembrane are at least partially, and preferably entirely, filled with a filler material which is also a semiconductor, but which differs from the nanomembrane semiconductor materials in composition, crystal orientation, or both.

Cellulose binding domain fusion proteins

Science.gov (United States)

Shoseyov, Oded; Shpiegl, Itai; Goldstein, Marc A.; Doi, Roy H.

1998-01-01

A cellulose binding domain (CBD) having a high affinity for crystalline cellulose and chitin is disclosed, along with methods for the molecular cloning and recombinant production thereof. Fusion products comprising the CBD and a second protein are likewise described. A wide range of applications are contemplated for both the CBD and the fusion products, including drug delivery, affinity separations, and diagnostic techniques.

Circuit design techniques for non-crystalline semiconductors

CERN Document Server

Sambandan, Sanjiv

2012-01-01

Despite significant progress in materials and fabrication technologies related to non-crystalline semiconductors, fundamental drawbacks continue to limit real-world application of these devices in electronic circuits. To help readers deal with problems such as low mobility and intrinsic time variant behavior, Circuit Design Techniques for Non-Crystalline Semiconductors outlines a systematic design approach, including circuit theory, enabling users to synthesize circuits without worrying about the details of device physics. This book: Offers examples of how self-assembly can be used as a powerf

Soft and Hard Textured Wheat Differ in Starch Properties as Indicated by Trimodal Distribution, Morphology, Thermal and Crystalline Properties.

Directory of Open Access Journals (Sweden)

Rohit Kumar

Full Text Available Starch and proteins are major components in the wheat endosperm that affect its end product quality. Between the two textural classes of wheat i.e. hard and soft, starch granules are loosely bound with the lipids and proteins in soft wheat due to higher expression of interfering grain softness proteins. It might have impact on starch granules properties. In this work for the first time the physiochemical and structural properties of different sized starch granules (A-, B- and C-granules were studied to understand the differences in starches with respect to soft and hard wheat. A-, B- and C-type granules were separated with >95% purity. Average number and proportion of A-, B-, and C-type granules was 18%, 56%, 26% and 76%, 19%, 5% respectively. All had symmetrical birefringence pattern with varied intensity. All displayed typical A-type crystallites. A-type granules also showed V-type crystallinity that is indicative of starch complexes with lipids and proteins. Granules differing in gelatinization temperature (ΔH and transition temperature (ΔT, showed different enthalpy changes during heating. Substitution analysis indicated differences in relative substitution pattern of different starch granules. Birefringence, percentage crystallinity, transmittance, gelatinization enthalpy and substitution decreased in order of A>B>C being higher in hard wheat than soft wheat. Amylose content decreased in order of A>B>C being higher in soft wheat than hard wheat. Reconstitution experiment showed that starch properties could be manipulated by changing the composition of starch granules. Addition of A-granules to total starch significantly affected its thermal properties. Effect of A-granule addition was higher than B- and C-granules. Transmittance of the starch granules paste showed that starch granules of hard wheat formed clear paste. These results suggested that in addition to differences in protein concentration, hard and soft wheat lines have

Synthesis and crystalline properties of CdS incorporated polyvinylidene fluoride (PVDF) composite film

Science.gov (United States)

Patel, Arunendra Kumar; Sunder, Aishwarya; Mishra, Shweta; Bajpai, Rakesh

2018-05-01

This paper gives an insight on the synthesis and crystalline properties of Polyvinylidene Fluoride (PVDF) (host matrix) composites impregnated with Cadmium Sulphide (CdS) using Dimethyl formamide (DMF) as the base, prepared by the well known solvent casting technique. The effect of doping concentration of CdS in to the PVDF matrix was studied using X-ray diffraction technique. The structural properties like crystallinity Cr, interplanar distance d, average size of the crystalline region (D), and average inter crystalline separation (R) have been estimated for the developed composite. The crystallinity index, crystallite size and inter crystalline separation is increasing with increase in the concentration of CdS in to the PVDF matrix while the interplanar distance d is decreasing.

A Systematic Protein Refolding Screen Method using the DGR Approach Reveals that Time and Secondary TSA are Essential Variables

NARCIS (Netherlands)

Wang, Yuanze; van Oosterwijk, Niels; Ali, Ameena M; Adawy, Alaa; Anindya, Atsarina L; Dömling, Alexander S S; Groves, Matthew R

2017-01-01

Refolding of proteins derived from inclusion bodies is very promising as it can provide a reliable source of target proteins of high purity. However, inclusion body-based protein production is often limited by the lack of techniques for the detection of correctly refolded protein. Thus, the

BAG3 Is a Modular, Scaffolding Protein that physically Links Heat Shock Protein 70 (Hsp70) to the Small Heat Shock Proteins.

Science.gov (United States)

Rauch, Jennifer N; Tse, Eric; Freilich, Rebecca; Mok, Sue-Ann; Makley, Leah N; Southworth, Daniel R; Gestwicki, Jason E

2017-01-06

Small heat shock proteins (sHsps) are a family of ATP-independent molecular chaperones that are important for binding and stabilizing unfolded proteins. In this task, the sHsps have been proposed to coordinate with ATP-dependent chaperones, including heat shock protein 70 (Hsp70). However, it is not yet clear how these two important components of the chaperone network are linked. We report that the Hsp70 co-chaperone, BAG3, is a modular, scaffolding factor to bring together sHsps and Hsp70s. Using domain deletions and point mutations, we found that BAG3 uses both of its IPV motifs to interact with sHsps, including Hsp27 (HspB1), αB-crystallin (HspB5), Hsp22 (HspB8), and Hsp20 (HspB6). BAG3 does not appear to be a passive scaffolding factor; rather, its binding promoted de-oligomerization of Hsp27, likely by competing for the self-interactions that normally stabilize large oligomers. BAG3 bound to Hsp70 at the same time as Hsp22, Hsp27, or αB-crystallin, suggesting that it might physically bring the chaperone families together into a complex. Indeed, addition of BAG3 coordinated the ability of Hsp22 and Hsp70 to refold denatured luciferase in vitro. Together, these results suggest that BAG3 physically and functionally links Hsp70 and sHsps. Copyright © 2016 Elsevier Ltd. All rights reserved.

Inclusion: Conceptualization and measurement

NARCIS (Netherlands)

Jansen, W.S.; Otten, S.; Van der Zee, K.I.; Jans, L.

2014-01-01

n the present research, we introduced a conceptual framework of inclusion and subsequently used this as a starting point to develop and validate a scale to measure perceptions of inclusion. Departing from existing work on inclusion and complementing this with theoretical insights from optimal

Inclusion : Conceptualization and measurement

NARCIS (Netherlands)

Jansen, Wiebren S.; Otten, Sabine; van der Zee, Karen I.; Jans, Lise

In the present research, we introduced a conceptual framework of inclusion and subsequently used this as a starting point to develop and validate a scale to measure perceptions of inclusion. Departing from existing work on inclusion and complementing this with theoretical insights from optimal

Inclusion: Conceptualization and measurement

NARCIS (Netherlands)

Jansens, Wiebren S.; Otten, Sabine; van der Zee, Karen; Jans, Lise

2014-01-01

In the present research, we introduced a conceptual framework of inclusion and subsequently used this as a starting point to develop and validate a scale to measure perceptions of inclusion. Departing from existing work on inclusion and complementing this with theoretical insights from optimal

On the determination of crystallinity and cellulose content in plant fibres

DEFF Research Database (Denmark)

Thygesen, Anders; Oddershede, Jette; Lilholt, Hans

2005-01-01

A comparative study of cellulose crystallinity based on the sample crystallinity and the cellulose content in plant fibres was performed for samples of different origin. Strong acid hydrolysis was found superior to agricultural fibre analysis and comprehensive plant fibre analysis for a consistent...... determination of the cellulose content. Crystallinity determinations were based on X-ray powder diffraction methods using side-loaded samples in reflection (Bragg-Brentano) mode. Rietveld refinements based on the recently published crystal structure of cellulose I beta followed by integration of the crystalline...... and 60 - 70 g/ 100 g cellulose in wood based fibres. These findings are significant in relation to strong fibre composites and bio-ethanol production....

Liquid crystallinity driven highly aligned large graphene oxide composites

Energy Technology Data Exchange (ETDEWEB)

Lee, Kyung Eun; Oh, Jung Jae; Yun, Taeyeong [Center for Nanomaterials and Chemical Reactions, Institute for Basic Science (IBS), Daejeon 305-701 (Korea, Republic of); Department of Chemistry, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701 (Korea, Republic of); Kim, Sang Ouk, E-mail: sangouk.kim@kaist.ac.kr [Center for Nanomaterials and Chemical Reactions, Institute for Basic Science (IBS), Daejeon 305-701 (Korea, Republic of); Department of Chemistry, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701 (Korea, Republic of)

2015-04-15

Graphene is an emerging graphitic carbon materials, consisting of sp{sup 2} hybridized two dimensinal honeycomb structure. It has been widely studied to incorporate graphene with polymer to utilize unique property of graphene and reinforce electrical, mechanical and thermal property of polymer. In composite materials, orientation control of graphene significantly influences the property of composite. Until now, a few method has been developed for orientation control of graphene within polymer matrix. Here, we demonstrate facile fabrication of high aligned large graphene oxide (LGO) composites in polydimethylsiloxane (PDMS) matrix exploiting liquid crystallinity. Liquid crystalline aqueous dispersion of LGO is parallel oriented within flat confinement geometry. Freeze-drying of the aligned LGO dispersion and subsequent infiltration with PDMS produce highly aligned LGO/PDMS composites. Owing to the large shape anisotropy of LGO, liquid crystalline alignment occurred at low concentration of 2 mg/ml in aqueous dispersion, which leads to the 0.2 wt% LGO loaded composites. - Graphical abstract: Liquid crystalline LGO aqueous dispersions are spontaneous parallel aligned between geometric confinement for highly aligned LGO/polymer composite fabrication. - Highlights: • A simple fabrication method for highly aligned LGO/PDMS composites is proposed. • LGO aqueous dispersion shows nematic liquid crystalline phase at 0.8 mg/ml. • In nematic phase, LGO flakes are highly aligned by geometric confinement. • Infiltration of PDMS into freeze-dried LGO allows highly aligned LGO/PDMS composites.

pH induced protein-scaffold biosynthesis of tunable shape gold nanoparticles

International Nuclear Information System (INIS)

Zhang Xiaorong; He Xiaoxiao; Wang Kemin; Ren Fang; Qin Zhihe

2011-01-01

In this paper, a pH-inductive protein-scaffold biosynthesis of shape-tunable crystalline gold nanoparticles at room temperature has been developed. By simple manipulation of the reaction solution's pH, anisotropic gold nanoparticles including spheres, triangles and cubes could be produced by incubating an aqueous solution of sodium tetrachloroaurate with Dolichomitriopsis diversiformis biomasses after immersion in ultrapure Millipore water overnight. A moss protein with molecular weight of about 71 kDa and pI of 4.9 was the primary biomolecule involved in the biosynthesis of gold nanoparticles. The secondary configuration of the proteins by CD spectrum implied that the moss protein could display different secondary configurations including random coil, α-helix and intermediate conformations between random coil and α-helix for the experimental pH solution. The growth process of gold nanoparticles further showed that the moss protein with different configurations provided the template scaffold for the shape-controlled biosynthesis of gold nanoparticles. The constrained shape of the gold nanoparticles, however, disappeared in boiled moss extract. The gold nanoparticles with designed morphology were successfully reconstructed using the moss protein purified from the gold nanoparticles. Structural characterizations by SEM, TEM and SAED showed that the triangular and cubic gold nanoparticles were single crystalline.

New photo-convertible reactions of blue-fluorescent calf α-crystallin

International Nuclear Information System (INIS)

Fujimori, E.

1979-01-01

Both native blue fluorescent α-crystalline from calf lenses and UV (300 nm)-irradiated blue-fluorescent α-crystalline, when further irradiated with 365 nm-UV light, produce photo-products capable of emitting a new fluorescence at 455 nm. Illumination of the photo-products with 420 nm visible light regenerates the original fluorescence at 420-425 nm. In addition, another fluorescence at 400 nm has also been found in UV (300 nm)-irradiated blue-fluorescent α-crystallin, when exposed to 365 nm-UV light. (author)

Effects of γ-irradiation and thermal treatment of crystallinity of drawn HDPE

International Nuclear Information System (INIS)

Liu Zhanjun; Silverman, J.

1997-01-01

The effect of absorbed dose irradiated in vacuum and air on the crystallinity of drawn HDPE was studied. Experimental results show that up to 250 kGy of absorbed dose when irradiated in vacuum, the crystallinity of drawn HDPE is decreased from about 75% to about 71%, and then the increase of absorbed dose until 1000 kGy has no further effect in lowering the crystallinity; when irradiated in air, an absorbed dose of 1000 kGy has no effect on the crystallinity of drawn HDPE. The effect of temperature of thermal treatment on the crystallinity of unirradiated drawn HDPE was also investigated. At first, the crystallinity is increased with the increase of temperature of thermal treatment, at about 120 degree C, it reaches the maximum value, and then it is rapidly lowered with the further increase of temperature of thermal treatment. Based on the existence of a lot of voids and lattice defects inside the drawn HDPE, the above experimental results were explained

A potential role for endogenous proteins as sacrificial sunscreens and antioxidants in human tissues

Directory of Open Access Journals (Sweden)

Sarah A. Hibbert

2015-08-01

Full Text Available Excessive ultraviolet radiation (UVR exposure of the skin is associated with adverse clinical outcomes. Although both exogenous sunscreens and endogenous tissue components (including melanins and tryptophan-derived compounds reduce UVR penetration, the role of endogenous proteins in absorbing environmental UV wavelengths is poorly defined. Having previously demonstrated that proteins which are rich in UVR-absorbing amino acid residues are readily degraded by broadband UVB-radiation (containing UVA, UVB and UVC wavelengths here we hypothesised that UV chromophore (Cys, Trp and Tyr content can predict the susceptibility of structural proteins in skin and the eye to damage by physiologically relevant doses (up to 15.4 J/cm2 of solar UVR (95% UVA, 5% UVB. We show that: i purified suspensions of UV-chromophore-rich fibronectin dimers, fibrillin microfibrils and β- and γ-lens crystallins undergo solar simulated radiation (SSR-induced aggregation and/or decomposition and ii exposure to identical doses of SSR has minimal effect on the size or ultrastructure of UV chromophore-poor tropoelastin, collagen I, collagen VI microfibrils and α-crystallin. If UV chromophore content is a factor in determining protein stability in vivo, we would expect that the tissue distribution of Cys, Trp and Tyr-rich proteins would correlate with regional UVR exposure. From bioinformatic analysis of 244 key structural proteins we identified several biochemically distinct, yet UV chromophore-rich, protein families. The majority of these putative UV-absorbing proteins (including the late cornified envelope proteins, keratin associated proteins, elastic fibre-associated components and β- and γ-crystallins are localised and/or particularly abundant in tissues that are exposed to the highest doses of environmental UVR, specifically the stratum corneum, hair, papillary dermis and lens. We therefore propose that UV chromophore-rich proteins are localised in regions of high UVR

A potential role for endogenous proteins as sacrificial sunscreens and antioxidants in human tissues.

Science.gov (United States)

Hibbert, Sarah A; Watson, Rachel E B; Gibbs, Neil K; Costello, Patrick; Baldock, Clair; Weiss, Anthony S; Griffiths, Christopher E M; Sherratt, Michael J

2015-08-01

Excessive ultraviolet radiation (UVR) exposure of the skin is associated with adverse clinical outcomes. Although both exogenous sunscreens and endogenous tissue components (including melanins and tryptophan-derived compounds) reduce UVR penetration, the role of endogenous proteins in absorbing environmental UV wavelengths is poorly defined. Having previously demonstrated that proteins which are rich in UVR-absorbing amino acid residues are readily degraded by broadband UVB-radiation (containing UVA, UVB and UVC wavelengths) here we hypothesised that UV chromophore (Cys, Trp and Tyr) content can predict the susceptibility of structural proteins in skin and the eye to damage by physiologically relevant doses (up to 15.4 J/cm(2)) of solar UVR (95% UVA, 5% UVB). We show that: i) purified suspensions of UV-chromophore-rich fibronectin dimers, fibrillin microfibrils and β- and γ-lens crystallins undergo solar simulated radiation (SSR)-induced aggregation and/or decomposition and ii) exposure to identical doses of SSR has minimal effect on the size or ultrastructure of UV chromophore-poor tropoelastin, collagen I, collagen VI microfibrils and α-crystallin. If UV chromophore content is a factor in determining protein stability in vivo, we would expect that the tissue distribution of Cys, Trp and Tyr-rich proteins would correlate with regional UVR exposure. From bioinformatic analysis of 244 key structural proteins we identified several biochemically distinct, yet UV chromophore-rich, protein families. The majority of these putative UV-absorbing proteins (including the late cornified envelope proteins, keratin associated proteins, elastic fibre-associated components and β- and γ-crystallins) are localised and/or particularly abundant in tissues that are exposed to the highest doses of environmental UVR, specifically the stratum corneum, hair, papillary dermis and lens. We therefore propose that UV chromophore-rich proteins are localised in regions of high UVR exposure

Evidence on Inclusion

DEFF Research Database (Denmark)

Dyssegaard, Camilla Brørup; Larsen, Michael Søgaard

The purpose of this publication is to examine existing research on inclusion to identify strategies of inclusion that have generated positive effects. To do so it is necessary to understand the effect of the applied strategies. One approach, which is being discussed, is to use evidence to determine...... which methods have proven more effective than others. The desire to gain insight into research on inclusion forms the basis of the current systematic review. The task was to determine which strategies primary research has found to be most effective for inclusion purposes. We have solved this task...... by addressing the existing research with the following question: What is the effect of including children with special needs in mainstream teaching in basic school, and which of the applied educational methods have proved to have a positive effect?...

Influence of the inclusion of cooked cereals and pea starch in diets based on soy or pea protein concentrate on nutrient digestibility and performance of young pigs.

Science.gov (United States)

Parera, N; Lázaro, R P; Serrano, M P; Valencia, D G; Mateos, G G

2010-02-01

An experiment was conducted to compare different dietary vegetable sources of starch and protein on the coefficient of apparent total tract digestibility (CATTD) of energy and nutrients and performance of piglets from 29 to 60 d of age. The experiment was completely randomized with 6 treatments arranged factorially with 3 sources of starch (cooked-flaked corn, cooked-flaked rice, and pea starch) and 2 sources of protein [soy protein concentrate (SPC) and pea protein concentrate (PPC)]. The pea starch and the PPC used were obtained by dehulling and grinding pea seeds to a mean particle size of 30 microm. Each treatment was replicated 6 times (6 pigs per pen). For the entire experiment, piglets fed cooked rice had greater ADG than piglets fed pea starch with piglets fed cooked corn being intermediate (471, 403, and 430 g/d, respectively; P Protein source did not have any effect on piglet performance. The CATTD of DM, OM, and GE were greater (P pea starch being intermediate. Crude protein digestibility was not affected by source of starch but was greater for the diets based on SPC than for diets based on PPC (0.836 vs. 0.821; P Protein source did not affect the digestibility of any of the other dietary components. It is concluded that cooked rice is an energy source of choice in diets for young pigs. The inclusion of PPC in the diet reduced protein digestibility but had no effects on energy digestibility or piglet performance. Therefore, the finely ground starch and protein fractions of peas can be used in substitution of cooked corn or SPC, respectively, in diets for young pigs.

Crystalline beams: Theory, experiments, and proposals

International Nuclear Information System (INIS)

Ruggiero, A.G.

1995-01-01

Crystalline Beams are an ordered state of an ensemble of ions, circulating in a storage ring, with very small velocity fluctuations. They can be obtained from ordinary warm ion beams with the application of intense cooling techniques (stochastic, electron, laser). A phase transition occurs when sufficiently small velocity spreads are reached, freezing the particle-to-particle spacing in strings, Zigzags, and helices ... The properties and the feasibility of Crystalline Beams depend on the choice of the lattice of the Storage Ring. There are three issues closely related to the design of the Storage Ring; namely: the determination of Equilibrium Configurations, Confinement Conditions, and Stability Conditions. Of particular concern is the effect of the trajectory curvature and of the beam momentum spread, since they set the requirements on the amount of momentum cooling, on the focussing, and on the distribution of bending in the lattice of the storage ring. The practical demonstration of Crystalline Beams may create the basis for an advanced technology for particle accelerators, where the limitations due to Coulomb intrabeam scattering and space-charge forces would finally be brought under control, so that beams of ions, more dense than normal, can be achieved for a variety of new applications

Bacterial adhesion on amorphous and crystalline metal oxide coatings

International Nuclear Information System (INIS)

Almaguer-Flores, Argelia; Silva-Bermudez, Phaedra; Galicia, Rey; Rodil, Sandra E.

2015-01-01

Several studies have demonstrated the influence of surface properties (surface energy, composition and topography) of biocompatible materials on the adhesion of cells/bacteria on solid substrates; however, few have provided information about the effect of the atomic arrangement or crystallinity. Using magnetron sputtering deposition, we produced amorphous and crystalline TiO 2 and ZrO 2 coatings with controlled micro and nanoscale morphology. The effect of the structure on the physical–chemical surface properties was carefully analyzed. Then, we studied how these parameters affect the adhesion of Escherichia coli and Staphylococcus aureus. Our findings demonstrated that the nano-topography and the surface energy were significantly influenced by the coating structure. Bacterial adhesion at micro-rough (2.6 μm) surfaces was independent of the surface composition and structure, contrary to the observation in sub-micron (0.5 μm) rough surfaces, where the crystalline oxides (TiO 2 > ZrO 2 ) surfaces exhibited higher numbers of attached bacteria. Particularly, crystalline TiO 2 , which presented a predominant acidic nature, was more attractive for the adhesion of the negatively charged bacteria. The information provided by this study, where surface modifications are introduced by means of the deposition of amorphous or crystalline oxide coatings, offers a route for the rational design of implant surfaces to control or inhibit bacterial adhesion. - Highlights: • Amorphous (a) and crystalline (c) TiO 2 and ZrO 2 coatings were deposited. • The atomic ordering influences the coatings surface charge and nano-topography. • The atomic ordering modifies the bacterial adhesion for the same surface chemistry. • S. aureus adhesion was lower on a-TiO 2 and a-ZrO 2 than on their c-oxide counterpart. • E. coli adhesion on a-TiO 2 was lower than on the c-TiO 2

Bacterial adhesion on amorphous and crystalline metal oxide coatings

Energy Technology Data Exchange (ETDEWEB)

Almaguer-Flores, Argelia [Facultad de Odontología, División de Estudios de Posgrado e Investigación, Universidad Nacional Autónoma de México, Circuito exterior s/n, Ciudad Universitaria, 04510 México D.F. (Mexico); Silva-Bermudez, Phaedra, E-mail: suriel21@yahoo.com [Unidad de Ingeniería de Tejidos, Terapia Celular y Medicina Regenerativa, Instituto Nacional de Rehabilitación, Calzada México-Xochimilco No. 289, Col. Arenal de Guadalupe, 14389 México D.F. (Mexico); Instituto de Investigaciones en Materiales, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Ciudad Universitaria, 04510 México D.F. (Mexico); Galicia, Rey; Rodil, Sandra E. [Instituto de Investigaciones en Materiales, Universidad Nacional Autónoma de México, Circuito Exterior s/n, Ciudad Universitaria, 04510 México D.F. (Mexico)

2015-12-01

Several studies have demonstrated the influence of surface properties (surface energy, composition and topography) of biocompatible materials on the adhesion of cells/bacteria on solid substrates; however, few have provided information about the effect of the atomic arrangement or crystallinity. Using magnetron sputtering deposition, we produced amorphous and crystalline TiO{sub 2} and ZrO{sub 2} coatings with controlled micro and nanoscale morphology. The effect of the structure on the physical–chemical surface properties was carefully analyzed. Then, we studied how these parameters affect the adhesion of Escherichia coli and Staphylococcus aureus. Our findings demonstrated that the nano-topography and the surface energy were significantly influenced by the coating structure. Bacterial adhesion at micro-rough (2.6 μm) surfaces was independent of the surface composition and structure, contrary to the observation in sub-micron (0.5 μm) rough surfaces, where the crystalline oxides (TiO{sub 2} > ZrO{sub 2}) surfaces exhibited higher numbers of attached bacteria. Particularly, crystalline TiO{sub 2}, which presented a predominant acidic nature, was more attractive for the adhesion of the negatively charged bacteria. The information provided by this study, where surface modifications are introduced by means of the deposition of amorphous or crystalline oxide coatings, offers a route for the rational design of implant surfaces to control or inhibit bacterial adhesion. - Highlights: • Amorphous (a) and crystalline (c) TiO{sub 2} and ZrO{sub 2} coatings were deposited. • The atomic ordering influences the coatings surface charge and nano-topography. • The atomic ordering modifies the bacterial adhesion for the same surface chemistry. • S. aureus adhesion was lower on a-TiO{sub 2} and a-ZrO{sub 2} than on their c-oxide counterpart. • E. coli adhesion on a-TiO{sub 2} was lower than on the c-TiO{sub 2}.

Determination of cellulose I crystallinity by FT-Raman spectroscopy

Science.gov (United States)

Umesh P. Agarwal; Richard S. Reiner; Sally A. Ralph

2009-01-01

Two new methods based on FT-Raman spectroscopy, one simple, based on band intensity ratio, and the other, using a partial least-squares (PLS) regression model, are proposed to determine cellulose I crystallinity. In the simple method, crystallinity in semicrystalline cellulose I samples was determined based on univariate regression that was first developed using the...

Electrochemical synthesis of self-organized TiO2 crystalline nanotubes without annealing

Science.gov (United States)

Giorgi, Leonardo; Dikonimos, Theodoros; Giorgi, Rossella; Buonocore, Francesco; Faggio, Giuliana; Messina, Giacomo; Lisi, Nicola

2018-03-01

This work demonstrates that upon anodic polarization in an aqueous fluoride-containing electrolyte, TiO2 nanotube array films can be formed with a well-defined crystalline phase, rather than an amorphous one. The crystalline phase was obtained avoiding any high temperature annealing. We studied the formation of nanotubes in an HF/H2O medium and the development of crystalline grains on the nanotube wall, and we found a facile way to achieve crystalline TiO2 nanotube arrays through a one-step anodization. The crystallinity of the film was influenced by the synthesis parameters, and the optimization of the electrolyte composition and anodization conditions (applied voltage and time) were carried out. For comparison purposes, crystalline anatase TiO2 nanotubes were also prepared by thermal treatment of amorphous nanotubes grown in an organic bath (ethylene glycol/NH4F/H2O). The morphology and the crystallinity of the nanotubes were studied by field emission gun-scanning electron microscopy (FEG-SEM) and Raman spectroscopy, whereas the electrochemical and semiconducting properties were analyzed by means of linear sweep voltammetry, impedance spectroscopy, and Mott-Schottky plots. X-ray photoelectron spectroscopy (XPS) and ultraviolet photoelectron spectroscopy (UPS) allowed us to determine the surface composition and the electronic structure of the samples and to correlate them with the electrochemical data. The optimal conditions to achieve a crystalline phase with high donor concentration are defined.

Solid State Characterization of Commercial Crystalline and Amorphous Atorvastatin Calcium Samples

OpenAIRE

Shete, Ganesh; Puri, Vibha; Kumar, Lokesh; Bansal, Arvind K.

2010-01-01

Atorvastatin calcium (ATC), an anti-lipid BCS class II drug, is marketed in crystalline and amorphous solid forms. The objective of this study was to perform solid state characterization of commercial crystalline and amorphous ATC drug samples available in the Indian market. Six samples each of crystalline and amorphous ATC were characterized using X-ray powder diffractometry (XRPD), differential scanning calorimetry (DSC), thermogravimetric analysis, Karl Fisher titrimetry, microscopy (hot s...

The effect of modified ijuk fibers to crystallinity of polypropylene composite

Science.gov (United States)

Prabowo, I.; Nur Pratama, J.; Chalid, M.

2017-07-01

Nowadays, plastics becomes concern associated with its degradation and environmental issues. It has led studies to develop an environmental-friendly material. To minimize the impact of those problems, recently the usage of natural fibers as a filler are introduced because of biodegradability and availability. The promising natural fiber is “ijuk” fiber from Arenga pinnata plant as a filler and polypropylene (PP) polymer as a matrix. Unfortunately, the natural fibers and polymers have the different properties on which polymers are polar while natural fibers are non-polar so that reducing the compatibility and resulting the poor crystallinity. To enhance the compatibility and crystallinity, ijuk fibers were prepared by multistage treatments including alkalinization with 5 and 10% sodium hydroxide (NaOH), oxidation with 3 and 6% sodium hypochlorite (NaClO) and hydrolysis with 20% sulphuric acid (H2SO4) in sequences. The purposes of multistage treatments are to remove the components such as lignin, wax, hemicellulose, to cause an oxidative fragmentation of remaining lignin and to annihilate the amorphous parts respectively. Fourier-Transform Infrared (FTIR) confirms the compatibility meanwhile Differential Scanning Calorimetry (DSC) reveals the crystallinity and Scanning Electron Microscope (SEM) displays surface morphology of polypropylene. The experiments were revealing that the effects of “ijuk” fibers by the multistage treatments of 5 and 10% NaOH resulting the crystallinity of polypropylene around 31.2 and 27.64% respectively compared to the crystallinity before adding the “ijuk” fibers for 16.8%. It indicates that the entire treatments increasing the compatibility and crystallinity of polypropylene. In addition, the use of 5% NaOH offers the better crystallinity than non-treated polypropylene. The experiments conclude that by adding alkalinized “ijuk” fibers of multistage treatments can increase the compatibility and crystallinity of polypropylene.

Proteomic characterization of an isolated fraction of synthetic proteasome inhibitor (PSI-induced inclusions in PC12 cells might offer clues to aggresomes as a cellular defensive response against proteasome inhibition by PSI

Directory of Open Access Journals (Sweden)

Li Xing'an

2010-08-01

Full Text Available Abstract Background Cooperation of constituents of the ubiquitin proteasome system (UPS with chaperone proteins in degrading proteins mediate a wide range of cellular processes, such as synaptic function and neurotransmission, gene transcription, protein trafficking, mitochondrial function and metabolism, antioxidant defence mechanisms, and apoptotic signal transduction. It is supposed that constituents of the UPS and chaperone proteins are recruited into aggresomes where aberrant and potentially cytotoxic proteins may be sequestered in an inactive form. Results To determinate the proteomic pattern of synthetic proteasome inhibitor (PSI-induced inclusions in PC12 cells after proteasome inhibition by PSI, we analyzed a fraction of PSI-induced inclusions. A proteomic feature of the isolated fraction was characterized by identification of fifty six proteins including twenty previously reported protein components of Lewy bodies, twenty eight newly identified proteins and eight unknown proteins. These proteins, most of which were recognized as a profile of proteins within cellular processes mediated by the UPS, a profile of constituents of the UPS and a profile of chaperone proteins, are classed into at least nine accepted categories. In addition, prolyl-4-hydroxylase beta polypeptide, an endoplasmic reticulum member of the protein disulfide isomerase family, was validated in the developmental process of PSI-induced inclusions in the cells. Conclusions It is speculated that proteomic characterization of an isolated fraction of PSI-induced inclusions in PC12 cells might offer clues to appearance of aggresomes serving as a cellular defensive response against proteasome inhibition.

Laterally inherently thin amorphous-crystalline silicon heterojunction photovoltaic cell

Energy Technology Data Exchange (ETDEWEB)

Chowdhury, Zahidur R., E-mail: zr.chowdhury@utoronto.ca; Kherani, Nazir P., E-mail: kherani@ecf.utoronto.ca [Department of Electrical and Computer Engineering, University of Toronto, 10 King' s College Road, Toronto, Ontario M5S 3G4 (Canada)

2014-12-29

This article reports on an amorphous-crystalline silicon heterojunction photovoltaic cell concept wherein the heterojunction regions are laterally narrow and distributed amidst a backdrop of well-passivated crystalline silicon surface. The localized amorphous-crystalline silicon heterojunctions consisting of the laterally thin emitter and back-surface field regions are precisely aligned under the metal grid-lines and bus-bars while the remaining crystalline silicon surface is passivated using the recently proposed facile grown native oxide–plasma enhanced chemical vapour deposited silicon nitride passivation scheme. The proposed cell concept mitigates parasitic optical absorption losses by relegating amorphous silicon to beneath the shadowed metallized regions and by using optically transparent passivation layer. A photovoltaic conversion efficiency of 13.6% is obtained for an untextured proof-of-concept cell illuminated under AM 1.5 global spectrum; the specific cell performance parameters are V{sub OC} of 666 mV, J{sub SC} of 29.5 mA-cm{sup −2}, and fill-factor of 69.3%. Reduced parasitic absorption, predominantly in the shorter wavelength range, is confirmed with external quantum efficiency measurement.

Confinement and stability of crystalline beams in storage rings

International Nuclear Information System (INIS)

Haffmans, A.F.

1995-01-01

We present a fully analytical approach to the study of the confinement and stability of open-quote open-quote Crystalline Beams close-quote close-quote in storage rings, in terms of such fundamental accelerator concepts as tune shift and stopband. We consider a open-quote open-quote Crystalline Beam close-quote close-quote consisting of substrings, arranged symmetrically around the reference trajectory, and we examine the motion of a slightly perturbed test particle on one of them. Our approach quite naturally leads to the conclusion, that (a) storage rings need to be operated below the transition energy, and (b) the open-quote open-quote Crystalline Beam close-quote close-quote has the same periodicity as the storage ring. Each open-quote open-quote Crystalline Beam close-quote close-quote has an upper and lower limit of the spacing between the ions. The upper limit is determined by condition (b), and the lower limit is set by the stability of the test particle motion around the equilibrium. copyright 1995 American Institute of Physics

Footstep towards Inclusive Education

Science.gov (United States)

Abbas, Faiza; Zafar, Aneeka; Naz, Tayyaba

2016-01-01

Inclusive education is a rising trend in the world. The first step towards inclusive education is providing the awareness to the general education teachers. This study focused to investigate the general education teachers of primary and secondary level awareness about the special education and inclusive education. This study is descriptive method…

More Policies, Greater Inclusion? Exploring the Contradictions of New Labour Inclusive Education Policy

Science.gov (United States)

Roulstone, Alan; Prideaux, Simon

2008-01-01

The era of New Labour government has witnessed unprecedented growth in inclusive education policies. There is, however, limited evidence that policies have increased disabled children's inclusion. This article explores reasons for this contradiction. Drawing on sociological insights, it is argued that New Labour policies on inclusive education…

Anisotropy-based crystalline oxide-on-semiconductor material

Science.gov (United States)

McKee, Rodney Allen; Walker, Frederick Joseph

2000-01-01

A semiconductor structure and device for use in a semiconductor application utilizes a substrate of semiconductor-based material, such as silicon, and a thin film of a crystalline oxide whose unit cells are capable of exhibiting anisotropic behavior overlying the substrate surface. Within the structure, the unit cells of the crystalline oxide are exposed to an in-plane stain which influences the geometric shape of the unit cells and thereby arranges a directional-dependent quality of the unit cells in a predisposed orientation relative to the substrate. This predisposition of the directional-dependent quality of the unit cells enables the device to take beneficial advantage of characteristics of the structure during operation. For example, in the instance in which the crystalline oxide of the structure is a perovskite, a spinel or an oxide of similarly-related cubic structure, the structure can, within an appropriate semiconductor device, exhibit ferroelectric, piezoelectric, pyroelectric, electro-optic, ferromagnetic, antiferromagnetic, magneto-optic or large dielectric properties that synergistically couple to the underlying semiconductor substrate.

Endogenous α-crystallin inhibits expression of caspase-3 induced by hypoxia in retinal neurons.

Science.gov (United States)

Ying, Xi; Peng, Yanli; Zhang, Jiaping; Wang, Xingli; Wu, Nan; Zeng, Yuxiao; Wang, Yi

2014-08-28

To investigate the expression of endogenous, hypoxic stress-induced α-crystallin and caspase-3 in rat retinal neurons in vitro. Retinal neurons were cultured from Long-Evans rats. The expression of endogenous α-crystallin was analyzed by immunohistochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR). Furthermore, hypoxic exposure was performed in cultured cells, and the expression of endogenous α-crystallin and caspase-3 was assayed by Western blotting. Positive α-crystallin staining was observed in cultured retinal neurons, and expression of endogenous α-crystallin mRNA peaked 3-5d after inoculation (Pendogenous, hypoxic stress-induced α-crystallin expression increased gradually, peaking 6h after hypoxia. The expression was more abundant compared to the control (Pendogenous α-crystallin in retinal neurons, especially over-expression induced by hypoxic stress, results in the down regulation of caspase-3. The data suggest that endogenous α-crystallin may act as an endogenous neuroprotective factor in retinal neurons. Copyright © 2014 Elsevier Inc. All rights reserved.

Definitions of terms relating to crystalline polymers (IUPAC Recommendations 2011)

Czech Academy of Sciences Publication Activity Database

Meille, S. V.; Allegra, G.; Geil, P. H.; He, J.; Hess, M.; Jin, J.-I.; Kratochvíl, Pavel; Mormann, W.; Stepto, R.

2011-01-01

Roč. 83, č. 10 (2011), s. 1831-1871 ISSN 0033-4545 Institutional research plan: CEZ:AV0Z40500505 Keywords : IUPAC Polymer Division * crystalline polymers * crystalline polymer conformation Subject RIV: CD - Macromolecular Chemistry Impact factor: 2.789, year: 2011

Linguistic Diversity and Social Inclusion

Science.gov (United States)

Piller, Ingrid; Takahashi, Kimie

2011-01-01

This introduction provides the framework for the special issue by describing the social inclusion agenda of neoliberal market democracies. While the social inclusion agenda has been widely adopted, social inclusion policies are often blind to the ways in which language proficiency and language ideologies mediate social inclusion in linguistically…

Inclusive pedagogy

DEFF Research Database (Denmark)

Korsgaard, Morten Timmermann; Mortensen, Stig Skov

of education. The article explores the relationship between Continental and Anglo-American educational theory, and why they seem to have developed in such distinct directions. Beginning with the Anglo-American perspective it is outlined how pedagogy and the so called educational interest became replaced...... and the politicisation of inclusive education, and a positive aim in the form of an argument for a move towards constructing a pedagogical ideal of inclusion....

Solid state characterization of commercial crystalline and amorphous atorvastatin calcium samples.

Science.gov (United States)

Shete, Ganesh; Puri, Vibha; Kumar, Lokesh; Bansal, Arvind K

2010-06-01

Atorvastatin calcium (ATC), an anti-lipid BCS class II drug, is marketed in crystalline and amorphous solid forms. The objective of this study was to perform solid state characterization of commercial crystalline and amorphous ATC drug samples available in the Indian market. Six samples each of crystalline and amorphous ATC were characterized using X-ray powder diffractometry (XRPD), differential scanning calorimetry (DSC), thermogravimetric analysis, Karl Fisher titrimetry, microscopy (hot stage microscopy, scanning electron microscopy), contact angle, and intrinsic dissolution rate (IDR). All crystalline ATC samples were found to be stable form I, however one sample possessed polymorphic impurity, evidenced in XRPD and DSC analysis. Amongst the amorphous ATC samples, XRPD demonstrated five samples to be amorphous 'form 27', while, one matched amorphous 'form 23'. Thermal behavior of amorphous ATC samples was compared to amorphous ATC generated by melt quenching in DSC. ATC was found to be an excellent glass former with T(g)/T(m) of 0.95. Residual crystallinity was detected in two of the amorphous samples by complementary use of conventional and modulated DSC techniques. The wettability and IDR of all amorphous samples was found to be higher than the crystalline samples. In conclusion, commercial ATC samples exhibited diverse solid state behavior that can impact the performance and stability of the dosage forms.

Hemin as a generic and potent protein misfolding inhibitor

Energy Technology Data Exchange (ETDEWEB)

Liu, Yanqin [School of Chemistry and Physics, The University of Adelaide, Adelaide, SA 5005 (Australia); Carver, John A. [Discipline of Pharmacology, The University of Adelaide, Adelaide, SA 5005 (Australia); Ho, Lam H.; Elias, Abigail K. [School of Chemistry and Physics, The University of Adelaide, Adelaide, SA 5005 (Australia); Musgrave, Ian F. [Research School of Chemistry, The Australian National University, Canberra, ACT 0200 (Australia); Pukala, Tara L., E-mail: tara.pukala@adelaide.edu.au [School of Chemistry and Physics, The University of Adelaide, Adelaide, SA 5005 (Australia)

2014-11-14

Highlights: • Hemin prevents Aβ42, α-synuclein and RCM-κ-casein forming amyloid fibrils. • Hemin inhibits the β-sheet structure formation of Aβ42. • Hemin reduces the cell toxicity caused by fibrillar Aβ42. • Hemin dissociates partially formed Aβ42 fibrils. • Hemin prevents amorphous aggregation by ADH, catalase and γs-crystallin. - Abstract: Protein misfolding causes serious biological malfunction, resulting in diseases including Alzheimer’s disease, Parkinson’s disease and cataract. Molecules which inhibit protein misfolding are a promising avenue to explore as therapeutics for the treatment of these diseases. In the present study, thioflavin T fluorescence and transmission electron microscopy experiments demonstrated that hemin prevents amyloid fibril formation of kappa-casein, amyloid beta peptide and α-synuclein by blocking β-sheet structure assembly which is essential in fibril aggregation. Further, inhibition of fibril formation by hemin significantly reduces the cytotoxicity caused by fibrillar amyloid beta peptide in vitro. Interestingly, hemin degrades partially formed amyloid fibrils and prevents further aggregation to mature fibrils. Light scattering assay results revealed that hemin also prevents protein amorphous aggregation of alcohol dehydrogenase, catalase and γs-crystallin. In summary, hemin is a potent agent which generically stabilises proteins against aggregation, and has potential as a key molecule for the development of therapeutics for protein misfolding diseases.

αA crystallin may protect against geographic atrophy-meta-analysis of cataract vs. cataract surgery for geographic atrophy and experimental studies.

Directory of Open Access Journals (Sweden)

Peng Zhou

Full Text Available BACKGROUND: Cataract and geographic atrophy (GA, also called advanced "dry" age-related macular degeneration are the two major causes of visual impairment in the developed world. The association between cataract surgery and the development of GA was controversial in previous studies. METHODS/PRINCIPAL FINDINGS: We performed a meta-analysis by pooling the current evidence in literature and found that cataract is associated with an increased risk of geographic atrophy with a summary odds ratio (OR of 3.75 (95% CI: 95% CI: 1.84-7.62. However, cataract surgery is not associated with the risk of geographic atrophy (polled OR=3.23, 95% CI: 0.63-16.47. Further experiments were performed to analyze how the αA-crystallin, the major component of the lens, influences the development of GA in a mouse model. We found that theαA-crystallin mRNA and protein expression increased after oxidative stress induced by NaIO(3 in immunohistochemistry of retinal section and western blot of posterior eyecups. Both functional and histopathological evidence confirmed that GA is more severe in αA-crystallin knockout mice compared to wild-type mice. CONCLUSIONS: Therefore, αA-crystallin may protect against geographic atrophy. This study provides a better understanding of the relationship between cataract, cataract surgery, and GA.

Effect of the UV modification of α-crystallin on its ability to suppress nonspecific aggregation

International Nuclear Information System (INIS)

Ellozy, A.R.; Ceger, Patricia; Wang, R.H.; Dillon, James

1996-01-01

Recent studies have shown that structural modifications of α-crystallin during lens aging decrease it's effectiveness as a molecular chaperone. Some of these post-translational modifications have been linked to UV radiation, and this study was undertaken to investigate the effect of UV irradiation on the ability of α-crystallin to suppress nonspecific aggregation. The effect of 3-hydroxykynurenine (3-HK) was also investigated as a model for its glucoside (3-HKG), a main lens chromophore that has been linked to photochemical changes in the human lens. Alpha- and γ-crystallin solutions (1 mg/mL, 1:0.125 wt/wt) were photolyzed (transmission above 295nm) for various time intervals. Thermal denaturation of γ-crystallin with or without α-crystallin was carried out at 70 o C and increases in light scattering were measured at 360 nm. We found that (1) irradiation of γ-crystallin increased its susceptibility to heat-induced scattering. The addition of α-crystallin protects it against thermal denaturation, although its ability to do so decreases the longer γ-crystallin is irradiated and (2) irradiation of α-crystallin decreases its ability to suppress nonspecific aggregation and the presence of 3-HK during irradiation decreases its further. Our results indicate that post-translational modifications of α-crystallin due to UV irradiation affect the sites and mechanisms by which it interacts with γ-crystallin. The kinetics of γ-crystallin unfolding during thermal denaturation were also analyzed. We found that a simple two state model applied for nonirradiated γ-crystallin. This model does not hold when γ-crystallin is irradiated in the prescence or absence of α-crystallin. In these cases, two step or multistep mechanisms are more likely. (Author)

Inclusion in the East: Chinese Students' Attitudes towards Inclusive Education

Science.gov (United States)

Malinen, Olli-Pekka; Savolainen, Hannu

2008-01-01

A sample of 523 Chinese university students was given a questionnaire on their attitudes towards the inclusion of children with disabilities into regular classrooms. Factor analysis, analysis of variance, t-test and correlations were used to assess the respondents' general attitude towards inclusion, the factor structure of the attitudes, the…

Homogenisation of sulphide inclusions within diamonds: A new approach to diamond inclusion geochemistry

Science.gov (United States)

McDonald, Iain; Hughes, Hannah S. R.; Butler, Ian B.; Harris, Jeffrey W.; Muir, Duncan

2017-11-01

Base metal sulphide (BMS) inclusions in diamonds provide a unique insight into the chalcophile and highly siderophile element composition of the mantle. Entombed within their diamond hosts, these provide a more robust (closed system) sample, from which to determine the trace element, Re-Os and S-isotopic compositions of the mantle than mantle xenoliths or orogenic peridotites, as they are shielded from alteration during ascent to the Earth's crust and subsequent surface weathering. However, at temperatures below 1100 °C some BMS inclusions undergo subsolidus re-equilibration from an original monosulphide solid solution (Mss) and this causes fractionation of the major and trace elements within the inclusions. Thus to study the subjects noted above, current techniques require the entire BMS inclusion to be extracted for analyses. Unfortunately, 'flaking' of inclusions during break-out is a frequent occurrence and hence the risk of accidentally under-sampling a portion of the BMS inclusion is inherent in current practices. This loss may have significant implications for Re-Os isotope analyses where incomplete sampling of a Re-rich phase, such as chalcopyrite that typically occurs at the outer margins of BMS inclusions, may induce significant bias in the Re-Os and 187Os/188Os measurements and resulting model and isochron ages. We have developed a method for the homogenisation of BMS inclusions in diamond prior to their break-out from the host stone. Diamonds are heated to 1100 °C and then quenched to chemically homogenise any sulphide inclusions for both major and trace elements. Using X-ray Computed Microtomography (μCT) we determine the shape and spatial setting of multiple inclusions within a host stone and crucially show that the volume of a BMS inclusion is the same both before and after homogenisation. We show that the homogenisation process significantly reduces the inherent variability of in situ analysis when compared with unhomogenised BMS, thereby

Autophobicity and layering behavior of thin liquid-crystalline polymer films.

NARCIS (Netherlands)

Wielen, van der M.W.J.; Cohen Stuart, M.A.; Fleer, G.J.

1998-01-01

The stability against breaking-up of thin spin-coated films of liquid-crystalline polymers depends on the film thickness and annealing temperature. This study concerns side-chain liquid-crystalline polymers, based on alternating copolymers of maleic anhydride and mesogenic alkenes. The mesogenic

Limitations of inclusive fitness.

Science.gov (United States)

Allen, Benjamin; Nowak, Martin A; Wilson, Edward O

2013-12-10

Until recently, inclusive fitness has been widely accepted as a general method to explain the evolution of social behavior. Affirming and expanding earlier criticism, we demonstrate that inclusive fitness is instead a limited concept, which exists only for a small subset of evolutionary processes. Inclusive fitness assumes that personal fitness is the sum of additive components caused by individual actions. This assumption does not hold for the majority of evolutionary processes or scenarios. To sidestep this limitation, inclusive fitness theorists have proposed a method using linear regression. On the basis of this method, it is claimed that inclusive fitness theory (i) predicts the direction of allele frequency changes, (ii) reveals the reasons for these changes, (iii) is as general as natural selection, and (iv) provides a universal design principle for evolution. In this paper we evaluate these claims, and show that all of them are unfounded. If the objective is to analyze whether mutations that modify social behavior are favored or opposed by natural selection, then no aspect of inclusive fitness theory is needed.

[Crystalline lens photodisruption using femtosecond laser: experimental study].

Science.gov (United States)

Chatoux, O; Touboul, D; Buestel, C; Balcou, P; Colin, J

2010-09-01

The aim of this study was to analyze the interactions during femtosecond (fs) laser photodisruption in ex vivo porcine crystalline lenses and to study the parameters for laser interaction optimization. An experimental femtosecond laser was used. The laser characteristics were: 1030 nm wavelength; pulse duration, 400 fs; and numerical aperture, 0.13. Specific software was created to custom and monitor any type of photoablation pattern for treatment purposes. Porcine crystalline lenses were placed in an open sky holder filled with physiological liquid (BSS) covered by a glass plate. A numerical camera was associated with metrological software in order to magnify and quantify the results. Transmission electron microscopy (TEM) was performed on some samples to identify the microscopic plasma interactions with the lens. The optimization of parameters was investigated in terms of the optical breakdown threshold, the sizing of interactions, and the best pattern for alignments. More than 150 crystalline lenses of freshly enucleated pigs were treated. The optical breakdown threshold (OBT) was defined as the minimal energy level per pulse necessary to observe a physical interaction. In our study, the OBT varied according to the following parameters: the crystalline lens itself, varying from 4.2 to 7.6 μJ (mean, 5.1 μJ), and the depth of laser focus, varying up to 1 μJ, increasing in the depth of the tissue. Analyzing the distance between impacts, we observed that the closer the impacts were the less power was needed to create a clear well-drawn defect pattern (lines), i.e., with a 4-μJ optimized OBT, when the impacts were placed every 2 μm for the x,y directions and 60 μm for the z direction. Coalescent bubbles created by plasma formation always disappeared in less than 24h. The nonthermal effect of plasma and the innocuousness on surrounding tissues were proven by the TEM results. The crystalline lens photodisruption by the femtosecond laser seems an innovative

[Expression, purification and antibody preparation of recombinat SARS-CoV X5 protein].

Science.gov (United States)

Wang, Li-Na; Kong, Jian-Qiang; Zhu, Ping; Du, Guan-Hua; Wang, Wei; Cheng, Ke-Di

2008-11-01

X5 protein is one of the putative unknown proteins of SARS-CoV. The recombinant protein has been successfully expressed in E. coli in the form of insoluble inclusion body. The inclusion body was dissolved in high concentration of urea. Affinity Chromatography was preformed to purify the denatured protein, and then the product was refolded in a series of gradient solutions of urea. The purified protein was obtained with the purity of > 95% and the yield of 93.3 mg x L(-1). Polyclonal antibody of this protein was obtained, and Western blotting assay indicated that the X5 protein has the strong property of antigen. Sixty-eight percent of the recombinant protein sequence was confirmed by LC-ESI-MS/MS analysis.

High pH solubilization and chromatography-based renaturation and purification of recombinant human granulocyte colony-stimulating factor from inclusion bodies.

Science.gov (United States)

Li, Ming; Fan, Hua; Liu, Jiahua; Wang, Minhong; Wang, Lili; Wang, Chaozhan

2012-03-01

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is a very efficient therapeutic protein drug which has been widely used in human clinics to treat cancer patients suffering from chemotherapy-induced neutropenia. In this study, rhG-CSF was solubilized from inclusion bodies by using a high-pH solution containing low concentration of urea. It was found that solubilization of the rhG-CSF inclusion bodies greatly depended on the buffer pH employed; alkalic pH significantly favored the solubilization. In addition, when small amount of urea was added to the solution at high pH, the solubilization was further enhanced. After solubilization, the rhG-CSF was renatured with simultaneous purification by using weak anion exchange, strong anion exchange, and hydrophobic interaction chromatography, separately. The results indicated that the rhG-CSF solubilized by the high-pH solution containing low concentration of urea had much higher mass recovery than the one solubilized by 8 M urea when using anyone of the three refolding methods employed in this work. In the case of weak anion exchange chromatography, the high pH solubilized rhG-CSF could get a mass recovery of 73%. The strategy of combining solubilization of inclusion bodies at high pH with refolding of protein using liquid chromatography may become a routine method for protein production from inclusion bodies.

Electromagnetic processes in strong crystalline fields

CERN Multimedia

2007-01-01

We propose a number of new investigations on aspects of radiation from high energy electron and positron beams (10-300 GeV) in single crystals and amorphous targets. The common heading is radiation emission by electrons and positrons in strong electromagnetic fields, but as the setup is quite versatile, other related phenomena in radiation emission can be studied as well. The intent is to clarify the role of a number of important aspects of radiation in strong fields as e.g. observed in crystals. We propose to measure trident 'Klein-like' production in strong crystalline fields, 'crystalline undulator' radiation, 'sandwich' target phenomena, LPM suppression of pair production as well as axial and planar effects in contributions of spin to the radiation.

Age-related changes in spectral transmittance of the human crystalline lens in situ.

Science.gov (United States)

Sakanishi, Yoshihito; Awano, Masakazu; Mizota, Atsushi; Tanaka, Minoru; Murakami, Akira; Ohnuma, Kazuhiko

2012-01-01

It was the aim of this study to measure spectral transmission of the human crystalline lens in situ. The crystalline lens was illuminated by one of four light-emitting diodes of different colors. The relative spectral transmittance of the human crystalline lens was measured with the Purkinje-Sanson mirror images over a wide range of ages. The study evaluated 36 crystalline lenses of 28 subjects aged 21-76 years. There was a significant correlation between the age and spectral transmittance for blue light. Spectral transmittance of the crystalline lens in situ could be measured with Purkinje-Sanson mirror images. Copyright © 2012 S. Karger AG, Basel.

Preparation of GST Fusion Proteins.

Science.gov (United States)

Einarson, Margret B; Pugacheva, Elena N; Orlinick, Jason R

2007-04-01

INTRODUCTIONThis protocol describes the preparation of glutathione-S-transferase (GST) fusion proteins, which have had a wide range of applications since their introduction as tools for synthesis of recombinant proteins in bacteria. GST was originally selected as a fusion moiety because of several desirable properties. First and foremost, when expressed in bacteria alone, or as a fusion, GST is not sequestered in inclusion bodies (in contrast to previous fusion protein systems). Second, GST can be affinity-purified without denaturation because it binds to immobilized glutathione, which provides the basis for simple purification. Consequently, GST fusion proteins are routinely used for antibody generation and purification, protein-protein interaction studies, and biochemical analysis.

Nonlinear laser pulse response in a crystalline lens.

Science.gov (United States)

Sharma, R P; Gupta, Pradeep Kumar; Singh, Ram Kishor; Strickland, D

2016-04-01

The propagation characteristics of a spatial Gaussian laser pulse have been studied inside a gradient-index structured crystalline lens with constant-density plasma generated by the laser-tissue interaction. The propagation of the laser pulse is affected by the nonlinearities introduced by the generated plasma inside the crystalline lens. Owing to the movement of plasma species from a higher- to a lower-temperature region, an increase in the refractive index occurs that causes the focusing of the laser pulse. In this study, extended paraxial approximation has been applied to take into account the evolution of the radial profile of the Gaussian laser pulse. To examine the propagation characteristics, variation of the beam width parameter has been observed as a function of the laser power and initial beam radius. The cavitation bubble formation, which plays an important role in the restoration of the elasticity of the crystalline lens, has been investigated.

Mixing induced reactive transport in fractured crystalline rocks

International Nuclear Information System (INIS)

Martinez-Landa, Lurdes; Carrera, Jesus; Dentz, Marco; Fernàndez-Garcia, Daniel; Nardí, Albert; Saaltink, Maarten W.

2012-01-01

In this paper the solute retention properties of crystalline fractured rocks due to mixing-induced geochemical reactions are studied. While fractured media exhibit paths of fast flow and transport and thus short residence times for conservative solutes, at the same time they promote mixing and dilution due to strong heterogeneity, which leads to sharp concentration contrasts. Enhanced mixing and dilution have a double effect that favors crystalline fractured media as a possible host medium for nuclear waste disposal. Firstly, peak radionuclide concentrations are attenuated and, secondly, mixing-induced precipitation reactions are enhanced significantly, which leads to radionuclide immobilization. An integrated framework is presented for the effective modeling of these flow, transport and reaction phenomena, and the interaction between them. In a simple case study, the enhanced dilution and precipitation potential of fractured crystalline rocks are systematically studied and quantified and contrasted it to retention and attenuation in an equivalent homogeneous formation.

Biochemical, immunological and toxicological characteristics of the crystal proteins of Bacillus thuringiensis subsp. medellin

Directory of Open Access Journals (Sweden)

Sergio Orduz

1996-04-01

Full Text Available Characterization of the insecticidal and hemolytic activity of solubilized crystal proteins of Bacillus thuringiensis (Bt subsp. medellin (Btmed was performed and compared to solubilized crystal proteins of isolates 1884 of B. thuringiensis subsp. israelensis (Bti and isolate PG-14 of B. thuringiensis subsp. morrisoni (Btm. In general, at acid pH values solubilization of the Bt crystalline parasporal inclusions (CPI was lower than at alkaline pH. The larvicidal activity demonstrated by the CPI of Btmed indicated that optimal solubilization of CPI takes place at a pH value of 11.3, in Bti at pH values from 5.03 to 11.3 and in Btm at pH values from 9.05 to 11.3. Hemolytic activity against sheep red blood cells was mainly found following extraction at pH 11.3 in all Bt strains tested. Polyacrylamide gel electrophoresis under denaturing conditions revealed that optimal solubilization of the CPI in all Bt strains takes place at the alkaline pH values from 9.05 to 11.3. An enriched preparation of Btmed crystals was obtained, solubilized and crystal proteins were separated on a size exclusion column (Sephacryl S-200. Three main protein peaks were observed on the chromatogram. The first peak had two main proteins that migrate between 90 to 100 kDa. These proteins are apparently not common to other Bt strains isolated to date. The second and third peaks obtained from the size exclusion column yielded polypeptides of 68 and 28-30 kDa, respectively. Each peak independently, showed toxicity against 1st instar Culex quinquefasciatus larvae. Interestingly, combinations of the fractions corresponding to the 68 and 30 kDa protein showed an increased toxicity. These results suggest that the 94 kDa protein is an important component of the Btmed toxins with the highest potency to kill mosquito larvae. When crystal proteins of Bti were probed with antisera raised independently against the three main protein fractions of Btmed, the only crystal protein that showed

Characterization of Small HSPs from Anemonia viridis Reveals Insights into Molecular Evolution of Alpha Crystallin Genes among Cnidarians

OpenAIRE

Nicosia, Aldo; Maggio, Teresa; Mazzola, Salvatore; Gianguzza, Fabrizio; Cuttitta, Angela; Costa, Salvatore

2014-01-01

Gene family encoding small Heat-Shock Proteins (sHSPs containing α-crystallin domain) are found both in prokaryotic and eukaryotic organisms; however, there is limited knowledge of their evolution. In this study, two small HSP genes termed AvHSP28.6 and AvHSP27, both organized in one intron and two exons, were characterised in the Mediterranean snakelocks anemone Anemonia viridis. The release of the genome sequence of Hydra magnipapillata and Nematostella vectensis enabled a comprehensive stu...

Inclusive innovation: a research project on the inclusion of social responsibility

NARCIS (Netherlands)

Nijhof, A.H.J.; Fisscher, O.A.M.; de Bakker, F.G.A.

2001-01-01

The research project 'Inclusive Innovation' aims at stimulating dialogue about inclusion of social responsibility based on universal human rights in the conduct of business. This dialogue concerns both the communication within the organisation and between the organisation and interested parties in

Reaction Front Evolution during Electrochemical Lithiation of Crystalline Silicon Nanopillars

KAUST Repository

Lee, Seok Woo

2012-12-01

The high theoretical specific capacity of Si as an anode material is attractive in lithium-ion batteries, although the issues caused by large volume changes during cycling have been a major challenge. Efforts have been devoted to understanding how diffusion-induced stresses cause fracture, but recent observations of anisotropic volume expansion in single-crystalline Si nanostructures require new theoretical considerations of expansion behavior during lithiation. Further experimental investigation is also necessary to better understand the anisotropy of the lithiation process. Here, we present a method to reveal the crystalline core of partially lithiated Si nanopillars with three different crystallographic orientations by using methanol to dissolve the Li atoms from the amorphous Li-Si alloy. The exposed crystalline cores have flat {110} surfaces at the pillar sidewalls; these surfaces represent the position of the reaction front between the crystalline core and the amorphous Li-Si alloy. It was also found that an amorphous Si structure remained on the flat surfaces of the crystalline core after dissolution of the Li, which was presumed to be caused by the accumulation of Si atoms left over from the removal of Li from the Li-Si alloy. © 2012 Wiley-VCH Verlag GmbH &Co. KGaA, Weinheim.

Reaction Front Evolution during Electrochemical Lithiation of Crystalline Silicon Nanopillars

KAUST Repository

Lee, Seok Woo; Berla, Lucas A.; McDowell, Matthew T.; Nix, William D.; Cui, Yi

2012-01-01

The high theoretical specific capacity of Si as an anode material is attractive in lithium-ion batteries, although the issues caused by large volume changes during cycling have been a major challenge. Efforts have been devoted to understanding how diffusion-induced stresses cause fracture, but recent observations of anisotropic volume expansion in single-crystalline Si nanostructures require new theoretical considerations of expansion behavior during lithiation. Further experimental investigation is also necessary to better understand the anisotropy of the lithiation process. Here, we present a method to reveal the crystalline core of partially lithiated Si nanopillars with three different crystallographic orientations by using methanol to dissolve the Li atoms from the amorphous Li-Si alloy. The exposed crystalline cores have flat {110} surfaces at the pillar sidewalls; these surfaces represent the position of the reaction front between the crystalline core and the amorphous Li-Si alloy. It was also found that an amorphous Si structure remained on the flat surfaces of the crystalline core after dissolution of the Li, which was presumed to be caused by the accumulation of Si atoms left over from the removal of Li from the Li-Si alloy. © 2012 Wiley-VCH Verlag GmbH &Co. KGaA, Weinheim.

Human Metapneumovirus Induces Formation of Inclusion Bodies for Efficient Genome Replication and Transcription.

Science.gov (United States)

Cifuentes-Muñoz, Nicolás; Branttie, Jean; Slaughter, Kerri Beth; Dutch, Rebecca Ellis

2017-12-15

Human metapneumovirus (HMPV) causes significant upper and lower respiratory disease in all age groups worldwide. The virus possesses a negative-sense single-stranded RNA genome of approximately 13.3 kb encapsidated by multiple copies of the nucleoprotein (N), giving rise to helical nucleocapsids. In addition, copies of the phosphoprotein (P) and the large RNA polymerase (L) decorate the viral nucleocapsids. After viral attachment, endocytosis, and fusion mediated by the viral glycoproteins, HMPV nucleocapsids are released into the cell cytoplasm. To visualize the subsequent steps of genome transcription and replication, a fluorescence in situ hybridization (FISH) protocol was established to detect different viral RNA subpopulations in infected cells. The FISH probes were specific for detection of HMPV positive-sense RNA (+RNA) and viral genomic RNA (vRNA). Time course analysis of human bronchial epithelial BEAS-2B cells infected with HMPV revealed the formation of inclusion bodies (IBs) from early times postinfection. HMPV IBs were shown to be cytoplasmic sites of active transcription and replication, with the translation of viral proteins being closely associated. Inclusion body formation was consistent with an actin-dependent coalescence of multiple early replicative sites. Time course quantitative reverse transcription-PCR analysis suggested that the coalescence of inclusion bodies is a strategy to efficiently replicate and transcribe the viral genome. These results provide a better understanding of the steps following HMPV entry and have important clinical implications. IMPORTANCE Human metapneumovirus (HMPV) is a recently discovered pathogen that affects human populations of all ages worldwide. Reinfections are common throughout life, but no vaccines or antiviral treatments are currently available. In this work, a spatiotemporal analysis of HMPV replication and transcription in bronchial epithelial cell-derived immortal cells was performed. HMPV was shown to

Continuous processing of recombinant proteins: Integration of inclusion body solubilization and refolding using simulated moving bed size exclusion chromatography with buffer recycling.

Science.gov (United States)

Wellhoefer, Martin; Sprinzl, Wolfgang; Hahn, Rainer; Jungbauer, Alois

2013-12-06

An integrated process which combines continuous inclusion body dissolution with NaOH and continuous matrix-assisted refolding based on closed-loop simulated moving bed size exclusion chromatography was designed and experimentally evaluated at laboratory scale. Inclusion bodies from N(pro) fusion pep6His and N(pro) fusion MCP1 from high cell density fermentation were continuously dissolved with NaOH, filtered and mixed with concentrated refolding buffer prior to refolding by size exclusion chromatography (SEC). This process enabled an isocratic operation of the simulated moving bed (SMB) system with a closed-loop set-up with refolding buffer as the desorbent buffer and buffer recycling by concentrating the raffinate using tangential flow filtration. With this continuous refolding process, we increased the refolding and cleavage yield of both model proteins by 10% compared to batch dilution refolding. Furthermore, more than 99% of the refolding buffer of the raffinate could be recycled which reduced the buffer consumption significantly. Based on the actual refolding data, we compared throughput, productivity, and buffer consumption between two batch dilution refolding processes - one using urea for IB dissolution, the other one using NaOH for IB dissolution - and our continuous refolding process. The higher complexity of the continuous refolding process was rewarded with higher throughput and productivity as well as significantly lower buffer consumption compared to the batch dilution refolding processes. Copyright © 2013 Elsevier B.V. All rights reserved.

Spatial distribution of radiation damage to crystalline proteins at 25–300 K

Energy Technology Data Exchange (ETDEWEB)

Warkentin, Matthew; Badeau, Ryan; Hopkins, Jesse B.; Thorne, Robert E., E-mail: ret6@cornell.edu [Cornell University, Ithaca, NY 14853 (United States)

2012-09-01

Dose-dependent atomic B factors are used to determine the average spatial distribution of radiation damage to crystalline thaumatin and urease. The spatial distribution of radiation damage (assayed by increases in atomic B factors) to thaumatin and urease crystals at temperatures ranging from 25 to 300 K is reported. The nature of the damage changes dramatically at approximately 180 K. Above this temperature the role of solvent diffusion is apparent in thaumatin crystals, as solvent-exposed turns and loops are especially sensitive. In urease, a flap covering the active site is the most sensitive part of the molecule and nearby loops show enhanced sensitivity. Below 180 K sensitivity is correlated with poor local packing, especially in thaumatin. At all temperatures, the component of the damage that is spatially uniform within the unit cell accounts for more than half of the total increase in the atomic B factors and correlates with changes in mosaicity. This component may arise from lattice-level, rather than local, disorder. The effects of primary structure on radiation sensitivity are small compared with those of tertiary structure, local packing, solvent accessibility and crystal contacts.

Reversibility of neurofilamentous inclusion formation following repeated sublethal intracisternal inoculums of AlCl3 in New Zealand white rabbits.

Science.gov (United States)

Strong, M J; Gaytan-Garcia, S; Jakowec, D M

1995-01-01

In this report, we describe the clinical, topographical and immunohistochemical characteristics of neurofilament (NF) inclusion formation induced by the intracisternal inoculation of young adult New Zealand white rabbits at 28-day intervals with 100 micrograms AlCl3 over the course of 267 days. The ability to recover following cessation of aluminum exposure has also been assessed. The extent of neurofilamentous inclusion formation was proportionate to the cumulative amount of AlCl3 inoculated and initially consisted of fusiform axonal distention in the ventral spinal cord at day 51 following the initial inoculum. Spinal motor neuron perikaryal inclusions and discrete axonal spheroids were observed at day 107 and supraspinal neurofilamentous pathology by day 156. Perikaryal inclusions were immunoreactive to antibodies recognizing both poorly phosphorylated (SMI 32) and more highly phosphorylated high molecular weight NF (NFH). In contrast, axonal spheroids were intensely immunoreactive at all stages with antibodies recognizing highly phosphorylated NFH and an age-dependent NFH phosphorylation state (SMI 34) with only faint SMI 32 immunoreactivity. Immunoreactivity to an antibody recognizing ubiquitin-protein conjugates did not appear until day 156, whereas inclusions were not immunoreactive to antibodies recognizing either phosphatase-dependent or -independent microtubule-associated protein tau at any stage. Upon withdrawal from further AlCl3 exposure after intervals of 51, 107 or 156 days following the initial inoculum, clinical recovery ensued in all rabbits. In all but the most severely affected rabbits, perikaryal neurofilamentous inclusions resolved. However, axonal spheroids continued to be prominent. These studies demonstrate that the repetitive intracisternal inoculation of AlCl3 in New Zealand white rabbits induces a reversible process of neurofilamentous inclusion formation that preferentially affects motor neurons, and in which recovery will occur in

Excimer fluorescence of liquid crystalline systems

Science.gov (United States)

Sakhno, Tamara V.; Khakhel, Oleg A.; Barashkov, Nikolay N.; Korotkova, Irina V.

1996-04-01

The method of synchronous scanning fluorescence spectroscopy shows a presence of dimers of pyrene in a polymeric matrix. The results suggest that excimer formation takes place with dimers in liquid crystalline systems.

Financial inclusion: Policies and practices

Directory of Open Access Journals (Sweden)

Thankom Arun

2015-12-01

Full Text Available As a key enabler for development, financial inclusion is firmly placed on the agenda of most governments as a key policy priority. Against this background, this round table provides a global and regional perspective on the policies and practices of financial inclusion. Using macro data, the collection reveals the diversity in the efforts towards achieving financial inclusion and the need for a progressive approach in financial inclusion. Further to this, the round table provides the regional perspectives on the policies and practices of financial inclusion in India, South Africa, and Australia.

Crystalline lens power and refractive error.

Science.gov (United States)

Iribarren, Rafael; Morgan, Ian G; Nangia, Vinay; Jonas, Jost B

2012-02-01

To study the relationships between the refractive power of the crystalline lens, overall refractive error of the eye, and degree of nuclear cataract. All phakic participants of the population-based Central India Eye and Medical Study with an age of 50+ years were included. Calculation of the refractive lens power was based on distance noncycloplegic refractive error, corneal refractive power, anterior chamber depth, lens thickness, and axial length according to Bennett's formula. The study included 1885 subjects. Mean refractive lens power was 25.5 ± 3.0 D (range, 13.9-36.6). After adjustment for age and sex, the standardized correlation coefficients (β) of the association with the ocular refractive error were highest for crystalline lens power (β = -0.41; P lens opacity grade (β = -0.42; P lens power (β = -0.95), lower corneal refractive power (β = -0.76), higher lens thickness (β = 0.30), deeper anterior chamber (β = 0.28), and less marked nuclear lens opacity (β = -0.05). Lens thickness was significantly lower in eyes with greater nuclear opacity. Variations in refractive error in adults aged 50+ years were mostly influenced by variations in axial length and in crystalline lens refractive power, followed by variations in corneal refractive power, and, to a minor degree, by variations in lens thickness and anterior chamber depth.

International Collaboration on Spent Fuel Disposition in Crystalline Media: FY17 Progress Report

Energy Technology Data Exchange (ETDEWEB)

Wang, Yifeng [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Hadgu, Teklu [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Kainina, Elena [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Jove-Colon, Carlos [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

2017-09-01

Active participation in international R&D is crucial for achieving the Spent Fuel Waste Science & Technology (SFWST) long-term goals of conducting “experiments to fill data needs and confirm advanced modeling approaches” and of having a “robust modeling and experimental basis for evaluation of multiple disposal system options” (by 2020). DOE’s Office of Nuclear Energy (NE) has developed a strategic plan to advance cooperation with international partners. The international collaboration on the evaluation of crystalline disposal media at Sandia National Laboratories (SNL) in FY17 focused on the collaboration through the Development of Coupled Models and their Validation against Experiments (DECOVALEX-2019) project. The DECOVALEX project is an international research and model comparison collaboration, initiated in 1992, for advancing the understanding and modeling of coupled thermo-hydro-mechanical-chemical (THMC) processes in geological systems. SNL has been participating in three tasks of the DECOVALEX project: Task A. Modeling gas injection experiments (ENGINEER), Task C. Modeling groundwater recovery experiment in tunnel (GREET), and Task F. Fluid inclusion and movement in the tight rock (FINITO).

A metastable liquid melted from a crystalline solid under decompression

Science.gov (United States)

Lin, Chuanlong; Smith, Jesse S.; Sinogeikin, Stanislav V.; Kono, Yoshio; Park, Changyong; Kenney-Benson, Curtis; Shen, Guoyin

2017-01-01

A metastable liquid may exist under supercooling, sustaining the liquid below the melting point such as supercooled water and silicon. It may also exist as a transient state in solid-solid transitions, as demonstrated in recent studies of colloidal particles and glass-forming metallic systems. One important question is whether a crystalline solid may directly melt into a sustainable metastable liquid. By thermal heating, a crystalline solid will always melt into a liquid above the melting point. Here we report that a high-pressure crystalline phase of bismuth can melt into a metastable liquid below the melting line through a decompression process. The decompression-induced metastable liquid can be maintained for hours in static conditions, and transform to crystalline phases when external perturbations, such as heating and cooling, are applied. It occurs in the pressure-temperature region similar to where the supercooled liquid Bi is observed. Akin to supercooled liquid, the pressure-induced metastable liquid may be more ubiquitous than we thought.

Crystalline anhydrous {alpha},{alpha}-trehalose (polymorph {beta}) and crystalline dihydrate {alpha},{alpha}-trehalose: A calorimetric study

Energy Technology Data Exchange (ETDEWEB)

Pinto, Susana S. [Centro de Quimica Estrutural, Complexo Interdisciplinar, Instituto Superior Tecnico, 1049-001 Lisbon (Portugal)]. E-mail: susanapinto@ist.utl.pt; Diogo, Herminio P. [Centro de Quimica Estrutural, Complexo Interdisciplinar, Instituto Superior Tecnico, 1049-001 Lisbon (Portugal)]. E-mail: hdiogo@ist.utl.pt; Moura-Ramos, Joaquim J. [Centro de Quimica-Fisica Molecular, Complexo Interdisciplinar, Instituto Superior Tecnico, 1049-001 Lisbon (Portugal)]. E-mail: mouraramos@ist.utl.pt

2006-09-15

The mean values of the standard massic energy of combustion of crystalline anhydrous {alpha},{alpha}-trehalose (C{sub 12}H{sub 22}O{sub 11}, polymorph {beta}) and crystalline dihydrate {alpha},{alpha}-trehalose (C{sub 12}H{sub 26}O{sub 13}) measured by static-bomb combustion calorimetry in oxygen, at the temperature T=298.15K, are {delta}{sub c}u{sup o}=-(16434.05+/-4.50)J.g{sup -1} and {delta}{sub c}u{sup o}=-(14816.05+/-3.52)J.g{sup -1}, respectively. The standard (p{sup o}=0.1MPa) molar enthalpy of formation of these compounds were derived from the corresponding standard molar enthalpies of combustion, respectively, {delta}{sub f}H{sub m}{sup o} (C{sub 12}H{sub 22}O{sub 11},cr)=-(2240.9+/-3.9)kJ.mol{sup -1}, and {delta}{sub f}H{sub m}{sup o} (C{sub 12}H{sub 26}O{sub 13},cr)=-(2832.6+/-3.6)kJ.mol{sup -1}. The values of the standard enthalpies of formation obtained in this work, together with data on enthalpies of solution at infinite dilution ({delta}{sub sol}H{sup {approx}}) for crystalline dihydrate and amorphous anhydrous trehalose, allow a better insight on the thermodynamic description of the trehalose system which can provide, together with the future research on the subject, a contribution for understanding the metabolism in several organisms, as well as the phase transition between the different polymorphs.

INCLUSIVE CULTURE IN PRE-SCHOOL INSTITUTIONS

Directory of Open Access Journals (Sweden)

Irena NOVACHEVSKA

2011-04-01

Full Text Available Inclusive education is a rational concept that refers to the overall and long-term transformation of institutional systems in society, especially in education. Along with the transformation, a number of important and unresolved issues still appear in both theory and practice, as the duty of pre-school institutions and schools is to educate every student in the mainstream education system. One of the most important aspects of inclusion is the inclusive culture. Regardless of the good inclusive policy and practice, one cannot talk about successful inclusion without a properly developed inclusive institutional culture.This paper is a contribution to the research considering the development of inclusive culture in three preschool institutions. It is based on the thinking and attitudes of the pre­school staff toward the necessity of developing and nurturing an inclusive culture. Successful inclusion of pupils with special needs in the mainstream school system cannot be conceived without an inclusive culture.

Effects of dietary Tenebrio molitor meal inclusion in free-range chickens.

Science.gov (United States)

Biasato, I; De Marco, M; Rotolo, L; Renna, M; Lussiana, C; Dabbou, S; Capucchio, M T; Biasibetti, E; Costa, P; Gai, F; Pozzo, L; Dezzutto, D; Bergagna, S; Martínez, S; Tarantola, M; Gasco, L; Schiavone, A

2016-12-01

Insects are currently being considered as a novel protein source for animal feeds, because they contain a large amount of protein. The larvae of Tenebrio molitor (TM) have been shown to be an acceptable protein source for broiler chickens in terms of growth performance, but till now, no data on histological or intestinal morphometric features have been reported. This study has had the aim of evaluating the effects of dietary TM inclusion on the performance, welfare, intestinal morphology and histological features of free-range chickens. A total of 140 medium-growing hybrid female chickens were free-range reared and randomly allotted to two dietary treatments: (i) a control group and (ii) a TM group, in which TM meal was included at 75 g/kg. Each group consisted of five pens as replicates, with 14 chicks per pen. Growth performance, haematological and serum parameters and welfare indicators were evaluated, and the animals were slaughtered at the age of 97 days. Two birds per pen (10 birds/treatment) were submitted to histological (liver, spleen, thymus, bursa of Fabricius, kidney, heart, glandular stomach and gut) and morphometric (duodenum, jejunum and ileum) investigations. The inclusion of TM did not affect the growth performance, haematological or serum parameters. The morphometric and histological features were not significantly affected either, thus suggesting no influence on nutrient metabolization, performance or animal health. Glandular stomach alterations (chronic flogosis with epithelial squamous metaplasia) were considered paraphysiological in relation to free-range farming. The observed chronic intestinal flogosis, with concomitant activation of the lymphoid tissue, was probably due to previous parasitic infections, which are very frequently detected in free-range chickens. In conclusion, the findings of this study show that yellow mealworm inclusion does not affect the welfare, productive performances or morphological features of free-range chickens

Production of recombinant proteins in Escherichia coli tagged with the fusion protein CusF3H.

Science.gov (United States)

Vargas-Cortez, Teresa; Morones-Ramirez, Jose Ruben; Balderas-Renteria, Isaias; Zarate, Xristo

2017-04-01

Recombinant protein expression in the bacterium Escherichia coli still is the number one choice for large-scale protein production. Nevertheless, many complications can arise using this microorganism, such as low yields, the formation of inclusion bodies, and the requirement for difficult purification steps. Most of these problems can be solved with the use of fusion proteins. Here, the use of the metal-binding protein CusF3H+ is described as a new fusion protein for recombinant protein expression and purification in E. coli. We have previously shown that CusF produces large amounts of soluble protein, with low levels of formation of inclusion bodies, and that proteins can be purified using IMAC resins charged with Cu(II) ions. CusF3H+ is an enhanced variant of CusF, formed by the addition of three histidine residues at the N-terminus. These residues then can bind Ni(II) ions allowing improved purity after affinity chromatography. Expression and purification of Green Fluorescent Protein tagged with CusF3H+ showed that the mutation did not alter the capacity of the fusion protein to increase protein expression, and purity improved considerably after affinity chromatography with immobilized nickel ions; high yields are obtained after tag-removal since CusF3H+ is a small protein of just 10 kDa. Furthermore, the results of experiments involving expression of tagged proteins having medium to large molecular weights indicate that the presence of the CusF3H+ tag improves protein solubility, as compared to a His-tag. We therefore endorse CusF3H+ as a useful alternative fusion protein/affinity tag for production of recombinant proteins in E. coli. Copyright © 2017 Elsevier Inc. All rights reserved.

Cryo-electron microscopy of membrane proteins.

Science.gov (United States)

Goldie, Kenneth N; Abeyrathne, Priyanka; Kebbel, Fabian; Chami, Mohamed; Ringler, Philippe; Stahlberg, Henning

2014-01-01

Electron crystallography is used to study membrane proteins in the form of planar, two-dimensional (2D) crystals, or other crystalline arrays such as tubular crystals. This method has been used to determine the atomic resolution structures of bacteriorhodopsin, tubulin, aquaporins, and several other membrane proteins. In addition, a large number of membrane protein structures were studied at a slightly lower resolution, whereby at least secondary structure motifs could be identified.In order to conserve the structural details of delicate crystalline arrays, cryo-electron microscopy (cryo-EM) allows imaging and/or electron diffraction of membrane proteins in their close-to-native state within a lipid bilayer membrane.To achieve ultimate high-resolution structural information of 2D crystals, meticulous sample preparation for electron crystallography is of outmost importance. Beam-induced specimen drift and lack of specimen flatness can severely affect the attainable resolution of images for tilted samples. Sample preparations that sandwich the 2D crystals between symmetrical carbon films reduce the beam-induced specimen drift, and the flatness of the preparations can be optimized by the choice of the grid material and the preparation protocol.Data collection in the cryo-electron microscope using either the imaging or the electron diffraction mode has to be performed applying low-dose procedures. Spot-scanning further reduces the effects of beam-induced drift. Data collection using automated acquisition schemes, along with improved and user-friendlier data processing software, is increasingly being used and is likely to bring the technique to a wider user base.

Synthesis of single-crystalline Al layers in sapphire

International Nuclear Information System (INIS)

Schlosser, W.; Lindner, J.K.N.; Zeitler, M.; Stritzker, B.

1999-01-01

Single-crystalline, buried aluminium layers were synthesized by 180 keV high-dose Al + ion implantation into sapphire at 500 deg. C. The approximately 70 nm thick Al layers exhibit in XTEM investigations locally abrupt interfaces to the single-crystalline Al 2 O 3 top layer and bulk, while thickness and depth position are subjected to variations. The layers grow by a ripening process of oriented Al precipitates, which at low doses exist at two different orientations. With increasing dose, precipitates with one out of the two orientations are observed to exist preferentially, finally leading to the formation of a single-crystalline layer. Al outdiffusion to the surface and the formation of spherical Al clusters at the surface are found to be competing processes to buried layer formation. The formation of Al layers is described by Rutherford Backscattering Spectroscopy (RBS), Cross-section transmission electron microscopy (XTEM) and Scanning electron microscopy (SEM) studies as a function of dose, temperature and substrate orientation

Disorder-induced localization in crystalline phase-change materials.

Science.gov (United States)

Siegrist, T; Jost, P; Volker, H; Woda, M; Merkelbach, P; Schlockermann, C; Wuttig, M

2011-03-01

Localization of charge carriers in crystalline solids has been the subject of numerous investigations over more than half a century. Materials that show a metal-insulator transition without a structural change are therefore of interest. Mechanisms leading to metal-insulator transition include electron correlation (Mott transition) or disorder (Anderson localization), but a clear distinction is difficult. Here we report on a metal-insulator transition on increasing annealing temperature for a group of crystalline phase-change materials, where the metal-insulator transition is due to strong disorder usually associated only with amorphous solids. With pronounced disorder but weak electron correlation, these phase-change materials form an unparalleled quantum state of matter. Their universal electronic behaviour seems to be at the origin of the remarkable reproducibility of the resistance switching that is crucial to their applications in non-volatile-memory devices. Controlling the degree of disorder in crystalline phase-change materials might enable multilevel resistance states in upcoming storage devices.

The physics of large deformation of crystalline solids

CERN Document Server

Bell, James F

1968-01-01

Historically, a major problem for the study of the large deformation of crystalline solids has been the apparent lack of unity in experimentally determined stress-strain functions. The writer's discovery in 1949 of the unexpectedly high velocity of incremental loading waves in pre-stressed large deformation fields emphasized to him the pressing need for the independent, systematic experimental study of the subject, to provide a firm foundation upon which physically plausible theories for the finite deformation of crystalline solids could be constructed. Such a study undertaken by the writer at that time and continued uninterruptedly to the present, led in 1956 to the development of the diffraction grating experiment which permitted, for the first time, the optically accurate determination of the strain-time detail of non-linear finite amplitude wave fronts propagating into crystalline solids whose prior history was precisely known. These experimental diffraction grating studies during the past decade have led...

Aggregation of Trp > Glu point mutants of human gamma-D crystallin provides a model for hereditary or UV-induced cataract.

Science.gov (United States)

Serebryany, Eugene; Takata, Takumi; Erickson, Erika; Schafheimer, Nathaniel; Wang, Yongting; King, Jonathan A

2016-06-01

Numerous mutations and covalent modifications of the highly abundant, long-lived crystallins of the eye lens cause their aggregation leading to progressive opacification of the lens, cataract. The nature and biochemical mechanisms of the aggregation process are poorly understood, as neither amyloid nor native-state polymers are commonly found in opaque lenses. The βγ-crystallin fold contains four highly conserved buried tryptophans, which can be oxidized to more hydrophilic products, such as kynurenine, upon UV-B irradiation. We mimicked this class of oxidative damage using Trp→Glu point mutants of human γD-crystallin. Such substitutions may represent a model of UV-induced photodamage-introduction of a charged group into the hydrophobic core generating "denaturation from within." The effects of Trp→Glu substitutions were highly position dependent. While each was destabilizing, only the two located in the bottom of the double Greek key fold-W42E and W130E-yielded robust aggregation of partially unfolded intermediates at 37°C and pH 7. The αB-crystallin chaperone suppressed aggregation of W130E, but not W42E, indicating distinct aggregation pathways from damage in the N-terminal vs C-terminal domain. The W130E aggregates had loosely fibrillar morphology, yet were nonamyloid, noncovalent, showed little surface hydrophobicity, and formed at least 20°C below the melting temperature of the native β-sheets. These features are most consistent with domain-swapped polymerization. Aggregation of partially destabilized crystallins under physiological conditions, as occurs in this class of point mutants, could provide a simple in vitro model system for drug discovery and optimization. © 2016 The Protein Society.

Liquid crystalline thermosetting polymers as protective coatings for aerospace

OpenAIRE

Guerriero, G.L.

2012-01-01

Environmental regulations are driving the development of new aerospace coating systems, mainly to eliminate chromates and reduce volatile organic compound (VOC) emissions. Among the various potential options for new coating materials, liquid crystalline polymers (LCPs) are attractive due to their unique combination of mechanical properties and chemical resistance. Their use, however, has been limited mainly due to poor adhesion properties. Thermotropic liquid crystalline thermosets displayed ...

Active protein aggregates induced by terminally attached self-assembling peptide ELK16 in Escherichia coli

Directory of Open Access Journals (Sweden)

Zhou Bihong

2011-02-01

Full Text Available Abstract Background In recent years, it has been gradually realized that bacterial inclusion bodies (IBs could be biologically active. In particular, several proteins including green fluorescent protein, β-galactosidase, β-lactamase, alkaline phosphatase, D-amino acid oxidase, polyphosphate kinase 3, maltodextrin phosphorylase, and sialic acid aldolase have been successfully produced as active IBs when fused to an appropriate partner such as the foot-and-mouth disease virus capsid protein VP1, or the human β-amyloid peptide Aβ42(F19D. As active IBs may have many attractive advantages in enzyme production and industrial applications, it is of considerable interest to explore them further. Results In this paper, we report that an ionic self-assembling peptide ELK16 (LELELKLK2 was able to effectively induce the formation of cytoplasmic inclusion bodies in Escherichia coli (E. coli when attached to the carboxyl termini of four model proteins including lipase A, amadoriase II, β-xylosidase, and green fluorescent protein. These aggregates had a general appearance similar to the usually reported cytoplasmic inclusion bodies (IBs under transmission electron microscopy or fluorescence confocal microscopy. Except for lipase A-ELK16 fusion, the three other fusion protein aggregates retained comparable specific activities with the native counterparts. Conformational analyses by Fourier transform infrared spectroscopy revealed the existence of newly formed antiparallel beta-sheet structures in these ELK16 peptide-induced inclusion bodies, which is consistent with the reported assembly of the ELK16 peptide. Conclusions This has been the first report where a terminally attached self-assembling β peptide ELK16 can promote the formation of active inclusion bodies or active protein aggregates in E. coli. It has the potential to render E. coli and other recombinant hosts more efficient as microbial cell factories for protein production. Our observation might

Growth and Brilliant Photo-Emission of Crystalline Hexagonal Column of Alq3 Microwires

Directory of Open Access Journals (Sweden)

Seokho Kim

2018-03-01

Full Text Available We report the growth and nanoscale luminescence characteristics of 8-hydroxyquinolinato aluminum (Alq3 with a crystalline hexagonal column morphology. Pristine Alq3 nanoparticles (NPs were prepared using a conventional reprecipitation method. Crystal hexagonal columns of Alq3 were grown by using a surfactant-assisted self-assembly technique as an adjunct to the aforementioned reprecipitation method. The formation and structural properties of the crystalline and non-crystalline Alq3 NPs were analyzed with scanning electron microscopy and X-ray diffraction. The nanoscale photoluminescence (PL characteristics and the luminescence color of the Alq3 single NPs and their crystal microwires (MWs were evaluated from color charge-coupled device images acquired using a high-resolution laser confocal microscope. In comparison with the Alq3 NPs, the crystalline MWs exhibited a very bright and sharp emission. This enhanced and sharp emission from the crystalline Alq3 single MWs originated from effective π-π stacking of the Alq3 molecules due to strong interactions in the crystalline structure.

The study of membrane-protein /detergent interactions by neutron crystallography

Energy Technology Data Exchange (ETDEWEB)

Timmins, P A; Penel, S [Institut Max von Laue - Paul Langevin (ILL), 38 - Grenoble (France); Pebay-Peyroula, E [IBS- UJF Grenoble (France)

1997-04-01

Proteins which are found embedded in membranes can usually only be purified and studied from the point of view of structure by dissolving them in detergents. The structure of the resulting mixed protein-detergent complexes are poorly understood. An important method for studying them is through neutron diffraction of the crystalline complexes. This allows us to understand better how the proteins behave in the natural membrane as well as allowing us to visualize and hopefully improve the crystallisation process. Studies on the pore-forming protein porin using data collected on the diffractometer DB21 are described. (author). 4 refs.

Crystallinity changes in wheat starch during the bread-making process: Starch crystallinity in the bread crust

NARCIS (Netherlands)

Primo-Martín, C.; Nieuwenhuijzen, N.H. van; Hamer, R.J.; Vliet, T. van

2007-01-01

The crystallinity of starch in crispy bread crust was quantified using several different techniques. Confocal scanning laser microscopy (CSLM) demonstrated the presence of granular starch in the crust and remnants of granules when moving towards the crumb. Differential scanning calorimetry (DSC)

Crystallinity changes in wheat starch during the bread-making process: starch crystallinity in the bread crust

NARCIS (Netherlands)

Primo-Martin, C.; Nieuwenhuijzen, van N.H.; Hamer, R.J.; Vliet, van T.

2007-01-01

The crystallinity of starch in crispy bread crust was quantified using several different techniques. Confocal scanning laser microscopy (CSLM) demonstrated the presence of granular starch in the crust and remnants of granules when moving towards the crumb. Differential scanning calorimetry (DSC)

Effect of Alchornea cordifolia leaf meal inclusion and enzyme supplementation on performance and digestibility of rabbits

Directory of Open Access Journals (Sweden)

S.O. Ayodele

2016-09-01

Full Text Available A feeding trial was conducted to study the performance, digestibility and health status of weaner rabbits fed diets including Alchornea cordifolia leaf meal (ALM: 18% crude protein [CP] and 12.9% crude fibre and supplemented with a multi-enzyme additive (cellulase, xylanase, β-glucanase, α-amylase, protease, lipase. Six experimental diets were arranged factorially: 3 levels of ALM (0, 5 and 10% substituting palm kernel cake: 16.3% CP and 39.1% neutral detergent fibre combined with 2 levels of enzyme supplementation (0 and 0.35 g/kg. One hundred and eighty healthy, 5-wk-old weaner rabbits of cross-breeds were randomly allotted to 6 dietary treatments (30 rabbits/treatment, 3 rabbits/replicate. Growth rate was not affected (P>0.05 by the main factors (exogenous enzyme and ALM inclusion and their interactions (13.5 g/d on av.. Daily feed intake and feed conversion ratio decreased (P=0.01 with the ALM inclusion by 8%, but did not affect faecal digestibility. However, enzyme supplementation improved crude protein and crude fibre digestibility (P<0.001 by 6%. In conclusion, ALM inclusion and enzyme supplementation had no adverse effect on the performance and digestibility of rabbits.

Fluid inclusion geothermometry

Science.gov (United States)

Cunningham, C.G.

1977-01-01

Fluid inclusions trapped within crystals either during growth or at a later time provide many clues to the histories of rocks and ores. Estimates of fluid-inclusion homogenization temperature and density can be obtained using a petrographic microscope with thin sections, and they can be refined using heating and freezing stages. Fluid inclusion studies, used in conjunction with paragenetic studies, can provide direct data on the time and space variations of parameters such as temperature, pressure, density, and composition of fluids in geologic environments. Changes in these parameters directly affect the fugacity, composition, and pH of fluids, thus directly influencing localization of ore metals. ?? 1977 Ferdinand Enke Verlag Stuttgart.

Syntheses, molecular and crystalline architectures, and ...

Indian Academy of Sciences (India)

Syntheses, molecular and crystalline architectures, and luminescence behaviour of terephthalate bridged heptacoordinated dinuclear lead(II) complexes containing a pentadentate N-donor Schiff base. SUBHASIS ROYa, SOMNATH CHOUBEYa, SUMITAVA KHANa, KISHALAY BHARa,. PARTHA MITRAb and BARINDRA ...

Atomically flat single-crystalline gold nanostructures for plasmonic nanocircuitry

Energy Technology Data Exchange (ETDEWEB)

Huang, J S; Geisler, P; Bruening, C; Kern, J; Prangsma, J C; Wu, X; Feichtner, Thorsten; Ziegler, J; Weinmann, P; Kamp, M; Forchel, A; Hecht, B [Wilhelm-Conrad-Roentgen-Center for Complex Material Systems, University of Wuerzburg (Germany); Biagioni, P [CNISM, Dipartimento di Fisica, Politecnico di Milano (Italy)

2011-07-01

Deep subwavelength integration of high-definition plasmonic nano-structures is of key importance for the development of future optical nanocircuitry. So far the experimental realization of proposed extended plasmonic networks remains challenging, mainly due to the multi-crystallinity of commonly used thermally evaporated gold layers. Resulting structural imperfections in individual circuit elements drastically reduce the yield of functional integrated nanocircuits. Here we demonstrate the use of very large but thin chemically grown single-crystalline gold flakes. After immobilization on any arbitrary surface, they serve as an ideal basis for focused-ion beam milling. We present high-definition ultra-smooth gold nanostructures with reproducible nanosized features over micrometer lengthscales. By comparing multi- and single-crystalline optical antennas we prove that the latter have superior optical properties which are in good agreement with numerical simulations.

Becoming Inclusive: A Code of Conduct for Inclusion and Diversity.

Science.gov (United States)

Schmidt, Bonnie J; MacWilliams, Brent R; Neal-Boylan, Leslie

There are increasing concerns about exclusionary behaviors and lack of diversity in the nursing profession. Exclusionary behaviors, which may include incivility, bullying, and workplace violence, discriminate and isolate individuals and groups who are different, whereas inclusive behaviors encourage diversity. To address inclusion and diversity in nursing, this article offers a code of conduct. This code of conduct builds on existing nursing codes of ethics and applies to nursing students and nurses in both educational and practice settings. Inclusive behaviors that are demonstrated in nurses' relationships with patients, colleagues, the profession, and society are described. This code of conduct provides a basis for measureable change, empowerment, and unification of the profession. Recommendations, implications, and a pledge to action are discussed. Copyright © 2016 Elsevier Inc. All rights reserved.

WORKSHOP: Crystalline beams

International Nuclear Information System (INIS)

Anon.

1989-01-01

Following pioneer work by specialists at the Soviet Novosibirsk Laboratory some ten years ago, interest developed in the possibility of 'freezing' ion beams in storage rings by pushing cooling (to smooth out beam behaviour) to its limits, the final goal being to lock the ions into a neat crystal pattern. After advances by groups working on laser cooled ions in traps, and with several cooling rings now in operation, a workshop on crystalline ion beams was organized recently by the GSI (Darmstadt) Laboratory and held at Wertheim in Germany

Postradiation reactions of free radicals in crystalline carbohydrates

International Nuclear Information System (INIS)

Yudin, I.V.; Filyanin, G.A.; Panasyuk, S.L.

1990-01-01

In order to determine the nature of the elementary stages of chain process of formation of molecular products in irradiated carbohydrates, the kinetics of their accumulation in crystalline matrices at 100-400 K were investigated. Chain formation of carbonyl products in xylose crystals irradiated at 100 K was identified at temperatures above 240 K and in saccharose, rhamnose, and arabinose crystals at T > 273 K. Chain formation of hydroxy acids with a radiochemical yield of ∼ 150 molecules/100 eV was confirmed in crystalline lactose

SUPPORT IN INCLUSIVE EDUCATION

OpenAIRE

Belma Čičkušić; Ševala Tulumović; Selma Bakić; Salem Bakić

2016-01-01

In order for inclusive class to be successful, associates are of great help to teachers. Besides associates, teachers' specialization can be accomplished through educational seminars on the inclusion topic. However, information about inclusion, working with children with special needs, can also be found in scientific journals that offer more information on methods of working with children with special needs, didactic materials customized according to abilities of children. Aim of ...

Proceedings of the scientific visit on crystalline rock repository development.

Energy Technology Data Exchange (ETDEWEB)

Mariner, Paul E.; Hardin, Ernest L.; Miksova, Jitka [RAWRA, Czech Republic

2013-02-01

A scientific visit on Crystalline Rock Repository Development was held in the Czech Republic on September 24-27, 2012. The visit was hosted by the Czech Radioactive Waste Repository Authority (RAWRA), co-hosted by Sandia National Laboratories (SNL), and supported by the International Atomic Energy Agency (IAEA). The purpose of the visit was to promote technical information exchange between participants from countries engaged in the investigation and exploration of crystalline rock for the eventual construction of nuclear waste repositories. The visit was designed especially for participants of countries that have recently commenced (or recommenced) national repository programmes in crystalline host rock formations. Discussion topics included repository programme development, site screening and selection, site characterization, disposal concepts in crystalline host rock, regulatory frameworks, and safety assessment methodology. Interest was surveyed in establishing a %E2%80%9Cclub,%E2%80%9D the mission of which would be to identify and address the various technical challenges that confront the disposal of radioactive waste in crystalline rock environments. The idea of a second scientific visit to be held one year later in another host country received popular support. The visit concluded with a trip to the countryside south of Prague where participants were treated to a tour of the laboratory and underground facilities of the Josef Regional Underground Research Centre.

Multi-genome identification and characterization of chlamydiae-specific type III secretion substrates: the Inc proteins