bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cu(II)‐Azabis(oxazoline)‐Complexes Immobilized on Superparamagnetic Magnetite@Silica‐Nanoparticles: A Highly Selective and Recyclable Catalyst for the Kinetic Resolution of 1,2‐Diols
Authors: Schätz Alexander , Hager Markus , Reiser Oliver 
Published: May 2009

AbstractTwo different types of azide functionalized magnetite@silica nanoparticles are synthesized, which are ideally suited as inexpensive supports for catalysts and reagents as demonstrated with the grafting of copper(II)‐azabis(oxazoline) complexes via a copper(I) catalyzed azide/alkyne cycloaddition (CuAAC) reaction. The potential of the immobilized complexes as catalysts is tested in the desymmetrization of racemic 1,2‐diols through asymmetric benzoylation. Compared to azabis(oxazolines) “clicked” to common polymeric supports such as MeOPEG or Merrifield resin, Fe3O4@SiO2 proves to be superior with respect to activity and selectivity, as exemplified by employing the catalysts in up to five runs with consistent high activity and selectivity. Recycling of the catalysts is achieved quantitatively by magnetic decantation.
Improved protein splicing reaction for low solubility protein fragments without insertion of native extein residues
Authors: Kamioka Tetsuya , Tawa Minako , Sohya Shihori , Yamazaki Toshio , Kuroda Yutaka 
Published: May 2009

AbstractApplication of trans protein splicing has been limited both by solubility problems and by the insertion of native extein residues (NERs) at the splicing site. Here, we report two simple methods for overcoming these problems and increasing the yield and activity of the spliced product. First, low solubility was alleviated by adding arginine to the reaction buffer and optimizing the splicing reaction condition. The protocol was demonstrated in the context of a Green Fluorescent Protein variant (GFPuv), and the final yield was increased by 1.9‐fold compared to control experiments performed under the same conditions but without addition of arginine. Second, the insertion of NERs was overcome by mutating, instead of inserting, a minimal number of residues in the target protein to amino acids required for the splicing reaction. We identified optimal splicing sites that conserve as much as possible the prerequisite NERs. As a result, the GFPuv residues 142–146 (EYNYN) were mutated to the reportedly minimal required NERs, EYCFN. GFPs spliced using this strategy had no NERs insertion and a fluorescence activity six times stronger than a control GFPuv with five NERs inserted at the splicing site (residue 145/6). In principle, the present protocol (Sw/oNI) can be applied to any target protein, even when no sequence similarity to NERs is present, though it will introduce up to five mutations at the splicing site. © 2009 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 92: 465–470, 2009.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com
Mechanistic basis for differential inhibition of the F1Fo‐ATPase by aurovertin
Authors: Johnson Kathryn M. , Swenson Lara , Opipari Anthony W. , Reuter Rolf , Zarrabi Nawid , Fierke Carol A. , Börsch Michael , Glick Gary D. 
Published: May 2009

AbstractThe mitochondrial F1Fo‐ATPase performs the terminal step of oxidative phosphorylation. Small molecules that modulate this enzyme have been invaluable in helping decipher F1Fo‐ATPase structure, function, and mechanism. Aurovertin is an antibiotic that binds to the β subunits in the F1 domain and inhibits F1Fo‐ATPase‐catalyzed ATP synthesis in preference to ATP hydrolysis. Despite extensive study and the existence of crystallographic data, the molecular basis of the differential inhibition and kinetic mechanism of inhibition of ATP synthesis by aurovertin has not been resolved. To address these questions, we conducted a series of experiments in both bovine heart mitochondria and E. coli membrane F1Fo‐ATPase. Aurovertin is a mixed, noncompetitive inhibitor of both ATP hydrolysis and synthesis with lower Ki values for synthesis. At low substrate concentrations, inhibition is cooperative suggesting a stoichiometry of two aurovertin per F1Fo‐ATPase. Furthermore, aurovertin does not completely inhibit the ATP hydrolytic activity at saturating concentrations. Single‐molecule experiments provide evidence that the residual rate of ATP hydrolysis seen in the presence of saturating concentrations of aurovertin results from a decrease in the binding change mechanism by hindering catalytic site interactions. The results from these studies should further the understanding of how the F1Fo‐ATPase catalyzes ATP synthesis and hydrolysis. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 830–840, 2009.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com
In vitro and in vivo RNA interference mediated suppression of Tn‐caspase‐1 for improved recombinant protein production in High FiveTMcell culture with the baculovirus expression vector system
Authors: Hebert Colin G. , Valdes James J. , Bentley William E. 
Published: May 2009

AbstractWhile traditional metabolic engineering generally relies on the augmentation of specific genes and pathways in order to increase the yield of target proteins, the advent of RNA interference (RNAi) as a biological tool has given metabolic engineers another tool capable of rationally altering the host cell's biological landscape in order to achieve a specific goal. Given its broad applicability and potent specificity, RNAi has the ability to suppress genes whose function is contrary to the desired phenotype. In this study, RNAi has been used to increase recombinant protein production in a Trichoplusia ni derived cell line (BTI‐TN‐5B1‐4—High Five™) using the Baculovirus Expression Vector System. The specific target investigated is Tn‐caspase‐1, a protease involved in apoptosis that is likely the principal effector caspase present in T. ni cells. Experiments were first conducted using in vitro synthesized dsRNA to verify silencing of Tn‐capase‐1 and increased protein production as a result. Subsequent experiments were conducted using a cell line stably expressing in vivo RNAi in the form of an inverted repeat that results in a hairpin upon transcription. Using this construct, Tn‐caspase‐1 transcript levels were decreased by 50% and caspase enzymatic activity was decreased by 90%. This cell line, designated dsTncasp‐2, demonstrates superior viability under low nutrient culture conditions and resulted in as much as two times the protein yield when compared to standard High Five cells. Biotechnol. Bioeng. 2009; 104: 390–399 © 2009 Wiley Periodicals, Inc.
Enrichment of anaerobic methanotrophs in sulfate‐reducing membrane bioreactors
Authors: Meulepas Roel J.W. , Jagersma Christian G. , Gieteling Jarno , Buisman Cees J.N. , Stams Alfons J.M. , Lens Piet N.L. 
Published: May 2009

AbstractAnaerobic oxidation of methane (AOM) in marine sediments is coupled to sulfate reduction (SR). AOM is mediated by distinct groups of archaea, called anaerobic methanotrophs (ANME). ANME co‐exist with sulfate‐reducing bacteria, which are also involved in AOM coupled SR. The microorganisms involved in AOM coupled to SR are extremely difficult to grow in vitro. Here, a novel well‐mixed submerged‐membrane bioreactor system is used to grow and enrich the microorganisms mediating AOM coupled to SR. Four reactors were inoculated with sediment sampled in the Eckernförde Bay (Baltic Sea) and operated at a methane and sulfate loading rate of 4.8 L L−1 day−1 (196 mmol L−1 day−1) and 3.0 mmol L−1 day−1. Two bioreactors were controlled at 15°C and two at 30°C, one reactor at 30°C contained also anaerobic granular sludge. At 15°C, the volumetric AOM and SR rates doubled approximately every 3.8 months. After 884 days, an enrichment culture was obtained with an AOM and SR rate of 1.0 mmol g day−1 (286 µmol g day−1). No increase in AOM and SR was observed in the two bioreactors operated at 30°C. The microbial community of one of the 15°C reactors was analyzed. ANME‐2a became the dominant archaea. This study showed that sulfate reduction with methane as electron donor is possible in well‐mixed bioreactors and that the submerged‐membrane bioreactor system is an excellent system to enrich slow‐growing microorganisms, like methanotrophic archaea. Biotechnol. Bioeng. 2009; 104: 458–470 © 2009 Wiley Periodicals, Inc.
Morphology of Barium Sulfate Crystals from Seeded Precipitation
Authors: Steyer C. , Sundmacher K. 
Published: May 2009

AbstractIt is shown that for seeded precipitation, the morphology and the location of active growth sites of the seed crystals depend on the conditions in which they were produced in terms of supersaturation, ion ratio of reactants, and feed rate. Further, the properties of the seed crystals influence the morphology and, hence, the size of the seeded precipitated particles. The reaction system is barium sulfate from barium chloride and potassium sulfate in a semi‐batch stirred tank reactor.
Penetration of Organic Liquids into Dry Portland Cement Mortar under Ambient and High Pressure
Authors: Seeger A. , Brickmann J. , Luft G. 
Published: May 2009

AbstractThe penetration depths (PDs) of 81 different organic liquids into uncracked dry cement mortar were measured by a suction test at ambient pressure. The physicochemical properties of the liquids (viscosity (η), surface tension (σ), density (ρ), partition coefficient (logP), and molecular volume) were correlated with their penetration depth at normal pressure. It was found that viscosity has the main impact on the penetration. It could be shown by dimensional analysis that the penetration depth at ambient pressure is proportional to the parameter (σρ/η2)1/4. Furthermore, a device to measure the penetration at high pressure up to 50 bar was set up. It was experimentally found that the natural logarithm of the PD at constant time is approximately proportional to the natural logarithm of the applied pressure. A method was elaborated to extrapolate the penetration depth at normal pressure from one single high‐pressure measurement. The advantage of this technique is the significantly shorter measurement time for the high‐pressure test of 15 min up to 1 h compared to the usual tests at normal pressure which require 72 h.
Enantioselective Recognition and Separation by a Homochiral Porous Lamellar Solid Based on Unsymmetrical Schiff Base Metal Complexes
Authors: Yuan Guozan , Zhu Chengfeng , Xuan Weimin , Cui Yong 
Published: May 2009

AbstractOur Schiff has come in: A homochiral porous lamellar solid based on chiral unsymmetrical Schiff base metal complexes was assembled by using coordination and hydrogen bonds (see picture). The host features a flexible helical framework and chiral amphiphilic channel surfaces capable of multiple interactions with guest species.magnified imageA homochiral lamellar supramolecular host system has been developed by treating 3‐[(E)‐{[(1R,2R)‐2‐aminocyclohexyl]imino}methyl]‐4‐hydroxybenzoic acid (H2L) with copper nitrate and characterized by a variety of techniques including microanalysis, IR spectroscopy, thermogravimetric analysis (TGA), circular dichroism (CD) spectroscopy, and powder and single‐crystal X‐ray diffraction. Tridentate Schiff base L ligands link adjacent metal centers to form 1D coordination polymeric chains using tridentate N2O donors and carboxylate groups, while interchain metal–oxygen and hydrogen‐bonding interactions further link 1D polymeric chains to a porous lamellar solid. The host features a flexible helical framework for guest inclusions and chiral amphiphilic channel surfaces lined with amine hydrogen atoms and aliphatic groups capable of multiple interactions with guest species. It allows for the development of an efficient approach for highly enantioselective separation of racemic secondary alcohols by inclusion crystallization followed by distillation (>99.5 % ee). In addition, the host including different alcohols exhibits guest‐responsive ferroelectric behavior.
Photonucleophilic Addition of the ε‐Amino Group of Lysine to a Triflusal Metabolite as a Mechanistic Key to Photoallergy Mediated by the Parent Drug
Authors: Montanaro Sara , Lhiaubet‐Vallet Virginie , Jiménez M. Consuelo , Blanca Miguel , Miranda Miguel Angel 
Published: May 2009

AbstractA mechanism for triflusal‐induced photoallergyinvolving complexation of 2‐hydroxy‐4‐trifluoromethylbenzoic acid with site I of human serum albumin and subsequent formation of a covalent adduct by photoreaction between a metabolite and a neighboring lysine residue is proposed. This is supported by the observed photobinding to poly‐L‐lysine. Thereby, a photoantigen is generated, which is a likely trigger of the immune response.magnified imageThe goal of the work presented herein is to gain deeper insight into the molecular basis of photoallergy mediated by triflusal through its active metabolite, 2‐hydroxy‐4‐trifluoromethylbenzoic acid (HTB). For this purpose, the interaction between HTB and human serum albumin (HSA) was investigated by fluorescence and laser flash photolysis to monitor inclusion into the protein binding sites through variation in the excited‐state properties. A remarkable lengthening of HTB triplet lifetime in the presence of HSA was observed. The use of oleic acid as a displacement probe clearly suggests the preference for dark binding in site I. The mechanism of photobinding was studied by irradiation of HTB in the presence of amino acids, and, in the case of lysine, a photoadduct was detected that arises from nucleophilic attack by the ε‐amino group to the trifluoromethyl substituent of HTB. Accordingly, photobinding of the metabolite to poly‐L‐lysine was also observed. Overall, these results are consistent with a mechanism for triflusal photoallergy involving complexation of HTB to site I of HSA and subsequent formation of a covalent photoadduct with one neighboring lysine residue.
Recent Advances in Solution NMR: Fast Methods and Heteronuclear Direct Detection
Authors: Felli Isabella C. , Brutscher Bernhard 
Published: May 2009

AbstractNMR spectroscopy for the molecular life sciences: Two areas of recent active research [fast methods and direct detection of low‐γ nuclei (see schematic)] are reviewed herein. These are powerful new tools for structural characterization of short‐lived molecules, large and intrinsically unstructured proteins, paramagnetic systems, as well as for the study of molecular kinetic processes at atomic resolution.magnified imageToday, NMR spectroscopy is the technique of choice to investigate molecular structure, dynamics, and interactions in solution at atomic resolution. A major limitation of NMR spectroscopy for the study of biological macromolecules such as proteins, nucleic acids, and their complexes, has always been its low sensitivity, a consequence of the weak magnetic spin interactions. Therefore many efforts have been invested in the last decade to improve NMR instrumentation in terms of experimental sensitivity. As a result of these efforts, the availability of high‐field magnets, cryogenically cooled probes, and probably in the near future hyperpolarization techniques, the intrinsic NMR sensitivity has increased by at least one order of magnitude. Stimulated by new challenges in the life sciences, these technical improvements have triggered the development of new NMR methods for the study of molecular systems of increasing size and complexity. Herein, we focus on two examples of recently developed NMR methodologies. First, advanced multidimensional data acquisition schemes provide a speed increase of several orders of magnitude. Second, NMR methods based on the direct detection of low‐gamma nuclei present a new spectroscopic tool, highly complementary to conventional NMR techniques. These new methods provide powerful new NMR tools for the study of short‐lived molecules, large and intrinsically unstructured proteins, paramagnetic systems, as well as for the characterization of molecular kinetic processes at atomic resolution. These examples illustrate how NMR is continuously adapting to the new challenges in the life sciences, with the focus shifting from the characterization of single biomolecules to an integrated view of interacting molecular networks observed at varying levels of biological organization.
Limitations of Induced Folding in Molecular Recognition by Intrinsically Disordered Proteins
Authors: Hazy Eszter , Tompa Peter 
Published: May 2009

AbstractIntrinsically disordered proteins exist and function without well‐defined three‐dimensional structures and are common in proteomes where they carry out essential functions. Herein, evidence for their lack of structure and the major functional benefits that this confers, are surveyed. An example is Tβ4, a sequence that is disordered until it binds to G‐actin, when it adopts an ordered conformation (see picture).magnified imageIntrinsically disordered proteins (IDPs) exist and function without well‐defined three‐dimensional structures, thus they defy the classical structure–function paradigm. These proteins are common in proteomes, and they carry out essential functions often related to signalling and regulation of transcription. Herein, the experimental evidence for their lack of structure and the major functional benefits that structural disorder confers, are surveyed. It is shown that IDPs often function by molecular recognition, in which either short motifs, or domain‐sized disordered segments are used for partner recognition. In both cases, the binding segment undergoes induced folding and it attains an ordered structure. This folding‐upon‐binding scenario suggests that the function of IDPs can be interpreted in terms of the static structural view of the classical paradigm. New developments in the field, however, suggest that folding upon binding is limited, and many IDPs preserve a significant level of disorder in the bound state, a phenomenon termed fuzziness. In addition, IDPs may structurally adapt to different partners with different functional outcomes, resulting in promiscuity in function termed moonlighting. It is suggested that a new model describing the structure–function relationship of IDPs has to encompass such structural and functional promiscuity inherent in the disordered state of IDPs.
Epitope‐tagging Math5 and Pou4f2: New tools to study retinal ganglion cell development in the mouse
Authors: Fu Xueyao , Kiyama Takae , Li Renzhong , Russell Mark , Klein William H. , Mu Xiuqian 
Published: May 2009

AbstractAlthough immunological detection of proteins is used extensively in retinal development, studies are often impeded because antibodies against crucial proteins cannot be generated or are not readily available. Here, we overcome these limitations by constructing genetically engineered alleles for Math5 and Pou4f2, two genes required for retinal ganglion cell (RGC) development. Sequences encoding a peptide epitope from haemagglutinin (HA) were added to Math5 or Pou4f2 in frame to generate Math5HA and Pou4f2HA alleles. We demonstrate that the tagged alleles recapitulated the wild‐type expression patterns of the two genes, and that the tags did not interfere with the function of the cognate proteins. In addition, by co‐staining, we found that Math5 and Pou4f2 were transiently co‐expressed in newly born RGCs, unequivocally demonstrating that Pou4f2 is immediately downstream of Math5 in RGC formation. The epitope‐tagged alleles provide new and useful tools for analyzing gene regulatory networks underlying RGC development. Developmental Dynamics 238:2309–2317, 2009. © 2009 Wiley‐Liss, Inc.
CTLA‐4 is required by CD4+CD25+ Treg to control CD4+ T‐cell lymphopenia‐induced proliferation
Authors: Sojka Dorothy K. , Hughson Angela , Fowell Deborah J. 
Published: May 2009

AbstractCTLA‐4 is constitutively expressed by CD4+CD25+Foxp3+ Treg but its precise role in Treg function is not clear. Although blockade of CTLA‐4 interferes with Treg function, studies using CTLA‐4‐deficient Treg have failed to reveal an essential requirement for CTLA‐4 in Treg suppression in vivo. Conditional deletion of CTLA‐4 in Foxp3+ T cells disrupts immune homeostasis in vivo but the immune processes disrupted by CTLA‐4 deletion have not been determined. We demonstrate that Treg expression of CTLA‐4 is essential for Treg control of lymphopenia‐induced CD4 T‐cell expansion. Despite IL‐10 expression, CTLA‐4‐deficient Treg were unable to control the expansion of CD4+ target cells in a lymphopenic environment. Moreover, unlike their WT counterparts, CTLA‐4‐deficient Treg failed to inhibit cytokine production associated with homeostatic expansion and were unable to prevent colitis. Thus, while Treg developing in the absence of CTLA‐4 appear to acquire some compensatory suppressive mechanisms in vitro, we identify a non‐redundant role for CTLA‐4 in Treg function in vivo.
Early simultaneous production of intranodal CD4 Th2 effectors and recirculating rapidly responding central‐memory‐like CD4 T cells
Authors: Serre Karine , Mohr Elodie , Toellner Kai‐Michael , Cunningham Adam F. , Bird Roger , Khan Mahmood , MacLennan Ian C. M. 
Published: May 2009

AbstractThis study characterizes the diversity of CD4 Th cells produced during a Th2 response in vivo. Kinetics of effector and memory cell differentiation by mouse OVA‐specific CD4 T cells was followed during primary responses to alum‐precipitated OVA. The complexity of the CD4 T response was assessed in nodes draining and distant from the site of immunization for phenotype, location and function. By 3 days IL‐4‐producing effector CD4 T cells developed in the draining node and a proportion of the responding cells had migrated to B‐cell follicles, while yet others had left the draining node. Some of these early migrant cells were recirculating cells with a central memory phenotype. These had divided four or more times in the draining node before migrating to distant nodes not exposed to antigen. We questioned the responsiveness of these early central‐memory‐like cells on secondary antigen challenge at sites distant to the primary immunization. They re‐entered cell cycle and migrated to B‐cell follicles, much more rapidly than naïve CD4 T cells and could still be induced to produce IL‐4. Their production and survival were independent of the starting frequency of antigen‐specific CD4 T cells. Thus intranodal effector cells and recirculating, rapidly responding central‐memory‐like cells emerged simultaneously from the third day of a primary Th2 response.
Antigens expressed by myelinating glia cells induce peripheral cross‐tolerance of endogenous CD8+ T cells
Authors: Schildknecht Anita , Probst Hans Christian , McCoy Kathy D. , Miescher Iris , Brenner Corinne , Leone Dino P. , Suter Ueli , Ohashi Pamela S. , van den Broek Maries 
Published: May 2009

AbstractAuto‐reactivity of T cells is largely prevented by central and peripheral tolerance. Nevertheless, immunization with certain self‐antigens emulsified in CFA induces autoimmunity in rodents, suggesting that tolerance to some self‐antigens is not robust. To investigate the fate of nervous system‐specific CD8+ T cells, which only recently came up as being important contributors for MS pathogenesis, we developed a mouse model that allows inducible expression of lymphocytic choriomeningitis virus‐derived CD8+ T‐cell epitopes specifically in oligodendrocytes and Schwann cells, the myelinating glia of the nervous system. These transgenic CD8+ T‐cell epitopes induced robust tolerance of endogenous auto‐reactive T cells, which proved thymus‐independent and was mediated by cross‐presenting bone‐marrow‐derived cells. Immunohistological staining of secondary lymphoid organs demonstrated the presence of glia‐derived antigens in DC, suggesting that peripheral tolerance of CD8+ T cells results from uptake and presentation by steady state DC.