Structure-Guided Functional Characterization of DUF1460 Reveals a Highly Specific NlpC/P60 Amidase Family

DOE PAGES

Xu, Qingping; Mengin-Lecreulx, Dominique; Patin, Delphine; ...

2014-11-20

GlcNAc-1,6-anhydro-MurNAc-tetrapeptide is a major peptidoglycan degradation intermediate and a cytotoxin. It is generated by lytic transglycosylases and further degraded and recycled by various enzymes. We have identified and characterized a novel, highly specific N-acetylmuramoyl-L-alanine amidase (AmiA) from Bacteroides uniformis, a member of the DUF1460 protein family, that hydrolyzes GlcNAc-1,6-anhydro-MurNAc-peptide into disaccharide and stem peptide. The high-resolution apo-structure at 1.15 Å resolution shows that AmiA is related to NlpC/P60 γ-D-Glu-meso-diaminopimelic acid amidases and shares a common catalytic core and cysteine peptidase-like active site. AmiA has evolved structural adaptations that reconfigure the substrate recognition site. The preferred substrates for AmiA were predictedmore » in silico based on structural and bioinformatics data, and were subsequently characterized experimentally. Ultimately, further crystal structures of AmiA in complexes with GlcNAc-1,6-anhydro-MurNAc and GlcNAc have enabled us to elucidate substrate recognition and specificity. DUF1460 is highly conserved in structure and defines a new amidase family.« less

Structure-guided functional characterization of DUF1460 reveals a highly specific NlpC/P60 amidase family.

PubMed

Xu, Qingping; Mengin-Lecreulx, Dominique; Patin, Delphine; Grant, Joanna C; Chiu, Hsiu-Ju; Jaroszewski, Lukasz; Knuth, Mark W; Godzik, Adam; Lesley, Scott A; Elsliger, Marc-André; Deacon, Ashley M; Wilson, Ian A

2014-12-02

GlcNAc-1,6-anhydro-MurNAc-tetrapeptide is a major peptidoglycan degradation intermediate and a cytotoxin. It is generated by lytic transglycosylases and further degraded and recycled by various enzymes. We have identified and characterized a highly specific N-acetylmuramoyl-L-alanine amidase (AmiA) from Bacteroides uniformis, a member of the DUF1460 protein family, that hydrolyzes GlcNAc-1,6-anhydro-MurNAc-peptide into disaccharide and stem peptide. The high-resolution apo structure at 1.15 Å resolution shows that AmiA is related to NlpC/P60 γ-D-Glu-meso-diaminopimelic acid amidases and shares a common catalytic core and cysteine peptidase-like active site. AmiA has evolved structural adaptations that reconfigure the substrate recognition site. The preferred substrates for AmiA were predicted in silico based on structural and bioinformatics data, and subsequently were characterized experimentally. Further crystal structures of AmiA in complexes with GlcNAc-1,6-anhydro-MurNAc and GlcNAc have enabled us to elucidate substrate recognition and specificity. DUF1460 is highly conserved in structure and defines another amidase family. Copyright © 2014 Elsevier Ltd. All rights reserved.

Structure-Guided Functional Characterization of DUF1460 Reveals a Highly Specific NlpC/P60 Amidase Family

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Qingping; Mengin-Lecreulx, Dominique; Patin, Delphine

GlcNAc-1,6-anhydro-MurNAc-tetrapeptide is a major peptidoglycan degradation intermediate and a cytotoxin. It is generated by lytic transglycosylases and further degraded and recycled by various enzymes. We have identified and characterized a novel, highly specific N-acetylmuramoyl-L-alanine amidase (AmiA) from Bacteroides uniformis, a member of the DUF1460 protein family, that hydrolyzes GlcNAc-1,6-anhydro-MurNAc-peptide into disaccharide and stem peptide. The high-resolution apo-structure at 1.15 Å resolution shows that AmiA is related to NlpC/P60 γ-D-Glu-meso-diaminopimelic acid amidases and shares a common catalytic core and cysteine peptidase-like active site. AmiA has evolved structural adaptations that reconfigure the substrate recognition site. The preferred substrates for AmiA were predictedmore » in silico based on structural and bioinformatics data, and were subsequently characterized experimentally. Ultimately, further crystal structures of AmiA in complexes with GlcNAc-1,6-anhydro-MurNAc and GlcNAc have enabled us to elucidate substrate recognition and specificity. DUF1460 is highly conserved in structure and defines a new amidase family.« less

A novel S-enantioselective amidase acting on 3,3,3-trifluoro-2-hydroxy-2-methylpropanamide from Arthrobacter sp. S-2.

PubMed

Fuhshuku, Ken-ichi; Watanabe, Shunsuke; Nishii, Tetsuro; Ishii, Akihiro; Asano, Yasuhisa

2015-01-01

A novel S-enantioselective amidase acting on 3,3,3-trifluoro-2-hydroxy-2-methylpropanamide was purified from Arthrobacter sp. S-2. The enzyme acted S-enantioselectively on 3,3,3-trifluoro-2-hydroxy-2-methylpropanamide to yield (S)-3,3,3-trifluoro-2-hydroxy-2-methylpropanoic acid. Based on the N-terminal amino acid sequence of this amidase, the gene coding S-amidase was cloned from the genomic DNA of Arthrobacter sp. S-2 and expressed in an Escherichia coli host. The recombinant S-amidase was purified and characterized. Furthermore, the purified recombinant S-amidase with the C-His6-tag, which was expressed in E. coli as the C-His6-tag fusion protein, was used in the kinetic resolution of (±)-3,3,3-trifluoro-2-hydroxy-2-methylpropanamide to obtain (S)-3,3,3-trifluoro-2-hydroxy-2-methylpropanoic acid and (R)-3,3,3-trifluoro-2-hydroxy-2-methylpropanamide.

Crystal structure analysis of a bacterial aryl acylamidase belonging to the amidase signature enzyme family

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Saeyoung; Park, Eun-Hye; Ko, Hyeok-Jin

2015-11-13

The atomic structure of a bacterial aryl acylamidase (EC 3.5.1.13; AAA) is reported and structural features are investigated to better understand the catalytic profile of this enzyme. Structures of AAA were determined in its native form and in complex with the analgesic acetanilide, p-acetaminophenol, at 1.70 Å and 1.73 Å resolutions, respectively. The overall structural fold of AAA was identified as an α/β fold class, exhibiting an open twisted β-sheet core surrounded by α-helices. The asymmetric unit contains one AAA molecule and the monomeric form is functionally active. The core structure enclosing the signature sequence region, including the canonical Ser-cisSer-Lys catalytic triad,more » is conserved in all members of the Amidase Signature enzyme family. The structure of AAA in a complex with its ligand reveals a unique organization in the substrate-binding pocket. The binding pocket consists of two loops (loop1 and loop2) in the amidase signature sequence and one helix (α10) in the non-amidase signature sequence. We identified two residues (Tyr{sup 136} and Thr{sup 330}) that interact with the ligand via water molecules, and a hydrogen-bonding network that explains the catalytic affinity over various aryl acyl compounds. The optimum activity of AAA at pH > 10 suggests that the reaction mechanism employs Lys{sup 84} as the catalytic base to polarize the Ser{sup 187} nucleophile in the catalytic triad. - Highlights: • We determined the first structure of a bacterial aryl acylamidase (EC 3.5.1.13). • Structure revealed spatially distinct architecture of the substrate-binding pocket. • Hydrogen-bonding with Tyr{sup 136} and Thr{sup 330} mediates ligand-binding and substrate.« less

Enabling cell-cell communication via nanopore formation: structure, function and localization of the unique cell wall amidase AmiC2 of Nostoc punctiforme.

PubMed

Büttner, Felix M; Faulhaber, Katharina; Forchhammer, Karl; Maldener, Iris; Stehle, Thilo

2016-04-01

To orchestrate a complex life style in changing environments, the filamentous cyanobacterium Nostoc punctiforme facilitates communication between neighboring cells through septal junction complexes. This is achieved by nanopores that perforate the peptidoglycan (PGN) layer and traverse the cell septa. The N-acetylmuramoyl-l-alanine amidase AmiC2 (Npun_F1846; EC 3.5.1.28) in N. punctiforme generates arrays of such nanopores in the septal PGN, in contrast to homologous amidases that mediate daughter cell separation after cell division in unicellular bacteria. Nanopore formation is therefore a novel property of AmiC homologs. Immunofluorescence shows that native AmiC2 localizes to the maturing septum. The high-resolution crystal structure (1.12 Å) of its catalytic domain (AmiC2-cat) differs significantly from known structures of cell splitting and PGN recycling amidases. A wide and shallow binding cavity allows easy access of the substrate to the active site, which harbors an essential zinc ion. AmiC2-cat exhibits strong hydrolytic activity in vitro. A single point mutation of a conserved glutamate near the zinc ion results in total loss of activity, whereas zinc removal leads to instability of AmiC2-cat. An inhibitory α-helix, as found in the Escherichia coli AmiC(E. coli) structure, is absent. Taken together, our data provide insight into the cell-biological, biochemical and structural properties of an unusual cell wall lytic enzyme that generates nanopores for cell-cell communication in multicellular cyanobacteria. The novel structural features of the catalytic domain and the unique biological function of AmiC2 hint at mechanisms of action and regulation that are distinct from other amidases. The AmiC2-cat structure has been deposited in the Protein Data Bank under accession number 5EMI. © 2016 Federation of European Biochemical Societies.

Structure-Function Analysis of Staphylococcus aureus Amidase Reveals the Determinants of Peptidoglycan Recognition and Cleavage*

PubMed Central

Büttner, Felix Michael; Zoll, Sebastian; Nega, Mulugeta; Götz, Friedrich; Stehle, Thilo

2014-01-01

The bifunctional major autolysin AtlA of Staphylococcus aureus cleaves the bacterium's peptidoglycan network (PGN) at two distinct sites during cell division. Deletion of the enzyme results in large cell clusters with disordered division patterns, indicating that AtlA could be a promising target for the development of new antibiotics. One of the two functions of AtlA is performed by the N-acetylmuramyl-l-alanine amidase AmiA, which cleaves the bond between the carbohydrate and the peptide moieties of PGN. To establish the structural requirements of PGN recognition and the enzymatic mechanism of cleavage, we solved the crystal structure of the catalytic domain of AmiA (AmiA-cat) in complex with a peptidoglycan-derived ligand at 1.55 Å resolution. The peptide stem is clearly visible in the structure, forming extensive contacts with protein residues by docking into an elongated groove. Less well defined electron density and the analysis of surface features indicate likely positions of the carbohydrate backbone and the pentaglycine bridge. Substrate specificity analysis supports the importance of the pentaglycine bridge for fitting into the binding cleft of AmiA-cat. PGN of S. aureus with l-lysine tethered with d-alanine via a pentaglycine bridge is completely hydrolyzed, whereas PGN of Bacillus subtilis with meso-diaminopimelic acid directly tethered with d-alanine is not hydrolyzed. An active site mutant, H370A, of AmiA-cat was completely inactive, providing further support for the proposed catalytic mechanism of AmiA. The structure reported here is not only the first of any bacterial amidase in which both the PGN component and the water molecule that carries out the nucleophilic attack on the carbonyl carbon of the scissile bond are present; it is also the first peptidoglycan amidase complex structure of an important human pathogen. PMID:24599952

Structure-function analysis of Staphylococcus aureus amidase reveals the determinants of peptidoglycan recognition and cleavage.

PubMed

Büttner, Felix Michael; Zoll, Sebastian; Nega, Mulugeta; Götz, Friedrich; Stehle, Thilo

2014-04-18

The bifunctional major autolysin AtlA of Staphylococcus aureus cleaves the bacterium's peptidoglycan network (PGN) at two distinct sites during cell division. Deletion of the enzyme results in large cell clusters with disordered division patterns, indicating that AtlA could be a promising target for the development of new antibiotics. One of the two functions of AtlA is performed by the N-acetylmuramyl-l-alanine amidase AmiA, which cleaves the bond between the carbohydrate and the peptide moieties of PGN. To establish the structural requirements of PGN recognition and the enzymatic mechanism of cleavage, we solved the crystal structure of the catalytic domain of AmiA (AmiA-cat) in complex with a peptidoglycan-derived ligand at 1.55 Å resolution. The peptide stem is clearly visible in the structure, forming extensive contacts with protein residues by docking into an elongated groove. Less well defined electron density and the analysis of surface features indicate likely positions of the carbohydrate backbone and the pentaglycine bridge. Substrate specificity analysis supports the importance of the pentaglycine bridge for fitting into the binding cleft of AmiA-cat. PGN of S. aureus with l-lysine tethered with d-alanine via a pentaglycine bridge is completely hydrolyzed, whereas PGN of Bacillus subtilis with meso-diaminopimelic acid directly tethered with d-alanine is not hydrolyzed. An active site mutant, H370A, of AmiA-cat was completely inactive, providing further support for the proposed catalytic mechanism of AmiA. The structure reported here is not only the first of any bacterial amidase in which both the PGN component and the water molecule that carries out the nucleophilic attack on the carbonyl carbon of the scissile bond are present; it is also the first peptidoglycan amidase complex structure of an important human pathogen.

Increasing plant growth by modulating omega-amidase expression in plants

DOEpatents

Unkefer, Pat J.; Anderson, Penelope S.; Knight, Thomas J.

2015-06-30

The present disclosure relates to compositions and methods for increasing the leaf-to-root ratio of the signal metabolite 2-oxoglutaramate and related proline molecules in plants by modulating levels of .omega.-amidase to increase nitrogen use efficiency, resulting in enhanced growth, faster growth rates, greater seed and fruit/pod yields, earlier and more productive flowering, increased tolerance to high salt conditions, and increased biomass yields.

A breakthrough in enzyme technology to fight penicillin resistance-industrial application of penicillin amidase.

PubMed

Buchholz, Klaus

2016-05-01

Enzymatic penicillin hydrolysis by penicillin amidase (also penicillin acylase, PA) represents a Landmark: the first industrially and economically highly important process using an immobilized biocatalyst. Resistance of infective bacteria to antibiotics had become a major topic of research and industrial activities. Solutions to this problem, the antibiotics resistance of infective microorganisms, required the search for new antibiotics, but also the development of derivatives, notably penicillin derivatives, that overcame resistance. An obvious route was to hydrolyse penicillin to 6-aminopenicillanic acid (6-APA), as a first step, for the introduction via chemical synthesis of various different side chains. Hydrolysis via chemical reaction sequences was tedious requiring large amounts of toxic chemicals, and they were cost intensive. Enzymatic hydrolysis using penicillin amidase represented a much more elegant route. The basis for such a solution was the development of techniques for enzyme immobilization, a highly difficult task with respect to industrial application. Two pioneer groups started to develop solutions to this problem in the late 1960s and 1970s: that of Günter Schmidt-Kastner at Bayer AG (Germany) and that of Malcolm Lilly of Imperial College London. Here, one example of this development, that at Bayer, will be presented in more detail since it illustrates well the achievement of a solution to the problems of industrial application of enzymatic processes, notably development of an immobilization method for penicillin amidase suitable for scale up to application in industrial reactors under economic conditions. A range of bottlenecks and technical problems of large-scale application had to be overcome. Data giving an inside view of this pioneer achievement in the early phase of the new field of biocatalysis are presented. The development finally resulted in a highly innovative and commercially important enzymatic process to produce 6-APA that

New generation of amino coumarin methyl sulfonate-based fluorogenic substrates for amidase assays in droplet-based microfluidic applications.

PubMed

Woronoff, Gabrielle; El Harrak, Abdeslam; Mayot, Estelle; Schicke, Olivier; Miller, Oliver J; Soumillion, Patrice; Griffiths, Andrew D; Ryckelynck, Michael

2011-04-15

Droplet-based microfluidics is a powerful tool for biology and chemistry as it allows the production and the manipulation of picoliter-size droplets acting as individual reactors. In this format, high-sensitivity assays are typically based on fluorescence, so fluorophore exchange between droplets must be avoided. Fluorogenic substrates based on the coumarin leaving group are widely used to measure a variety of enzymatic activities, but their application in droplet-based microfluidic systems is severely impaired by the fast transport of the fluorescent product between compartments. Here we report the synthesis of new amidase fluorogenic substrates based on 7-aminocoumarin-4-methanesulfonic acid (ACMS), a highly water-soluble dye, and their suitability for droplet-based microfluidics applications. Both substrate and product had the required spectral characteristics and remained confined in droplets from hours to days. As a model experiment, a phenylacetylated ACMS was synthesized and used as a fluorogenic substrate of Escherichia coli penicillin G acylase. Kinetic parameters (k(cat) and K(M)) measured in bulk and in droplets on-chip were very similar, demonstrating the suitability of this synthesis strategy to produce a variety of ACMS-based substrates for assaying amidase activities both in microtiter plate and droplet-based microfluidic formats. © 2011 American Chemical Society

The quaternary structure of the amidase from Geobacillus pallidus RAPc8 is revealed by its crystal packing

DOE Office of Scientific and Technical Information (OSTI.GOV)

Agarkar, Vinod B.; Kimani, Serah W.; Cowan, Donald A.

2006-12-01

The amidase from G. pallidus RAPc8, a moderate thermophile, converts amides to the corresponding acids and ammonia and has application as an industrial catalyst. RAPc8 amidase has been cloned, expressed and purified, and then crystallized using the hanging-drop vapour-diffusion method. The amidase from Geobacillus pallidus RAPc8, a moderate thermophile, is a member of the nitrilase enzyme superfamily. It converts amides to the corresponding acids and ammonia and has application as an industrial catalyst. RAPc8 amidase has been cloned and functionally expressed in Escherichia coli and has been purified by heat treatment and a number of chromatographic steps. The enzyme wasmore » crystallized using the hanging-drop vapour-diffusion method. Crystals produced in the presence of 1.2 M sodium citrate, 400 mM NaCl, 100 mM sodium acetate pH 5.6 were selected for X-ray diffraction studies. A data set having acceptable statistics to 1.96 Å resolution was collected under cryoconditions using an in-house X-ray source. The space group was determined to be primitive cubic P4{sub 2}32, with unit-cell parameter a = 130.49 (±0.05) Å. The structure was solved by molecular replacement using the backbone of the hypothetical protein PH0642 from Pyrococcus horikoshii (PDB code 1j31) with all non-identical side chains substituted with alanine as a probe. There is one subunit per asymmetric unit. The subunits are packed as trimers of dimers with D3 point-group symmetry around the threefold axis in such a way that the dimer interface seen in the homologues is preserved.« less

Amidase encapsulated O-carboxymethyl chitosan nanoparticles for vaccine delivery.

PubMed

Smitha, K T; Sreelakshmi, M; Nisha, N; Jayakumar, R; Biswas, Raja

2014-02-01

This work reports the development of amidase encapsulated O-carboxymethyl chitosan nanoparticles (Ami-O-CMC NPs) of 300±50 nm size by ionic cross-linking method. The prepared Ami-O-CMC NPs had an encapsulation efficiency of 55.39%. Haemolysis assay and cytotoxicity studies proved the hemocompatibility and cytocompatibility of the prepared NPs. The sustained release of Ami from the NPs is expected to prolong its immunogenicity and in turn lead to development of better protective immunity against Staphylococcus aureus infections. Copyright © 2013 Elsevier B.V. All rights reserved.

A peptidoglycan recognition protein from Sciaenops ocellatus is a zinc amidase and a bactericide with a substrate range limited to Gram-positive bacteria.

PubMed

Li, Mo-Fei; Zhang, Min; Wang, Chun-Lin; Sun, Li

2012-02-01

Peptidoglycan recognition proteins (PGRPs) are a family of innate immune molecules that recognize bacterial peptidoglycan. PGRPs are highly conserved in invertebrates and vertebrates including fish. However, the biological function of teleost PGRP remains largely uninvestigated. In this study, we identified a PGRP homologue, SoPGLYRP-2, from red drum (Sciaenops ocellatus) and analyzed its activity and potential function. The deduced amino acid sequence of SoPGLYRP-2 is composed of 482 residues and shares 46-94% overall identities with known fish PGRPs. SoPGLYRP-2 contains at the C-terminus a single zinc amidase domain with conserved residues that form the catalytic site. Quantitative RT-PCR analysis detected SoPGLYRP-2 expression in multiple tissues, with the highest expression occurring in liver and the lowest expression occurring in brain. Experimental bacterial infection upregulated SoPGLYRP-2 expression in kidney, spleen, and liver in time-dependent manners. To examine the biological activity of SoPGLYRP-2, purified recombinant proteins representing the intact SoPGLYRP-2 (rSoPGLYRP-2) and the amidase domain (rSoPGLYRP-AD) were prepared from Escherichia coli. Subsequent analysis showed that rSoPGLYRP-2 and rSoPGLYRP-AD (i) exhibited comparable Zn(2+)-dependent peptidoglycan-lytic activity and were able to recognize and bind to live bacterial cells, (ii) possessed bactericidal effect against Gram-positive bacteria and slight bacteriostatic effect against Gram-negative bacteria, (iii) were able to block bacterial infection into host cells. These results indicate that SoPGLYRP-2 is a zinc-dependent amidase and a bactericide that targets preferentially at Gram-positive bacteria, and that SoPGLYRP-2 is likely to play a role in host innate immune defense during bacterial infection. Copyright © 2011 Elsevier Ltd. All rights reserved.

Structural requirements of choline derivatives for 'conversion' of pneumococcal amidase. A new single-step procedure for purification of this autolysin.

PubMed

Sanz, J M; Lopez, R; Garcia, J L

1988-05-23

Tertiary amines appear to be the minimal structure needed to convert in vitro the inactive form (E-form) of pneumococcal amidase to the catalytic active form (C-form). Diethylethanolamine was one of the compounds that converted the E-form, a finding that has been used successfully to develop an affinity chromatography system in DEAE-cellulose for the rapid and efficient purification of lytic enzymes of pneumococcus and its bacteriophages.

Human butyrylcholinesterase components differ in aryl acylamidase activity.

PubMed

Montenegro, María F; María, T Moral-Naranjo; de la Cadena, María Páez; Campoy, Francisco J; Muñoz-Delgado, Encarnación; Vidal, Cecilio J

2008-04-01

Apart from its esterase activity, butyrylcholinesterase (BuChE) displays aryl acylamidase (AAA) activity able to hydrolyze o-nitroacetanilide (ONA) and its trifluoro-derivative (F-ONA). We report here that, despite amidase and esterase sites residing in the same protein, in human samples depleted of acetylcholinesterase the ratio of amidase to esterase activity varied depending on the source of BuChE. The much faster degradation of ONA and F-ONA by BuChE monomers (G1) of colon and kidney than by the tetramers (G4) suggests aggregation-driven differences in the AAA site between single and polymerized subunits. The similar ratio of F-ONAto butyrylthiocholine hydrolysis by serum G1 and G4 forms support structural differences in the amidase site according to the source of BuChE. The changing ratios of amidase to esterase activities in the human sources probably arise from post-translational modifications in BuChE subunits, the specific proportion of monomers and oligomers and the variable capacity of the tetramers for degrading ONA and F-ONA. The elevated amidase activity of BuChE monomers and the scant activity of the tetramers justify the occurrence of single BuChE subunits in cells as a means to sustain the AAA activity of BuChE which otherwise could be lost by tetramerization.

N-Acetylanthranilate Amidase from Arthrobacter nitroguajacolicus Rü61a, an α/β-Hydrolase-Fold Protein Active towards Aryl-Acylamides and -Esters, and Properties of Its Cysteine-Deficient Variant▿ †

PubMed Central

Kolkenbrock, Stephan; Parschat, Katja; Beermann, Bernd; Hinz, Hans-Jürgen; Fetzner, Susanne

2006-01-01

N-acetylanthranilate amidase (Amq), a 32.8-kDa monomeric amide hydrolase, is involved in quinaldine degradation by Arthrobacter nitroguajacolicus Rü61a. Sequence analysis and secondary structure predictions indicated that Amq is related to carboxylesterases and belongs to the α/β-hydrolase-fold superfamily of enzymes; inactivation of (His6-tagged) Amq by phenylmethanesulfonyl fluoride and diethyl pyrocarbonate and replacement of conserved residues suggested a catalytic triad consisting of S155, E235, and H266. Amq is most active towards aryl-acetylamides and aryl-acetylesters. Remarkably, its preference for ring-substituted analogues was different for amides and esters. Among the esters tested, phenylacetate was hydrolyzed with highest catalytic efficiency (kcat/Km = 208 mM−1 s−1), while among the aryl-acetylamides, o-carboxy- or o-nitro-substituted analogues were preferred over p-substituted or unsubstituted compounds. Hydrolysis by His6Amq of primary amides, lactams, N-acetylated amino acids, azocoll, tributyrin, and the acylanilide and urethane pesticides propachlor, propham, carbaryl, and isocarb was not observed; propanil was hydrolyzed with 1% N-acetylanthranilate amidase activity. The catalytic properties of the cysteine-deficient variant His6AmqC22A/C63A markedly differed from those of His6Amq. The replacements effected some changes in Kms of the enzyme and increased kcats for most aryl-acetylesters and some aryl-acetylamides by factors of about three to eight while decreasing kcat for the formyl analogue N-formylanthranilate by several orders of magnitude. Circular dichroism studies indicated that the cysteine-to-alanine replacements resulted in significant change of the overall fold, especially an increase in α-helicity of the cysteine-deficient protein. The conformational changes may also affect the active site and may account for the observed changes in kinetic properties. PMID:17041061

Enzyme immobilization techniques on poly(glycidyl methacrylate-co-ethylene dimethacrylate) carrier with penicillin amidase as model.

PubMed

Drobník, J; Saudek, V; Svec, F; Kálal, J; Vojtísek, V; Bárta, M

1979-08-01

Two types of bead-form macroporous carriers based on glycidyl methacrylate with ethylene dimethacrylate copolymers were used for the immobilization of penicillin amidase either directly or after chemical modification. Direct binding through oxirane groups, which is equally efficient at pH 4.2 and 7, is relatively slow and brings about an activity loss at low enzyme concentrations. The most efficient immobilization was achieved on glutaraldehyde-activated amino carrier, irrespective of whether the amino groups were formed by ammonia or 1,6-diaminohexane treatment of the original oxirane carrier. Hydrazine treatment gave lower immobilization yields. The same is true of the azide method independent of the length of the spacer. Most enzyme activity was preserved by coupling the carbodiimide-activated enzyme to the carrier with alkyl or arylamino groups at the end of a longer substituent. Immobilization on diazo-modified carrier gave average results. Rapid immobilization by a lysine-modified phosgene-treated carrier resulted in an activity loss. It is suggested that multipoint and very tight attachment of the enzyme molecule to the matrix decreased the activity. The immobilized activity is quite stable in solution and very stable upon lyophilization with sucrose.

Molecular characterization of a novel bacterial aryl acylamidase belonging to the amidase signature enzyme family.

PubMed

Ko, Hyeok-Jin; Lee, Eun Woo; Bang, Won-Gi; Lee, Cheol-Koo; Kim, Kyoung Heon; Choi, In-Geol

2010-05-01

In seeking aryl acylamidase (EC 3.5.1.13) acting on an amide bond in p-acetaminophenol (Tylenol), we identified a novel gene encoding 496 residues of a protein. The gene revealed a conserved amidase signature region with a canonical catalytic triad. The gene was expressed in E. coli and characterized for its biochemical properties. The optimum pH and temperature for the activity on p-acetaminophenol were 10 and 37 degrees C, respectively. The half-life of enzyme activity at 37 degrees C was 192 h and 90% of its activity remained after 3 h incubation at 40 degrees C. Divalent metals was found to inhibit the activity of enzyme. The K (m) values for various aryl acylamides such as 4-nitroacetanilide, p-acetaminophenol, phenacetin, 4-chloroacetanilide and acetanilide were 0.10, 0.32, 0.83, 1.9 and 19 mM, respectively. The reverse reaction activity (amide synthesis) was also examined using various chain lengths (C(1) approximately C(4) and C(10)) of carboxylic donors and aniline as substrates. These kinetic parameters and substrate specificity in forward and reverse reaction indicated that the aryl acylamidase in this study has a preference for aryl substrate having polar functional groups and hydrophobic carboxylic donors.

Role of Two Cell Wall Amidases in Septal Junction and Nanopore Formation in the Multicellular Cyanobacterium Anabaena sp. PCC 7120

PubMed Central

Bornikoel, Jan; Carrión, Alejandro; Fan, Qing; Flores, Enrique; Forchhammer, Karl; Mariscal, Vicente; Mullineaux, Conrad W.; Perez, Rebeca; Silber, Nadine; Wolk, C. Peter; Maldener, Iris

2017-01-01

Filamentous cyanobacteria have developed a strategy to perform incompatible processes in one filament by differentiating specialized cell types, N2-fixing heterocysts and CO2-fixing, photosynthetic, vegetative cells. These bacteria can be considered true multicellular organisms with cells exchanging metabolites and signaling molecules via septal junctions, involving the SepJ and FraCD proteins. Previously, it was shown that the cell wall lytic N-acetylmuramyl-L-alanine amidase, AmiC2, is essential for cell–cell communication in Nostoc punctiforme. This enzyme perforates the septal peptidoglycan creating an array of nanopores, which may be the framework for septal junction complexes. In Anabaena sp. PCC 7120, two homologs of AmiC2, encoded by amiC1 and amiC2, were identified and investigated in two different studies. Here, we compare the function of both AmiC proteins by characterizing different Anabaena amiC mutants, which was not possible in N. punctiforme, because there the amiC1 gene could not be inactivated. This study shows the different impact of each protein on nanopore array formation, the process of cell–cell communication, septal protein localization, and heterocyst differentiation. Inactivation of either amidase resulted in significant reduction in nanopore count and in the rate of fluorescent tracer exchange between neighboring cells measured by FRAP analysis. In an amiC1 amiC2 double mutant, filament morphology was affected and heterocyst differentiation was abolished. Furthermore, the inactivation of amiC1 influenced SepJ localization and prevented the filament-fragmentation phenotype that is characteristic of sepJ or fraC fraD mutants. Our findings suggest that both amidases are to some extent redundant in their function, and describe a functional relationship of AmiC1 and septal proteins SepJ and FraCD. PMID:28929086

Role of Two Cell Wall Amidases in Septal Junction and Nanopore Formation in the Multicellular Cyanobacterium Anabaena sp. PCC 7120.

PubMed

Bornikoel, Jan; Carrión, Alejandro; Fan, Qing; Flores, Enrique; Forchhammer, Karl; Mariscal, Vicente; Mullineaux, Conrad W; Perez, Rebeca; Silber, Nadine; Wolk, C Peter; Maldener, Iris

2017-01-01

Filamentous cyanobacteria have developed a strategy to perform incompatible processes in one filament by differentiating specialized cell types, N 2 -fixing heterocysts and CO 2 -fixing, photosynthetic, vegetative cells. These bacteria can be considered true multicellular organisms with cells exchanging metabolites and signaling molecules via septal junctions, involving the SepJ and FraCD proteins. Previously, it was shown that the cell wall lytic N -acetylmuramyl-L-alanine amidase, AmiC2, is essential for cell-cell communication in Nostoc punctiforme . This enzyme perforates the septal peptidoglycan creating an array of nanopores, which may be the framework for septal junction complexes. In Anabaena sp. PCC 7120, two homologs of AmiC2, encoded by amiC1 and amiC2 , were identified and investigated in two different studies. Here, we compare the function of both AmiC proteins by characterizing different Anabaena amiC mutants, which was not possible in N. punctiforme , because there the amiC1 gene could not be inactivated. This study shows the different impact of each protein on nanopore array formation, the process of cell-cell communication, septal protein localization, and heterocyst differentiation. Inactivation of either amidase resulted in significant reduction in nanopore count and in the rate of fluorescent tracer exchange between neighboring cells measured by FRAP analysis. In an amiC1 amiC2 double mutant, filament morphology was affected and heterocyst differentiation was abolished. Furthermore, the inactivation of amiC1 influenced SepJ localization and prevented the filament-fragmentation phenotype that is characteristic of sepJ or fraC fraD mutants. Our findings suggest that both amidases are to some extent redundant in their function, and describe a functional relationship of AmiC1 and septal proteins SepJ and FraCD.

Deduced catalytic mechanism of d-amino acid amidase from Ochrobactrum anthropi SV3

PubMed Central

Okazaki, Seiji; Suzuki, Atsuo; Komeda, Hidenobu; Asano, Yasuhisa; Yamane, Takashi

2008-01-01

d-Amino acid amidase (DAA) from Ochrobactrum anthropi SV3 catalyzes d-stereospecific hydrolysis of amino acid amides. DAA has attracted attention as a catalyst for the stereospecific production of d-amino acids, although the mechanism that drives the reaction has not been clear. Previously, the structure of DAA was classified into two types, a substrate-bound state with an ordered Ω loop, and a ground state with a disordered Ω loop. Because the binding of the substrate facilitates ordering, this transition was regarded to be induced fit motion. The angles and distances of hydrogen bonds at Tyr149 Oη, Ser60 Oγ and Lys63 Nζ revealed that Tyr149 Oη donates an H atom to a water molecule in the substrate-bound state, and that Tyr149 Oη donates an H atom to Ser60 Oγ or Lys63 Nζ in the ground state. Taking into consideration the locations of the H atoms of Tyr149 Oη, Ser60 Oγ and Lys63 Nζ, a catalytic mechanism of DAA activity is presented, wherein a shift of an H atom at Tyr149 Oη in the substrate-bound versus the ground state plays a significant role in the reaction. This mechanism explains well why acylation proceeds and deacylation does not proceed in the substrate-bound state. PMID:18421151

Cell wall amidase AmiC1 is required for cellular communication and heterocyst development in the cyanobacterium Anabaena PCC 7120 but not for filament integrity.

PubMed

Berendt, Susanne; Lehner, Josef; Zhang, Yao Vincent; Rasse, Tobias M; Forchhammer, Karl; Maldener, Iris

2012-10-01

Filamentous cyanobacteria of the order Nostocales display typical properties of multicellular organisms. In response to nitrogen starvation, some vegetative cells differentiate into heterocysts, where fixation of N(2) takes place. Heterocysts provide a micro-oxic compartment to protect nitrogenase from the oxygen produced by the vegetative cells. Differentiation involves fundamental remodeling of the gram-negative cell wall by deposition of a thick envelope and by formation of a neck-like structure at the contact site to the vegetative cells. Cell wall-hydrolyzing enzymes, like cell wall amidases, are involved in peptidoglycan maturation and turnover in unicellular bacteria. Recently, we showed that mutation of the amidase homologue amiC2 gene in Nostoc punctiforme ATCC 29133 distorts filament morphology and function. Here, we present the functional characterization of two amiC paralogues from Anabaena sp. strain PCC 7120. The amiC1 (alr0092) mutant was not able to differentiate heterocysts or to grow diazotrophically, whereas the amiC2 (alr0093) mutant did not show an altered phenotype under standard growth conditions. In agreement, fluorescence recovery after photobleaching (FRAP) studies showed a lack of cell-cell communication only in the AmiC1 mutant. Green fluorescent protein (GFP)-tagged AmiC1 was able to complement the mutant phenotype to wild-type properties. The protein localized in the septal regions of newly dividing cells and at the neck region of differentiating heterocysts. Upon nitrogen step-down, no mature heterocysts were developed in spite of ongoing heterocyst-specific gene expression. These results show the dependence of heterocyst development on amidase function and highlight a pivotal but so far underestimated cellular process, the remodeling of peptidoglycan, for the biology of filamentous cyanobacteria.

Synthesis, biological evaluation, and 3D QSAR study of 2-methyl-4-oxo-3-oxetanylcarbamic acid esters as N-acylethanolamine acid amidase (NAAA) inhibitors.

PubMed

Ponzano, Stefano; Berteotti, Anna; Petracca, Rita; Vitale, Romina; Mengatto, Luisa; Bandiera, Tiziano; Cavalli, Andrea; Piomelli, Daniele; Bertozzi, Fabio; Bottegoni, Giovanni

2014-12-11

N-(2-Oxo-3-oxetanyl)carbamic acid esters have recently been reported to be noncompetitive inhibitors of the N-acylethanolamine acid amidase (NAAA) potentially useful for the treatment of pain and inflammation. In the present study, we further explored the structure-activity relationships of the carbamic acid ester side chain of 2-methyl-4-oxo-3-oxetanylcarbamic acid ester derivatives. Additional favorable features in the design of potent NAAA inhibitors have been found together with the identification of a single digit nanomolar inhibitor. In addition, we devised a 3D QSAR using the atomic property field method. The model turned out to be able to account for the structural variability and was prospectively validated by designing, synthesizing, and testing novel inhibitors. The fairly good agreement between predictions and experimental potency values points to this 3D QSAR model as the first example of quantitative structure-activity relationships in the field of NAAA inhibitors.

Pleiotropic Effects of Cell Wall Amidase LytA on Streptococcus pneumoniae Sensitivity to the Host Immune Response

PubMed Central

Ramos-Sevillano, Elisa; Urzainqui, Ana; Campuzano, Susana; Moscoso, Miriam; González-Camacho, Fernando; Domenech, Mirian; Rodríguez de Córdoba, Santiago; Sánchez-Madrid, Francisco; Brown, Jeremy S.; García, Ernesto

2014-01-01

The complement system is a key component of the host immune response for the recognition and clearance of Streptococcus pneumoniae. In this study, we demonstrate that the amidase LytA, the main pneumococcal autolysin, inhibits complement-mediated immunity independently of effects on pneumolysin by a complex process of impaired complement activation, increased binding of complement regulators, and direct degradation of complement C3. The use of human sera depleted of either C1q or factor B confirmed that LytA prevented activation of both the classical and alternative pathways, whereas pneumolysin inhibited only the classical pathway. LytA prevented binding of C1q and the acute-phase protein C-reactive protein to S. pneumoniae, thereby reducing activation of the classical pathway on the bacterial surface. In addition, LytA increased recruitment of the complement downregulators C4BP and factor H to the pneumococcal cell wall and directly cleaved C3b and iC3b to generate degradation products. As a consequence, C3b deposition and phagocytosis increased in the absence of LytA and were markedly enhanced for the lytA ply double mutant, confirming that a combination of LytA and Ply is essential for the establishment of pneumococcal pneumonia and sepsis in a murine model of infection. These data demonstrate that LytA has pleiotropic effects on complement activation, a finding which, in combination with the effects of pneumolysin on complement to assist with pneumococcal complement evasion, confirms a major role of both proteins for the full virulence of the microorganism during septicemia. PMID:25404032

Thin-Layer Polymer Wrapped Enzymes Encapsulated in Hierarchically Mesoporous Silica with High Activity and Enhanced Stability

NASA Astrophysics Data System (ADS)

Zhang, Fang; Wang, Meitao; Liang, Chao; Jiang, Huangyong; Shen, Jian; Li, Hexing

2014-03-01

A novel soft-hard cooperative approach was developed to synthesize bioactive mesoporous composite by pre-wrapping Penicillin G amidase with poly(acrylaimde) nanogel skin and subsequently incorporating such Penicillin G amidase nanocapsules into hierarchically mesoporous silica. The as-received bioactive mesoporous composite exhibited comparable activity and extraordinarily high stability in comparison with native Penicillin G amidase and could be used repetitively in the water-medium hydrolysis of penicillin G potassium salt. Furthermore, this strategy could be extended to the synthesis of multifunctional bioactive mesoporous composite by simultaneously introducing glucose oxidase nanocapsules and horseradish peroxidase nanocapsules into hierarchically mesoporous silica, which demonstrated a synergic effect in one-pot tandem oxidation reaction. Improvements in the catalytic performances were attributed to the combinational unique structure from soft polymer skin and hard inorganic mesoporous silica shell, which cooperatively helped enzyme molecules to retain their appropriate geometry and simultaneously decreased the enzyme-support negative interaction and mass transfer limitation under heterogeneous conditions.

The level of aryl acylamidase activity displayed by human butyrylcholinesterase depends on its molecular distribution.

PubMed

Montenegro, M F; Moral-Naranjo, M T; Páez de la Cadena, M; Campoy, F J; Muñoz-Delgado, E; Vidal, C J

2008-09-25

Butyrylcholinesterase (BuChE) and acetylcholinesterase (AChE) display both esterase and aryl acylamidase (AAA) activities. Their AAA activity can be measured using o-nitroacetanilide (ONA). In human samples depleted of acetylcholinesterase, we noticed that the ratio of amidase to esterase activities varied depending on the source, despite both activities being due to BuChE. Searching for an explanation, we compared the activities of BuChE molecular forms in samples of human colon, kidney and serum, and observed that BuChE monomers (G(1)) hydrolyzed o-nitroacetanilide much faster than tetramers (G(4)). This fact suggested that association might cause differences in the AAA site between single and polymerized subunits. This and other post-translational modifications in BuChE subunits probably determine their level of AAA activity. The higher amidase activity of monomers could justify the presence of single BuChE subunits in cells as a way to preserve the AAA activity of BuChE, which could be lost by oligomerization.

New Enzymatic Method of Chiral Amino Acid Synthesis by Dynamic Kinetic Resolution of Amino Acid Amides: Use of Stereoselective Amino Acid Amidases in the Presence of α-Amino-ɛ-Caprolactam Racemase▿

PubMed Central

Yamaguchi, Shigenori; Komeda, Hidenobu; Asano, Yasuhisa

2007-01-01

d- and l-amino acids were produced from l- and d-amino acid amides by d-aminopeptidase from Ochrobactrum anthropi C1-38 and l-amino acid amidase from Pseudomonas azotoformans IAM 1603, respectively, in the presence of α-amino-ɛ-caprolactam racemase from Achromobacter obae as the catalyst by dynamic kinetic resolution of amino acid amides. PMID:17586677

Effect of mass transfer in a recirculation batch reactor system for immobilized penicillin amidase.

PubMed

Park, J M; Choi, C Y; Seong, B L; Han, M H

1982-10-01

The effect of external mass transfer resistance on the overall reaction rate of the immobilized whole cell penicillin amidase of E. coli in a recirculation batch reactor was investigated. The internal diffusional resistance was found negligible as indicated by the value of effectiveness factor, 0.95. The local environmental change in a column due to the pH drop was successfully overcome by employing buffer solution. The reaction rate was measured by pH-stat method and was found to follow the simple Michaelis-Menten law at the initial stage of the reaction. The values of the net reaction rate experimentally determined were used to calculate the substrate concentration at the external surface of the catalyst pellet and then to calculate the mass transfer coefficient, k(L), at various flow rates and substrate concentrations. The correlation proposed by Chilton and Colburn represented adequately the experimental data. The linear change of log j(D) at low log N(Re) with negative slope was ascribed to the fact that the external mass transfer approached the state of pure diffusion in the limit of zero superficial velocity.

Pseudomonas aeruginosa mutants resistant to urea inhibition of growth on acetanilide.

PubMed

Gregoriou, M; Brown, P R; Tata, R

1977-11-01

Pseudomonas aeruginosa AI 3 was able to grow in medium containing acetanilide (N-phenylacetamide) as a carbon source when NH4+ was the nitrogen source but not when urea was the nitrogen source. AIU mutants isolated from strain AI 3 grew on either medium. Urease levels in bacteria grown in the presence of urea were 10-fold lower when NH4+ or acetanilide was also in the medium, but there were no apparent differences in urease or its synthesis between strain AI 3 and mutant AIU 1N. The first metabolic step in the acetanilide utlization is catalyzed by an amidase. Amidases in several AIU strains showed altered physiochemical properties. Urea inhibited amidase in a time-dependent reaction, but the rates of the inhibitory reaction with amidases from the AIU mutants were slower than with AI 3 amidase. The purified amidase from AIU 1N showed a marked difference in its pH/activity profile from that obtained with purified AI 3 amidase. These observations indicate that the ability of strain AIU 1N and the other mutants to grow on acetanilide/urea medium is associated with a mutation in the amidase structural gene; this was confirmed for strain AIU 1N by transduction.

Pseudomonas aeruginosa mutants resistant to urea inhibition of growth on acetanilide.

PubMed Central

Gregoriou, M; Brown, P R; Tata, R

1977-01-01

Pseudomonas aeruginosa AI 3 was able to grow in medium containing acetanilide (N-phenylacetamide) as a carbon source when NH4+ was the nitrogen source but not when urea was the nitrogen source. AIU mutants isolated from strain AI 3 grew on either medium. Urease levels in bacteria grown in the presence of urea were 10-fold lower when NH4+ or acetanilide was also in the medium, but there were no apparent differences in urease or its synthesis between strain AI 3 and mutant AIU 1N. The first metabolic step in the acetanilide utlization is catalyzed by an amidase. Amidases in several AIU strains showed altered physiochemical properties. Urea inhibited amidase in a time-dependent reaction, but the rates of the inhibitory reaction with amidases from the AIU mutants were slower than with AI 3 amidase. The purified amidase from AIU 1N showed a marked difference in its pH/activity profile from that obtained with purified AI 3 amidase. These observations indicate that the ability of strain AIU 1N and the other mutants to grow on acetanilide/urea medium is associated with a mutation in the amidase structural gene; this was confirmed for strain AIU 1N by transduction. PMID:410788

Key role of amino acid residues in the dimerization and catalytic activation of the autolysin LytA, an important virulence factor in Streptococcus pneumoniae.

PubMed

Romero, Patricia; López, Rubens; García, Ernesto

2007-06-15

LytA, the main autolysin of Streptococcus pneumoniae, was the first member of the bacterial N-acetylmuramoyl-l-alanine amidase (NAM-amidase) family of proteins to be well characterized. This autolysin degrades the peptidoglycan bonds of pneumococcal cell walls after anchoring to the choline residues of the cell wall teichoic acids via its choline-binding module (ChBM). The latter is composed of seven repeats (ChBRs) of approximately 20 amino acid residues. The translation product of the lytA gene is the low-activity E-form of LytA (a monomer), which can be "converted" (activated) in vitro by choline into the fully active C-form at low temperature. The C-form is a homodimer with a boomerang-like shape. To study the structural requirements for the monomer-to-dimer modification and to clarify whether "conversion" is synonymous with dimerization, the biochemical consequences of replacing four key amino acid residues of ChBR6 and ChBR7 (the repeats involved in dimer formation) were determined. The results obtained with a collection of 21 mutated NAM-amidases indicate that Ile-315 is a key amino acid residue in both LytA activity and folding. Amino acids with a marginal position in the solenoid structure of the ChBM were of minor influence in dimer stability; neither the size, polarity, nor aromatic nature of the replacement amino acids affected LytA activity. In contrast, truncated proteins were drastically impaired in their activity and conversion capacity. The results indicate that dimerization and conversion are different processes, but they do not answer the questions of whether conversion can only be achieved after a dimer formation step.

In vitro effect of cell phone radiation on motility, DNA fragmentation and clusterin gene expression in human sperm.

PubMed

Zalata, Adel; El-Samanoudy, Ayman Z; Shaalan, Dalia; El-Baiomy, Youssef; Mostafa, Taymour

2015-01-01

Use of cellular phones emitting radiofrequency electromagnetic field (RF-EMF) has been increased exponentially and become a part of everyday life. This study aimed to investigate the effects of in vitro RF-EMF exposure emitted from cellular phones on sperm motility index, sperm DNA fragmentation and seminal clusterin (CLU) gene expression. In this prospective study, a total of 124 semen samples were grouped into the following main categories: i. normozoospermia (N, n=26), ii. asthenozoospermia (A, n=32), iii. asthenoteratozoospermia (AT, n=31) and iv. oligoasthenoteratozoospermia (OAT, n=35). The same semen samples were then divided into two portions non-exposed and exposed samples to cell phone radiation for 1 hour. Before and immediately after exposure, both aliquots were subjected to different assessments for sperm motility, acrosin activity, sperm DNA fragmentation and CLU gene expression. Statistical differences were analyzed using paired t student test for comparisons between two sub-groups where p<0.05 was set as significant. There was a significant decrease in sperm motility, sperm linear velocity, sperm linearity index, and sperm acrosin activity, whereas there was a significant increase in sperm DNA fragmentation percent, CLU gene expression and CLU protein levels in the exposed semen samples to RF-EMF compared with non-exposed samples in OAT>AT>A>N groups, respectively (p<0.05). Cell phone emissions have a negative impact on exposed sperm motility index, sperm acrosin activity, sperm DNA fragmentation and seminal CLU gene expression, especially in OAT cases.

Activity Augmentation of Amphioxus Peptidoglycan Recognition Protein BbtPGRP3 via Fusion with a Chitin Binding Domain

PubMed Central

Wang, Wen-Jie; Cheng, Wang; Luo, Ming; Yan, Qingyu; Yu, Hong-Mei; Li, Qiong; Cao, Dong-Dong; Huang, Shengfeng; Xu, Anlong; Mariuzza, Roy A.; Chen, Yuxing; Zhou, Cong-Zhao

2015-01-01

Peptidoglycan recognition proteins (PGRPs), which have been identified in most animals, are pattern recognition molecules that involve antimicrobial defense. Resulting from extraordinary expansion of innate immune genes, the amphioxus encodes many PGRPs of diverse functions. For instance, three isoforms of PGRP encoded by Branchiostoma belcheri tsingtauense, termed BbtPGRP1~3, are fused with a chitin binding domain (CBD) at the N-terminus. Here we report the 2.7 Å crystal structure of BbtPGRP3, revealing an overall structure of an N-terminal hevein-like CBD followed by a catalytic PGRP domain. Activity assays combined with site-directed mutagenesis indicated that the individual PGRP domain exhibits amidase activity towards both DAP-type and Lys-type peptidoglycans (PGNs), the former of which is favored. The N-terminal CBD not only has the chitin-binding activity, but also enables BbtPGRP3 to gain a five-fold increase of amidase activity towards the Lys-type PGNs, leading to a significantly broadened substrate spectrum. Together, we propose that modular evolution via domain shuffling combined with gene horizontal transfer makes BbtPGRP1~3 novel PGRPs of augmented catalytic activity and broad recognition spectrum. PMID:26479246

A novel aryl acylamidase from Nocardia farcinica hydrolyses polyamide.

PubMed

Heumann, Sonja; Eberl, Anita; Fischer-Colbrie, Gudrun; Pobeheim, Herbert; Kaufmann, Franz; Ribitsch, Doris; Cavaco-Paulo, Artur; Guebitz, Georg M

2009-03-01

An alkali stable polyamidase was isolated from a new strain of Nocardia farcinica. The enzyme consists of four subunits with a total molecular weight of 190 kDa. The polyamidase cleaved amide and ester bonds of water insoluble model substrates like adipic acid bishexylamide and bis(benzoyloxyethyl)terephthalate and hydrolyzed different soluble amides to the corresponding acid. Treatment of polyamide 6 with this amidase led to an increased hydrophilicity based on rising height and tensiometry measurements and evidence of surface hydrolysis of polyamide 6 is shown. In addition to amidase activity, the enzyme showed activity on p-nitrophenylbutyrate. On hexanoamide the amidase exhibited a K(m) value of 5.5 mM compared to 0.07 mM for p-nitroacetanilide. The polyamidase belongs to the amidase signature family and is closely related to aryl acylamidases from different strains/species of Nocardia and to the 6-aminohexanoate-cyclic dimer hydrolase (EI) from Arthrobacter sp. KI72.

Lipoamidase activity in normal and mutagenized pancreatic cholesterol esterase (bile salt-stimulated lipase).

PubMed Central

Hui, D Y; Hayakawa, K; Oizumi, J

1993-01-01

Purified human milk lipoamidase was digested with endoproteinase Lys-C and the digested peptides were subjected to gasphase microsequence analysis. The sequencing of three isolated peptides of human milk lipoamidase revealed the identity of this protein with human milk bile salt-stimulated lipase (pancreatic cholesterol esterase). The identity of the cholesterol esterase with lipoamidase was confirmed by expressing a recombinant form of rat pancreatic cholesterol esterase and testing for lipoamidase activity of the recombinant protein. The results showed that the recombinant cholesterol esterase displayed both lipolytic and lipoamidase activities and was capable of hydrolysing triacetin and lipoyl-4-aminobenzoate (LPAB). The mechanisms of the esterase and amidase activities of the enzyme were further tested by determining enzyme activity in a mutagenized cholesterol esterase with a His435-->Gln435 substitution. This mutation has been shown previously to abolish enzyme activity against esterase substrates [DiPersio, Fontaine and Hui (1991) J. Biol. Chem. 266, 4033-4036]. We showed that the mutagenized protein was effective in hydrolysing the amidase substrate LPAB and displayed similar enzyme kinetics to those of the native enzyme. These data indicate that the mechanism for the cholesterol esterase hydrolysis of lipoamides is different from that of the hydrolysis of substrates with an ester linkage. The presence of an enzyme in the gastrointestinal tract capable of both ester and amide hydrolysis suggests an important role for this protein in the digestion and absorption processes. PMID:8471055

Improved penicillin amidase production using a genetically engineered mutant of escherichia coli ATCC 11105

DOE Office of Scientific and Technical Information (OSTI.GOV)

Robas, N.; Zouheiry, H.; Branlant, G.

Penicillin G amidase (PGA) is a key enzyme for the industrial production of penicillin G derivatives used in therapeutics. Escherichia coli ATCC 11105 is the more commonly used strain for PGA production. To improve enzyme yield, the authors constructed various recombinant E. coli HB 101 and ATCC 11105 strains. For each strain, PGA production was determined for various concentrations of glucose and phenylacetic acid (PAA) in the medium. The E. coli strain, G271, was identified as the best performer (800 U NIPAB/L). This strain was obtained as follows: an E. coli ATCC 11105 mutant (E. coli G133) was first selectedmore » based on a low negative effect of glucose on PGA production. This mutant was then transformed with a pBR322 derivative containing the PGA gene. Various experiments were made to try to understand the reason for the high productivity of E. coli G271. The host strain, E. coli G133, was found to be mutated in one (or more) gene(s) whose product(s) act(s) in trans on the PGA gene expression. Its growth is not inhibited by high glucose concentration in the medium. Interestingly, whereas glucose still exerts some negative effect on the PGA production by E. coli G133, PGA production by its transformant (E. coli G271) is stimulated by glucose. The reason for this stimulation is discussed. Transformation of E. coli G133 with a pBR322 derivative containing the HindIII fragment of the PGA gene, showed that the performance of E. coli G271 depends both upon the host strain properties and the plasmid structure. Study of the production by the less efficient E. coli HB101 derivatives brought some light on the mechanism of regulation of the PGA gene.« less

Product-induced gene expression, a product-responsive reporter assay used to screen metagenomic libraries for enzyme-encoding genes.

PubMed

Uchiyama, Taku; Miyazaki, Kentaro

2010-11-01

A reporter assay-based screening method for enzymes, which we named product-induced gene expression (PIGEX), was developed and used to screen a metagenomic library for amidases. A benzoate-responsive transcriptional activator, BenR, was placed upstream of the gene encoding green fluorescent protein and used as a sensor. Escherichia coli sensor cells carrying the benR-gfp gene cassette fluoresced in response to benzoate concentrations as low as 10 μM but were completely unresponsive to the substrate benzamide. An E. coli metagenomic library consisting of 96,000 clones was grown in 96-well format in LB medium containing benzamide. The library cells were then cocultivated with sensor cells. Eleven amidase genes were recovered from 143 fluorescent wells; eight of these genes were homologous to known bacterial amidase genes while three were novel genes. In addition to their activity toward benzamide, the enzymes were active toward various substrates, including d- and l-amino acid amides, and displayed enantioselectivity. Thus, we demonstrated that PIGEX is an effective approach for screening novel enzymes based on product detection.

A supramolecular complex between proteinases and beta-cyclodextrin that preserves enzymatic activity: physicochemical characterization.

PubMed

Denadai, Angelo M L; Santoro, Marcelo M; Lopes, Miriam T P; Chenna, Angélica; de Sousa, Frederico B; Avelar, Gabriela M; Gomes, Marco R Túlio; Guzman, Fanny; Salas, Carlos E; Sinisterra, Rubén D

2006-01-01

Cyclodextrins are suitable drug delivery systems because of their ability to subtly modify the physical, chemical, and biological properties of guest molecules through labile interactions by formation of inclusion and/or association complexes. Plant cysteine proteinases from Caricaceae and Bromeliaceae are the subject of therapeutic interest, because of their anti-inflammatory, antitumoral, immunogenic, and wound-healing properties. In this study, we analyzed the association between beta-cyclodextrin (betaCD) and fraction P1G10 containing the bioactive proteinases from Carica candamarcensis, and described the physicochemical nature of the solid-state self-assembled complexes by Fourier transform infrared (FTIR) spectroscopy, thermogravimetry (TG), differential scanning calorimetry (DSC), X-ray powder diffraction (XRD), and nuclear magnetic resonance (NMR), as well as in solution by circular dichroism (CD), isothermal titration calorimetry (ITC), and amidase activity. The physicochemical analyses suggest the formation of a complex between P1G10 and betaCD. Higher secondary interactions, namely hydrophobic interactions, hydrogen bonding and van der Waals forces were observed at higher P1G10 : betaCD mass ratios. These results provide evidence of the occurrence of strong solid-state supramolecular non-covalent interactions between P1G10 and betaCD. Microcalorimetric analysis demonstrates that complexation results in a favorable enthalpic contribution, as has already been described during formation of similar betaCD inclusion compounds. The amidase activity of the complex shows that the enzyme activity is not readily available at 24 hours after dissolution of the complex in aqueous buffer; the proteinase becomes biologically active by the second day and remains stable until day 16, when a gradual decrease occurs, with basal activity attained by day 29. The reported results underscore the potential for betaCDs as candidates for complexing cysteine proteinases, resulting

Identification of bovine sperm acrosomal proteins that interact with a 32-kDa acrosomal matrix protein.

PubMed

Nagdas, Subir K; Smith, Linda; Medina-Ortiz, Ilza; Hernandez-Encarnacion, Luisa; Raychoudhury, Samir

2016-03-01

Mammalian fertilization is accomplished by the interaction between sperm and egg. Previous studies from this laboratory have identified a stable acrosomal matrix assembly from the bovine sperm acrosome termed the outer acrosomal membrane-matrix complex (OMC). This stable matrix assembly exhibits precise binding activity for acrosin and N-acetylglucosaminidase. A highly purified OMC fraction comprises three major (54, 50, and 45 kDa) and several minor (38-19 kDa) polypeptides. The set of minor polypeptides (38-19 kDa) termed "OMCrpf polypeptides" is selectively solubilized by high-pH extraction (pH 10.5), while the three major polypeptides (55, 50, and 45 kDa) remain insoluble. Proteomic identification of the OMC32 polypeptide (32 kDa polypeptide isolated from high-pH soluble fraction of OMC) yielded two peptides that matched the NCBI database sequence of acrosin-binding protein. Anti-OMC32 recognized an antigenically related family of polypeptides (OMCrpf polypeptides) in the 38-19-kDa range with isoelectric points ranging between 4.0 and 5.1. Other than glycohydrolases, OMC32 may also be complexed to other acrosomal proteins. The present study was undertaken to identify and localize the OMC32 binding polypeptides and to elucidate the potential role of the acrosomal protein complex in sperm function. OMC32 affinity chromatography of a detergent-soluble fraction of bovine cauda sperm acrosome followed by mass spectrometry-based identification of bound proteins identified acrosin, lactadherin, SPACA3, and IZUMO1. Co-immunoprecipitation analysis also demonstrated the interaction of OMC32 with acrosin, lactadherin, SPACA3, and IZUMO1. Our immunofluorescence studies revealed the presence of SPACA3 and lactadherin over the apical segment, whereas IZUMO1 is localized over the equatorial segment of Triton X-100 permeabilized cauda sperm. Immunoblot analysis showed that a significant portion of SPACA3 was released after the lysophosphatidylcholine (LPC)-induced acrosome

In vitro bactericidal and bacteriolytic activity of ceragenin CSA-13 against planktonic cultures and biofilms of Streptococcus pneumoniae and other pathogenic streptococci.

PubMed

Moscoso, Miriam; Esteban-Torres, María; Menéndez, Margarita; García, Ernesto

2014-01-01

Ceragenin CSA-13, a cationic steroid, is here reported to show a concentration-dependent bactericidal/bacteriolytic activity against pathogenic streptococci, including multidrug-resistant Streptococcus pneumoniae. The autolysis promoted by CSA-13 in pneumococcal cultures appears to be due to the triggering of the major S. pneumoniae autolysin LytA, an N-acetylmuramoyl-L-alanine amidase. CSA-13 also disintegrated pneumococcal biofilms in a very efficient manner, although at concentrations slightly higher than those required for bactericidal activity on planktonic bacteria. CSA-13 has little hemolytic activity which should allow testing its antibacterial efficacy in animal models.

Use of biochemical markers to evaluate the quality of fresh and cryopreserved semen from the arctic fox (Vulpes lagopus).

PubMed

Stasiak, K; Glogowski, J; Demianowicz, W; Kowalski, R; Nowak-Tkaczyk, A; Janicki, B

2014-01-01

The aim of this study was to use biochemical markers to evaluate the quality of fresh and cryopreserved semen from the arctic fox (Vulpes lagopus). Twenty-three manually collected ejaculates were analysed for the main indicators of semen quality (sperm concentration and ejaculate volume). Sperm motility and percentage of morphologically normal and abnormal spermatozoa were determined according to the stage of cryopreservation (fresh--measurement A; equilibrated--measurement B; frozen/thawed--measurement C). Furthermore, the seminal plasma and supernatants were analysed after equilibration and freeze/thawing for the activity of the enzymes alkaline phosphatase (ALP), acid phosphatase (AcP), lactate dehydrogenase (LDH) and aspartate aminotransferase (AspAT), and for the activity of acrosin inhibitors (AP). The mean concentration of sperm was 625.1 million/cm3, and ejaculate volume averaged 1.6 cm3. Seminal plasma was characterized by the highest activity of alkaline phosphatase (3.43 x 10(3) U/l) and lowest activity of acrosin inhibitors (4.55 x 10(3) U/l). After equilibration, the supernatants showed the highest activity of acid phosphatase (94.9 U/l) and after freeze-thawing, they showed a high activity of lactate dehydrogenase (535.8 U/l) and aspartate aminotransferase (577.1 U/l), which indicates that these proteins had leaked from spermatozoa into the extracellular medium during the biotechnique of semen cryopreservation. In addition, several significant relationships were found between some indicators of semen quality and plasma and/or supernatant enzyme activity.

SPINK2 deficiency causes infertility by inducing sperm defects in heterozygotes and azoospermia in homozygotes.

PubMed

Kherraf, Zine-Eddine; Christou-Kent, Marie; Karaouzene, Thomas; Amiri-Yekta, Amir; Martinez, Guillaume; Vargas, Alexandra S; Lambert, Emeline; Borel, Christelle; Dorphin, Béatrice; Aknin-Seifer, Isabelle; Mitchell, Michael J; Metzler-Guillemain, Catherine; Escoffier, Jessica; Nef, Serge; Grepillat, Mariane; Thierry-Mieg, Nicolas; Satre, Véronique; Bailly, Marc; Boitrelle, Florence; Pernet-Gallay, Karin; Hennebicq, Sylviane; Fauré, Julien; Bottari, Serge P; Coutton, Charles; Ray, Pierre F; Arnoult, Christophe

2017-08-01

Azoospermia, characterized by the absence of spermatozoa in the ejaculate, is a common cause of male infertility with a poorly characterized etiology. Exome sequencing analysis of two azoospermic brothers allowed the identification of a homozygous splice mutation in SPINK2, encoding a serine protease inhibitor believed to target acrosin, the main sperm acrosomal protease. In accord with these findings, we observed that homozygous Spink2 KO male mice had azoospermia. Moreover, despite normal fertility, heterozygous male mice had a high rate of morphologically abnormal spermatozoa and a reduced sperm motility. Further analysis demonstrated that in the absence of Spink2, protease-induced stress initiates Golgi fragmentation and prevents acrosome biogenesis leading to spermatid differentiation arrest. We also observed a deleterious effect of acrosin overexpression in HEK cells, effect that was alleviated by SPINK2 coexpression confirming its role as acrosin inhibitor. These results demonstrate that SPINK2 is necessary to neutralize proteases during their cellular transit toward the acrosome and that its deficiency induces a pathological continuum ranging from oligoasthenoteratozoospermia in heterozygotes to azoospermia in homozygotes. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

Ascorbic acid, catalase and chlorpromazine reduce cryopreservation-induced damages to crossbred bull spermatozoa.

PubMed

Paudel, K P; Kumar, S; Meur, S K; Kumaresan, A

2010-04-01

The present study evaluated the effectiveness of ascorbic acid, catalase, chlorpromazine and their combinations in reducing the cryodamages to crossbred bull (Bos taurus x Bos indicus) spermatozoa. A total of 32 ejaculates (eight each from four bulls) were diluted in Tris-citric acid-fructose-egg yolk-glycerol extender. Each ejaculate was split into six parts (five treatment and one control). Treatment groups included 10 mm ascorbic acid, 0.1 mm chlorpromazine, 200 IU/ml catalase, 10 mm ascorbic acid + 0.1 mm chlorpromazine or 200 IU/ml catalase + 0.1 mm chlorpromazine in the extender. Fluorescent probes (Fluorescein isothiocyanate--Pisum sativum agglutinin + Propidium iodide) were used for the assessment of spermatozoa viability and acrosomal status. The proportion of acrosome intact live (AIL), acrosome intact dead, acrosome reacted live and acrosome reacted dead sperm was assessed in fresh, equilibrated and frozen-thawed semen. The functional status of the sperm was assessed using hypo-osmotic sperm swelling test (HOSST). Activities of acrosin and hyaluronidase enzyme were also determined. Lipid peroxidation level was assayed based on the melonaldehyde (MDA) production. In cryopreserved semen, the values of AIL spermatozoa, HOSST response, hyaluronidase and acrosin activity were reduced by 53%, 47%, 34% and 54%, respectively from their initial values in fresh semen. However, MDA level was threefold higher in the frozen-thawed sperm compared with fresh sperm. Significant (p < 0.05) improvement in motility, viability, HOSST response, retention of hyaluonidase and acrosin and reduction in MDA was recorded in ascorbic acid, catalase, ascorbic acid + chlorpromazine and catalase + chlorpromazine incorporated groups. The percentage of AIL sperm was significantly (p < 0.05) higher in ascorbic acid, catalase and ascorbic acid + chlorpromazine incorporated groups compared with the control. Chlorpromazine alone did not improve the post-thaw semen quality but when combined

Detection of sperm-reactive antibodies in wild sika deer and identification of the sperm antigens.

PubMed

Kawase, Osamu; Jimbo, Mitsuru

2018-03-16

Antisperm antibodies potentially inhibit sperm functions causing the sterility in humans and experimentally treated animals. However, there is no information about antisperm antibodies emerging spontaneously in wildlife. In this study, we searched for the sperm-reactive antibodies, spontaneously produced in wild sika deer (Cervus nippon), and identified the sperm antigens. We collected 529 fecal masses of sika deer in Japanese cities, from which we extracted the mucosal antibodies to test them for reactivities to deer sperm proteins by ELISA. Two of the extracts contained IgAs that were highly reactive to the sperm proteins. The molecular weights of the active IgAs, partially purified by DEAE-sephadex A-50, were estimated at more than 100 kDa, suggesting that the IgAs evaded drastic digestion in the gastrointestinal tract. Two-dimensional electrophoresis and immunoblotting detected three major antigens, and the following LC-MS/MS analysis identified them as alpha-enolase, phosphoglycerate kinase 2 and acrosin-binding protein. The antibodies were cross-reactive to a recombinant human acrosin-binding protein. To our knowledge, this is the first research to find that the sperm-reactive antibodies are produced spontaneously in wildlife and they recognize a common antigen found in humans.

Hydrolysis of acetylthiocoline, o-nitroacetanilide and o-nitrotrifluoroacetanilide by fetal bovine serum acetylcholinesterase.

PubMed

Montenegro, María F; Moral-Naranjo, María T; Muñoz-Delgado, Encarnación; Campoy, Francisco J; Vidal, Cecilio J

2009-04-01

Besides esterase activity, acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) hydrolyze o-nitroacetanilides through aryl acylamidase activity. We have reported that BuChE tetramers and monomers of human blood plasma differ in o-nitroacetanilide (ONA) hydrolysis. The homology in quaternary structure and folding of subunits in the prevalent BuChE species (G4(H)) of human plasma and AChE forms of fetal bovine serum prompted us to study the esterase and amidase activities of fetal bovine serum AChE. The k(cat)/K(m) values for acetylthiocholine (ATCh), ONA and its trifluoro derivative N-(2-nitrophenyl)-trifluoroacetamide (F-ONA) were 398 x 10(6) M(-1) min(-1), 0.8 x 10(6) M(-1) min(-1), and 17.5 x 10(6) M(-1) min(-1), respectively. The lack of inhibition of amidase activity at high F-ONA concentrations makes it unlikely that there is a role for the peripheral anionic site (PAS) in F-ONA degradation, but the inhibition of ATCh, ONA and F-ONA hydrolysis by the PAS ligand fasciculin-2 points to the transit of o-nitroacetalinides near the PAS on their way to the active site. Sedimentation analysis confirmed substrate hydrolysis by tetrameric 10.9S AChE. As compared with esterase activity, amidase activity was less sensitive to guanidine hydrochloride. This reagent led to the formation of 9.3S tetramers with partially unfolded subunits. Their capacity to hydrolyze ATCh and F-ONA revealed that, despite the conformational change, the active site architecture and functionality of AChE were partially retained.

Subcellular Localization of Rice Leaf Aryl Acylamidase Activity 1

PubMed Central

Gaynor, John J.; Still, Cecil C.

1983-01-01

The intracellular localization of aryl acylamidase (aryl-acylamide amidohydrolase, EC 3.5.1.13) in rice (Oryza sativa L. var Starbonnet) leaves was investigated. The enzyme hydrolyzes and detoxifies the herbicide propanil (3,4-dichloropropionanilide) thereby accounting for immunity of the rice plant to herbicidal action. Fractionation of mesophyll protoplasts by differential centrifugation yielded the highest specific activity of amidase in the crude mitochondrial fraction. Further separation of density gradients of the silica sol Percoll also indicated that this enzyme was mitochondrial. By the use of biochemical markers, the purified mitochondrial fraction was shown to be substantially free of contamination from nuclei, chloroplasts, golgi, and plasma membranes. Subfractionation of the purified mitochondria suggests that this enzyme is located on the outer membrane. PMID:16662987

Liquid nitrogen vapor is comparable to liquid nitrogen for storage of cryopreserved human sperm: evidence from the characteristics of post-thaw human sperm.

PubMed

Hu, Jingmei; Zhao, Shidou; Xu, Chengyan; Zhang, Lin; Lu, Shaoming; Cui, Linlin; Ma, Jinlong; Chen, Zi-Jiang

2015-11-01

To compare the differences in the characteristics of post-thaw human sperm after storage in either liquid nitrogen (LN2; -196 °C) or LN2 vapor (-167 °C). Experimental study. University hospital. Thirty healthy volunteers who agreed to donate their normal semen samples for infertility or research were included in the study. Semen samples (n = 30) were divided into eight aliquots and frozen. Four aliquots of each human semen sample were stored in LN2 (-196 °C), and the other four aliquots were stored in LN2 vapor (-167 °C). After 1, 3, 6, or 12 months, samples were thawed and analyzed. The motility was evaluated by the manual counting method. The viability was estimated by eosin staining. The morphology was analyzed by Diff-Quik staining. The sperm DNA integrity was determined with acridine orange fluorescent staining, and acrosin activity was assayed by the modified Kennedy method. The characteristics of post-thaw human sperm, including motility, viability, morphology, DNA integrity, and acrosin activity, showed no significant difference between LN2 and LN2 vapor storage for the different time periods. LN2 vapor was comparable to LN2 in post-thaw sperm characteristics, suggesting that LN2 vapor may be substituted for LN2 for the long-term storage of human sperm. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

A new acylamidase from Rhodococcus erythropolis TA37 can hydrolyze N-substituted amides.

PubMed

Lavrov, K V; Zalunin, I A; Kotlova, E K; Yanenko, A S

2010-08-01

A new acylamidase was isolated from Rhodococcus erythropolis TA37 and characterized. N-Substituted acrylamides (isopropyl acrylamide, N,N-dimethyl-aminopropyl acrylamide, and methylene-bis-acrylamide), acid para-nitroanilides (4'-nitroacetanilide, Gly-pNA, Ala-pNA, Leu-pNA), and N-acetyl derivatives of glycine, alanine, and leucine are good substrates for this enzyme. Aliphatic amides (acetamide, acrylamide, isobutyramide, n-butyramide, and valeramide) are also used as substrates but with less efficiency. The enzyme subunit mass by SDS-PAGE is 55 kDa. Maximal activity is exhibited at pH 7-8 and 55°C. The enzyme is stable for 15 h at 22°C and for 0.5 h at 45°C. The Michaelis constant (K(m)) is 0.25 mM with Gly-pNA and 0.55 mM with Ala-pNA. The acylamidase activity is suppressed by inhibitors of serine proteases (phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate) but is not suppressed by inhibitors of aliphatic amidases (acetaldehyde and nitrophenyl disulfides). The N-terminal amino acid sequence of the acylamidase is highly homologous to those of two putative amidases detected from sequenced R. erythropolis genomes. It is suggested that the acylamidase together with the detected homologs forms a new class within the amidase signature family.

EFFECT OF CRYOPRESERVATION AND THEOPHYLLINE ON MOTILITY CHARACTERISTICS OF LAKE STURGEON (ACIPENSER FULVESCENS) SPERMATOZOA

EPA Science Inventory

Computer-assisted motility analysis (CASA) was used to evaluate the effect of cryopreservation and theophylline treatment on sperm motility of lake sturgeon (Acipenser fulvescens).Motility was recorded at 0 and 5 min postactivation.The effect of cryopreservation on sperm acrosin-...

A chimeric LysK-lysostaphin fusion enzyme lysing Staphylococcus aureus cells: a study of both kinetics of inactivation and specifics of interaction with anionic polymers

USDA-ARS?s Scientific Manuscript database

A staphylolytic fusion protein (K-L) was created, harboring three unique lytic activities comprised of the LysK CHAP endopeptidase, and amidase domains, and the lysostaphin glycyl-glycine endopeptidase domain. To assess the potential of possible therapeutic applications, the kinetic behavior of K-L...

Second-Generation Non-Covalent NAAA Inhibitors are Protective in a Model of Multiple Sclerosis.

PubMed

Migliore, Marco; Pontis, Silvia; Fuentes de Arriba, Angel Luis; Realini, Natalia; Torrente, Esther; Armirotti, Andrea; Romeo, Elisa; Di Martino, Simona; Russo, Debora; Pizzirani, Daniela; Summa, Maria; Lanfranco, Massimiliano; Ottonello, Giuliana; Busquet, Perrine; Jung, Kwang-Mook; Garcia-Guzman, Miguel; Heim, Roger; Scarpelli, Rita; Piomelli, Daniele

2016-09-05

Palmitoylethanolamide (PEA) and oleoylethanolamide (OEA) are endogenous lipid mediators that suppress inflammation. Their actions are terminated by the intracellular cysteine amidase, N-acylethanolamine acid amidase (NAAA). Even though NAAA may offer a new target for anti-inflammatory therapy, the lipid-like structures and reactive warheads of current NAAA inhibitors limit the use of these agents as oral drugs. A series of novel benzothiazole-piperazine derivatives that inhibit NAAA in a potent and selective manner by a non-covalent mechanism are described. A prototype member of this class (8) displays high oral bioavailability, access to the central nervous system (CNS), and strong activity in a mouse model of multiple sclerosis (MS). This compound exemplifies a second generation of non-covalent NAAA inhibitors that may be useful in the treatment of MS and other chronic CNS disorders. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Secreted Immunomodulatory Proteins of Staphylococcus aureus Activate Platelets and Induce Platelet Aggregation.

PubMed

Binsker, Ulrike; Palankar, Raghavendra; Wesche, Jan; Kohler, Thomas P; Prucha, Josephine; Burchhardt, Gerhard; Rohde, Manfred; Schmidt, Frank; Bröker, Barbara M; Mamat, Uwe; Pané-Farré, Jan; Graf, Anica; Ebner, Patrick; Greinacher, Andreas; Hammerschmidt, Sven

2018-04-01

Staphylococcus aureus can cause bloodstream infections associated with infective endocarditis (IE) and disseminated intravascular coagulopathy (DIC). Both complications involve platelets. In view of an increasing number of antibiotic-resistant strains, new approaches to control systemic S. aureus infection are gaining importance. Using a repertoire of 52 recombinant S. aureus proteins in flow cytometry-based platelet activation and aggregation assays, we identified, in addition to the extracellular adherence protein Eap, three secreted staphylococcal proteins as novel platelet activating proteins. Eap and the chemotaxis inhibitory protein of S. aureus (CHIPS), the formyl peptide receptor-like 1 inhibitory protein (FLIPr) and the major autolysin Atl induced P-selectin expression in washed platelets and platelet-rich plasma. Similarly, AtlA, CHIPS and Eap induced platelet aggregation in whole blood. Fluorescence microscopy illustrated that P-selectin expression is associated with calcium mobilization and re-organization of the platelet actin cytoskeleton. Characterization of the functionally active domains of the major autolysin AtlA and Eap indicates that the amidase domain of Atl and the tandem repeats 3 and 4 of Eap are crucial for platelet activation. These results provide new insights in S. aureus protein interactions with platelets and identify secreted proteins as potential treatment targets in case of antibiotic-resistant S. aureus infection. Schattauer GmbH Stuttgart.

Molecular and functional characterization of peptidoglycan-recognition protein SC2 (PGRP-SC2) from Nile tilapia (Oreochromis niloticus) involved in the immune response to Streptococcus agalactiae.

PubMed

Gan, Zhen; Chen, Shannan; Hou, Jing; Huo, Huijun; Zhang, Xiaolin; Ruan, Baiye; Laghari, Zubair Ahmed; Li, Li; Lu, Yishan; Nie, Pin

2016-07-01

PGRP-SC2, the member of PGRP family, plays an important role in regulation of innate immune response. In this paper, a PGRP-SC2 gene of Nile tilapia, Oreochromis niloticus (designated as On-PGRP-SC2) was cloned and its expression pattern under the infection of Streptococcus agalactiae was investigated. Sequence analysis showed main structural features required for amidase activity were detected in the deduced amino acid sequence of On-PGRP-SC2. In healthy tilapia, the On-PGRP-SC2 transcripts could be detected in all the examined tissues, with the most abundant expression in the muscle. When infected with S. agalactiae, there was a clear time-dependent expression pattern of On-PGRP-SC2 in the spleen, head kidney and brain. The assays for the amidase activity suggested that recombinant On-PGRP-SC2 protein had a Zn(2+)-dependent PGN-degrading activity. Moreover, our works showed that recombinant On-PGRP-SC2 protein could significantly reduce bacterial load in target organs attacked by S. agalactiae. These findings indicated that On-PGRP-SC2 may play important roles in the immune response to S. agalactiae in Nile tilapia. Copyright © 2016 Elsevier Ltd. All rights reserved.

Staphylococcus haemolyticus prophage ΦSH2 endolysin relies on Cysteine, Histidine-dependent Amidohydrolases/Peptidases activity for lysis ‘from without’

PubMed Central

Schmelcher, Mathias; Korobova, Olga; Schischkova, Nina; Kiseleva, Natalia; Kopylov, Paul; Pryamchuk, Sergey; Donovan, David M.; Abaev, Igor

2014-01-01

Staphylococcus aureus is an important pathogen, with methicillin-resistant (MRSA) and multi-drug resistant strains becoming increasingly prevalent in both human and veterinary clinics. S. aureus causing bovine mastitis yields high annual losses to the dairy industry. Conventional treatment of mastitis by broad range antibiotics is often not successful and may contribute to development of antibiotic resistance. Bacteriophage endolysins present a promising new source of antimicrobials. The endolysin of prophage ΦSH2 of Staphylococcus haemolyticus strain JCSC1435 (ΦSH2 lysin) is a peptidoglycan hydrolase consisting of two catalytic domains (CHAP and amidase) and an SH3b cell wall binding domain. In this work, we demonstrated its lytic activity against live staphylococcal cells and investigated the contribution of each functional module to bacterial lysis by testing a series of deletion constructs in zymograms and turbidity reduction assays. The CHAP domain exhibited three-fold higher activity than the full length protein and optimum activity in physiological saline. This activity was further enhanced by the presence of bivalent calcium ions. The SH3b domain was shown to be required for full activity of the complete ΦSH2 lysin. The full length enzyme and the CHAP domain showed activity against multiple staphylococcal strains, including MRSA strains, mastitis isolates, and CoNS. PMID:23026556

Characterization of an Indole-3-Acetamide Hydrolase from Alcaligenes faecalis subsp. parafaecalis and Its Application in Efficient Preparation of Both Enantiomers of Chiral Building Block 2,3-Dihydro-1,4-Benzodioxin-2-Carboxylic Acid.

PubMed

Mishra, Pradeep; Kaur, Suneet; Sharma, Amar Nath; Jolly, Ravinder S

2016-01-01

Both the enantiomers of 2,3-dihydro-1,4-benzodioxin-2-carboxylic acid are valuable chiral synthons for enantiospecific synthesis of therapeutic agents such as (S)-doxazosin mesylate, WB 4101, MKC 242, 2,3-dihydro-2-hydroxymethyl-1,4-benzodioxin, and N-[2,4-oxo-1,3-thiazolidin-3-yl]-2,3-dihydro-1,4-benzodioxin-2-carboxamide. Pharmaceutical applications require these enantiomers in optically pure form. However, currently available methods suffer from one drawback or other, such as low efficiency, uncommon and not so easily accessible chiral resolving agent and less than optimal enantiomeric purity. Our interest in finding a biocatalyst for efficient production of enantiomerically pure 2,3-dihydro-1,4-benzodioxin-2-carboxylic acid lead us to discover an amidase activity from Alcaligenes faecalis subsp. parafaecalis, which was able to kinetically resolve 2,3-dihydro-1,4-benzodioxin-2-carboxyamide with E value of >200. Thus, at about 50% conversion, (R)-2,3-dihydro-1,4-benzodioxin-2-carboxylic acid was produced in >99% e.e. The remaining amide had (S)-configuration and 99% e.e. The amide and acid were easily separated by aqueous (alkaline)-organic two phase extraction method. The same amidase was able to catalyse, albeit at much lower rate the hydrolysis of (S)-amide to (S)-acid without loss of e.e. The amidase activity was identified as indole-3-acetamide hydrolase (IaaH). IaaH is known to catalyse conversion of indole-3-acetamide (IAM) to indole-3-acetic acid (IAA), which is phytohormone of auxin class and is widespread among plants and bacteria that inhabit plant rhizosphere. IaaH exhibited high activity for 2,3-dihydro-1,4-benzodioxin-2-carboxamide, which was about 65% compared to its natural substrate, indole-3-acetamide. The natural substrate for IaaH indole-3-acetamide shared, at least in part a similar bicyclic structure with 2,3-dihydro-1,4-benzodioxin-2-carboxamide, which may account for high activity of enzyme towards this un-natural substrate. To the best of

Staphylococcus haemolyticus prophage ΦSH2 endolysin relies on cysteine, histidine-dependent amidohydrolases/peptidases activity for lysis 'from without'.

PubMed

Schmelcher, Mathias; Korobova, Olga; Schischkova, Nina; Kiseleva, Natalia; Kopylov, Paul; Pryamchuk, Sergey; Donovan, David M; Abaev, Igor

2012-12-31

Staphylococcus aureus is an important pathogen, with methicillin-resistant (MRSA) and multi-drug resistant strains becoming increasingly prevalent in both human and veterinary clinics. S. aureus causing bovine mastitis yields high annual losses to the dairy industry. Conventional treatment of mastitis by broad range antibiotics is often not successful and may contribute to development of antibiotic resistance. Bacteriophage endolysins present a promising new source of antimicrobials. The endolysin of prophage ΦSH2 of Staphylococcus haemolyticus strain JCSC1435 (ΦSH2 lysin) is a peptidoglycan hydrolase consisting of two catalytic domains (CHAP and amidase) and an SH3b cell wall binding domain. In this work, we demonstrated its lytic activity against live staphylococcal cells and investigated the contribution of each functional module to bacterial lysis by testing a series of deletion constructs in zymograms and turbidity reduction assays. The CHAP domain exhibited three-fold higher activity than the full length protein and optimum activity in physiological saline. This activity was further enhanced by the presence of bivalent calcium ions. The SH3b domain was shown to be required for full activity of the complete ΦSH2 lysin. The full length enzyme and the CHAP domain showed activity against multiple staphylococcal strains, including MRSA strains, mastitis isolates, and CoNS. Published by Elsevier B.V.

Discovery and Biochemical Characterization of PlyP56, PlyN74, and PlyTB40—Bacillus Specific Endolysins

PubMed Central

Etobayeva, Irina; Linden, Sara B.; Alem, Farhang; Rizkalla, Lucas; Temple, Louise; Hakami, Ramin M.

2018-01-01

Three Bacillus bacteriophage-derived endolysins, designated PlyP56, PlyN74, and PlyTB40, were identified, cloned, purified, and characterized for their antimicrobial properties. Sequence alignment reveals these endolysins have an N-terminal enzymatically active domain (EAD) linked to a C-terminal cell wall binding domain (CBD). PlyP56 has a Peptidase_M15_4/VanY superfamily EAD with a conserved metal binding motif and displays biological dependence on divalent ions for activity. In contrast, PlyN74 and PlyTB40 have T7 lysozyme-type Amidase_2 and carboxypeptidase T-type Amidase_3 EADs, respectively, which are members of the MurNAc-LAA superfamily, but are not homologs and thus do not have a shared protein fold. All three endolysins contain similar SH3-family CBDs. Although minor host range differences were noted, all three endolysins show relatively broad antimicrobial activity against members of the Bacillus cereus sensu lato group with the highest lytic activity against B. cereus ATCC 4342. Characterization studies determined the optimal lytic activity for these enzymes was at physiological pH (pH 7.0–8.0), over a broad temperature range (4–55 °C), and at low concentrations of NaCl (<50 mM). Direct comparison of lytic activity shows the PlyP56 enzyme to be twice as effective at lysing the cell wall peptidoglycan as PlyN74 or PlyTB40, suggesting PlyP56 is a good candidate for further antimicrobial development as well as bioengineering studies. PMID:29883383

Toxic effects of Pb2+ entering sperm through Ca2+ channels in the freshwater crab Sinopotamon henanense.

PubMed

Li, Na; Xu, Peng; Jing, Wei-Xin; Hwang, Jiang-Shiou; Wang, Lan

2017-11-01

Lead (Pb) is a heavy metal that can damage animal sperm. To study the effects of Pb on calcium homeostasis and calcium channel in the sperm of freshwater crab Sinopotamon henanense, the induction of acrosome reaction (AR) and acrosin activity were investigated when crabs were exposed to different Pb concentrations (0, 3.675, 7.35, 14.7, 29.4 and 58.8mg/L) for 3, 5 and 7 d separately. Fluorescent probe Fluo-3/AM was loaded into the sperm, and [Ca 2+ ] in the sperm was measured by fluorescence microscopy and using microplate reader. The calmodulin (CaM) concentration was measured by ELISA method. Verapamil (VRP), a calcium channel blocker, was used to evaluate whether Pb can enter the sperm through calcium channels leading to sperm damage. After sperm were exposed at 50μg/L VRP, 100μg/L Pb, 50μg/L VRP+100μg/L Pb, 1000μg/L Pb and 50μg/L VRP+1000μg/L Pb for 1h in vitro，sperm quality parameters (sperm survival and sperm DNA integrity) and levels of parameters indicating oxidative stress (protein carbonylation [PCO] and malondialdehyde [MDA]) were measured. Our data showed that Pb reduced the induction of acrosome reaction (AR), down-regulated the acrosin activity, decreased the intracellular concentration of Ca 2+ and elevated CaM concentration. Compared to controls, Pb alone induced significant stress, as reflected by decreasing sperm survival and sperm DNA integrity, and increasing PCO and MDA contents. In the presence of VRP, 100μg/L Pb-induced stresses were reduced, all the measured parameters in the sperm exposed at 100μg/L Pb returned to control levels. Our results indicate that Pb enters the sperm of the crab S. henanense through calcium channels, the inhibition of which blocks Pb-induced stresses such as sperm quality decline and oxidative damage. Copyright © 2017 Elsevier B.V. All rights reserved.

Characterization of Enterococcus faecium bacteriophage IME-EFm5 and its endolysin LysEFm5.

PubMed

Gong, Pengjuan; Cheng, Mengjun; Li, Xinwei; Jiang, Haiyan; Yu, Chuang; Kahaer, Nadire; Li, Juecheng; Zhang, Lei; Xia, Feifei; Hu, Liyuan; Sun, Changjiang; Feng, Xin; Lei, Liancheng; Han, Wenyu; Gu, Jingmin

2016-05-01

Due to the worldwide prevalence of antibiotic resistant strains, phages therapy has been revitalized recently. In this study, an Enterococcus faecium phage named IME-EFm5 was isolated from hospital sewage. Whole genomic sequence analysis demonstrated that IME-EFm5 belong to the Siphoviridae family, and has a double-stranded genome of 42,265bp (with a 35.51% G+C content) which contains 70 putative coding sequences. LysEFm5, the endolysin of IME-EFm5, contains an amidase domain in its N-terminal and has a wider bactericidal spectrum than its parental phage IME-EFm5, including 7 strains of vancomycin-resistant E. faecium. The mutagenesis analysis revealed that the zinc ion binding residues (H27, H132, and C140), E90, and T138 are required for the catalysis of LysEFm5. However, the antibacterial activity of LysEFm5 is zinc ion independent, which is inconsistent with most of other amidase members. The phage lysin LysEFm5 might be an alternative treatment strategy for infections caused by multidrug-resistant E. faecium. Copyright © 2016 Elsevier Inc. All rights reserved.

Ligand-Binding Properties and Conformational Dynamics of Autolysin Repeat Domains in Staphylococcal Cell Wall Recognition

PubMed Central

Zoll, Sebastian; Schlag, Martin; Shkumatov, Alexander V.; Rautenberg, Maren; Svergun, Dmitri I.; Götz, Friedrich

2012-01-01

The bifunctional major autolysin Atl plays a key role in staphylococcal cell separation. Processing of Atl yields catalytically active amidase (AM) and glucosaminidase (GL) domains that are each fused to repeating units. The two repeats of AM (R1 and R2) target the enzyme to the septum, where it cleaves murein between dividing cells. We have determined the crystal structure of R2, which reveals that each repeat folds into two half-open β-barrel subunits. We further demonstrate that lipoteichoic acid serves as a receptor for the repeats and that this interaction depends on conserved surfaces in each subunit. Small-angle X-ray scattering of the mature amidase reveals the presence of flexible linkers separating the AM, R1, and R2 units. Different levels of flexibility for each linker provide mechanistic insights into the conformational dynamics of the full-length protein and the roles of its components in cell wall association and catalysis. Our analysis supports a model in which the repeats direct the catalytic AM domain to the septum, where it can optimally perform the final step of cell division. PMID:22609916

Aryl acylamidase activity exhibited by butyrylcholinesterase is higher in chick than in horse, but much lower than in fetal calf serum.

PubMed

Weitnauer, E; Robitzki, A; Layer, P G

1998-10-02

Several side activities have been attributed to butyrylcholinesterase (BChE), including aryl acylamidase (AAA) activity, which is an amidase-like activity with unknown physiological function splitting the artificial substrate o-nitroacetanilide. For avians, extensive developmental data have pointed to neurogenetic functions of BChE, however, a possible AAA activity of BChE has not been studied. In this study, we first compare the relative levels of AAA exhibited by BChE in whole sera from chick, fetal calves (FCS) and horse. Remarkably, FCS exhibits a 400-fold higher ratio of AAA/BChE than horse and 80-fold higher than chick serum. We then show that an immunoisolated preparation of BChE from chicken serum presents significant activity for AAA. Both in sera and with the purified enzyme, the AAA activity is fully inhibited by anticholinesterase drugs, showing that AAA activity is exclusively conveyed by the BChE molecule. Noticeably, AAA inhibition even occurs at lower drug concentrations than that of BChE activity itself. Moreover, AAA is sensitive to serotonin. These data indicate that (1) AAA is a general feature of serum BChE of vertebrates including avians, (2) AAA is effectively inhibited by cholinergic and serotonergic agents, and (3) AAA may have a developmental role, since it is much pronounced in a serum from fetal animals. Functionally, deamination of neuropeptides, a link between cholinergic and serotonergic neurotransmitter systems, and roles in lipoprotein metabolism could be relevant.

METABOLIC ENGINEERING TO DEVELOP A PATHWAY FOR THE SELECTIVE CLEAVAGE OF CARBON-NITROGEN BONDS

DOE Office of Scientific and Technical Information (OSTI.GOV)

John J. Kilbane III

The objective of the project is to develop biochemical pathways for the selective cleavage of C-N bonds in molecules found in petroleum. The initial phase of the project will focus on the isolation or development of an enzyme capable of cleaving the C-N bond in aromatic amides, specifically 2-aminobiphenyl. The objective of the second phase of the research will be to construct a biochemical pathway for the selective removal of nitrogen from carbazole by combining the carA genes from Sphingomonas sp. GTIN11 with the gene(s) encoding an appropriate amidase. The objective of the final phase of the project will bemore » to develop derivative CN bond cleaving enzymes that have broader substrate ranges and to demonstrate the use of such strains to selectively remove nitrogen from petroleum. The project is on schedule and no major difficulties have been encountered. During the first year of the project (October, 2002-September, 2003) enrichment culture experiments have resulted in the isolation of promising cultures that may be capable of cleaving C-N bonds in aromatic amides, several amidase genes have been cloned and are currently undergoing directed evolution to obtain derivatives that can cleave C-N bonds in aromatic amides, and the carA genes from Sphingomonas sp. GTIN11, and Pseudomonas resinovorans CA10 were cloned in vectors capable of replicating in Escherichia coli. Future research will address expression of these genes in Rhodococcus erythropolis. Enrichment culture experiments and directed evolution experiments continue to be a main focus of research activity and further work is required to obtain an appropriate amidase that will selectively cleave C-N bonds in aromatic substrates. Once an appropriate amidase gene is obtained it must be combined with genes encoding an enzyme capable of converting carbazole to 2'aminobiphenyl-2,3-diol: specifically carA genes. The carA genes from two sources have been cloned and are ready for construction of C-N bond cleavage

The Enzymology of 2-Hydroxyglutarate, 2-Hydroxyglutaramate and 2-Hydroxysuccinamate and Their Relationship to Oncometabolites

PubMed Central

Hariharan, Vivek A.; Denton, Travis T.; Paraszcszak, Sarah; McEvoy, Kyle; Jeitner, Thomas M.; Krasnikov, Boris F.; Cooper, Arthur J. L.

2017-01-01

Many enzymes make “mistakes”. Consequently, repair enzymes have evolved to correct these mistakes. For example, lactate dehydrogenase (LDH) and mitochondrial malate dehydrogenase (mMDH) slowly catalyze the reduction of 2-oxoglutarate (2-OG) to the oncometabolite l-2-hydroxyglutarate (l-2-HG). l-2-HG dehydrogenase corrects this error by converting l-2-HG to 2-OG. LDH also catalyzes the reduction of the oxo group of 2-oxoglutaramate (2-OGM; transamination product of l-glutamine). We show here that human glutamine synthetase (GS) catalyzes the amidation of the terminal carboxyl of both the l- and d- isomers of 2-HG. The reaction of 2-OGM with LDH and the reaction of l-2-HG with GS generate l-2-hydroxyglutaramate (l-2-HGM). We also show that l-2-HGM is a substrate of human ω-amidase. The product (l-2-HG) can then be converted to 2-OG by l-2-HG dehydrogenase. Previous work showed that 2-oxosuccinamate (2-OSM; transamination product of l-asparagine) is an excellent substrate of LDH. Finally, we also show that human ω-amidase converts the product of this reaction (i.e., l-2-hydroxysuccinamate; l-2-HSM) to l-malate. Thus, ω-amidase may act together with hydroxyglutarate dehydrogenases to repair certain “mistakes” of GS and LDH. The present findings suggest that non-productive pathways for nitrogen metabolism occur in mammalian tissues in vivo. Perturbations of these pathways may contribute to symptoms associated with hydroxyglutaric acidurias and to tumor progression. Finally, methods for the synthesis of l-2-HGM and l-2-HSM are described that should be useful in determining the roles of ω-amidase/4- and 5-C compounds in photorespiration in plants. PMID:28358347

The Enzymology of 2-Hydroxyglutarate, 2-Hydroxyglutaramate and 2-Hydroxysuccinamate and Their Relationship to Oncometabolites.

PubMed

Hariharan, Vivek A; Denton, Travis T; Paraszcszak, Sarah; McEvoy, Kyle; Jeitner, Thomas M; Krasnikov, Boris F; Cooper, Arthur J L

2017-03-30

Many enzymes make "mistakes". Consequently, repair enzymes have evolved to correct these mistakes. For example, lactate dehydrogenase (LDH) and mitochondrial malate dehydrogenase (mMDH) slowly catalyze the reduction of 2-oxoglutarate (2-OG) to the oncometabolite l-2-hydroxyglutarate (l-2-HG). l-2-HG dehydrogenase corrects this error by converting l-2-HG to 2-OG. LDH also catalyzes the reduction of the oxo group of 2-oxoglutaramate (2-OGM; transamination product of l-glutamine). We show here that human glutamine synthetase (GS) catalyzes the amidation of the terminal carboxyl of both the l- and d- isomers of 2-HG. The reaction of 2-OGM with LDH and the reaction of l-2-HG with GS generate l-2-hydroxyglutaramate (l-2-HGM). We also show that l-2-HGM is a substrate of human ω-amidase. The product (l-2-HG) can then be converted to 2-OG by l-2-HG dehydrogenase. Previous work showed that 2-oxosuccinamate (2-OSM; transamination product of l-asparagine) is an excellent substrate of LDH. Finally, we also show that human ω-amidase converts the product of this reaction (i.e., l-2-hydroxysuccinamate; l-2-HSM) to l-malate. Thus, ω-amidase may act together with hydroxyglutarate dehydrogenases to repair certain "mistakes" of GS and LDH. The present findings suggest that non-productive pathways for nitrogen metabolism occur in mammalian tissues in vivo. Perturbations of these pathways may contribute to symptoms associated with hydroxyglutaric acidurias and to tumor progression. Finally, methods for the synthesis of l-2-HGM and l-2-HSM are described that should be useful in determining the roles of ω-amidase/4- and 5-C compounds in photorespiration in plants.

Functional characterization of a short peptidoglycan recognition protein, PGRP5 in grass carp Ctenopharyngodon idella.

PubMed

Li, Jun Hua; Chang, Ming Xian; Xue, Na Na; Nie, P

2013-08-01

Peptidoglycan recognition proteins (PGRPs), which are evolutionarily conserved from insects to mammals, recognize bacterial peptidoglycan (PGN) and function in antibacterial innate immunity. In this study, a short-form PGRP, designated as gcPGRP5 was identified from grass carp Ctenopharyngodon idella. The deduced amino acid sequence of gcPGRP5 is composed of 180 residues with a conserved PGRP domain at the C-terminus. The gcPGRP5 gene consists of four exons and three introns, spacing approximately 2.3 kb in genomic sequence. Phylogenetic analysis demonstrated that the gcPGRP5 is clustered with other PGRP-S identified in teleost fish. The gcPGRP5 is constitutively expressed in all organs/tissues examined, and its expression was significantly induced in CIK cells treated with lipoteichoic acid (LTA), polyinosinic polycytidylic acid (Poly I:C) and PGN. Fluorescence analysis showed that gcPGRP5 is distributed in cytoplasm of CIK cells, and cell lysates from CIK cells transfected with pTurbo-gcPGRP5-GFP and ptGFP1-gcPGRP5 plasmids display the binding activity and peptidoglycan-lytic amidase activity toward Lys-PGN from Staphylococcus aureus and Dap-PGN from Bacillus subtilis. Furthermore, heat-shock protein70 (Hsp70), and MyD88, an adaptor molecule in Toll-like receptor pathway, had an increased expression in CIK cells overexpressed with gcPGRP5. It is thus indicated that gcPGRP5 exhibits amidase activity, and also possesses roles in anti-stress, and in Toll-like receptor signaling pathway. Copyright © 2013 Elsevier Ltd. All rights reserved.

The structural basis of the difference in sensitivity for PNGase F in the de-N-glycosylation of the native bovine pancreatic ribonucleases B and BS.

PubMed

Blanchard, Véronique; Frank, Martin; Leeflang, Bas R; Boelens, Rolf; Kamerling, Johannis P

2008-03-18

In glycoanalysis protocols, N-glycans from glycoproteins are most frequently released with peptide- N (4)-( N-acetyl-beta-glucosaminyl)asparagine amidase F (PNGase F). As the enzyme is an amidase, it cleaves the NH-CO linkage between the Asn side chain and the Asn-bound GlcNAc residue. Usually, the enzyme has a low activity, or is not active at all, on native glycoproteins. A typical example is native bovine pancreatic ribonuclease B (RNase B) with oligomannose-type N-glycans at Asn-34. However, native RNase BS, generated by subtilisin digestion of native RNase B, which comprises amino acid residues 21-124 of RNase B, is sensitive to PNGase F digestion. The same holds for carboxymethylated RNase B (RNase B (cm)). In this study, NMR spectroscopy and molecular modeling have been used to explain the differences in PNGase F activity for native RNase B, native RNase BS, and RNase B (cm). NMR analysis combined with literature data clearly indicated that the N-glycan at Asn-34 is more mobile in RNase BS than in RNase B. MD simulations showed that the region around Asn-34 in RNase B is not very flexible, whereby the alpha-helix of the amino acid residues 1-20 has a stabilizing effect. In RNase BS, the alpha-helix formed by amino acid residues 23-32 is significantly more flexible. Using these data, the possibilities for complex formation of both RNase B and RNase BS with PNGase F were studied, and a model for the RNase BS-PNGase F complex is proposed.

Studies on the membrane integrity of human sperm treated with a new injectable male contraceptive.

PubMed

Chaudhury, K; Bhattacharyya, A K; Guha, S K

2004-08-01

The aim of this study was to evaluate the integrity of sperm surface characteristics in the presence of a new male contraceptive, RISUG [1 mg styrene maleic anhydride (SMA)/100 microl dimethylsulphoxide (DMSO) in 1 ml sperm solution]. Progressively motile human sperm were treated in vitro with RISUG. The cells were analysed for the release of 5'-nucleotidase (5'-NT) (a plasma membrane marker) using 3 mmol/l 5'-AMP and 3 mmol/l beta-glycerophosphate as substrates. Hyaluronidase (an acrosomal membrane marker) was analysed using hyaluronic acid as a substrate. The contents of free and total acrosin, and % proacrosin (all acrosome markers) were assayed using 0.5 mmol/l alpha-N-benzoyl-L-arginine ethylester (BAEE). RISUG caused almost complete disintegration of the plasma membrane leading to significant (P < 0.0001) release of 5'-NT into the surrounding media. Complete dissolution of the acrosome with concomitant vesiculation of the membrane system, as judged from the loss of hyaluronidase, was observed. Total acrosin content in the sperm was also reduced to almost 10%, and proacrosin dropped to 13.2% in the presence of RISUG in comparison to 90.2% in control (P < 0.0001), indicating dispersion of acrosomal contents. Under in vitro conditions, RISUG, at a concentration of 1 mg SMA dissolved in 100 microl of DMSO, caused significant damage to the acrosome and its contents, indicating loss of functional ability of sperm. Copyright 2004 European Society of Human Reproduction and Embryology

Remodeling of the plasma membrane in preparation for sperm–egg recognition: roles of acrosomal proteins

PubMed Central

Tanphaichitr, Nongnuj; Kongmanas, Kessiri; Kruevaisayawan, Hathairat; Saewu, Arpornrad; Sugeng, Clarissa; Fernandes, Jason; Souda, Puneet; Angel, Jonathan B; Faull, Kym F; Aitken, R John; Whitelegge, Julian; Hardy, Daniel; Berger, Trish; Baker, Mark

2015-01-01

The interaction of sperm with the egg's extracellular matrix, the zona pellucida (ZP) is the first step of the union between male and female gametes. The molecular mechanisms of this process have been studied for the past six decades with the results obtained being both interesting and confusing. In this article, we describe our recent work, which attempts to address two lines of questions from previous studies. First, because there are numerous ZP binding proteins reported by various researchers, how do these proteins act together in sperm–ZP interaction? Second, why do a number of acrosomal proteins have ZP affinity? Are they involved mainly in the initial sperm–ZP binding or rather in anchoring acrosome reacting/reacted spermatozoa to the ZP? Our studies reveal that a number of ZP binding proteins and chaperones, extracted from the anterior sperm head plasma membrane, coexist as high molecular weight (HMW) complexes, and that these complexes in capacitated spermatozoa have preferential ability to bind to the ZP. Zonadhesin (ZAN), known as an acrosomal protein with ZP affinity, is one of these proteins in the HMW complexes. Immunoprecipitation indicates that ZAN interacts with other acrosomal proteins, proacrosin/acrosin and sp32 (ACRBP), also present in the HMW complexes. Immunodetection of ZAN and proacrosin/acrosin on spermatozoa further indicates that both proteins traffic to the sperm head surface during capacitation where the sperm acrosomal matrix is still intact, and therefore they are likely involved in the initial sperm–ZP binding step. PMID:25994642

Contribution of the Staphylococcus aureus Atl AM and GL murein hydrolase activities in cell division, autolysis, and biofilm formation.

PubMed

Bose, Jeffrey L; Lehman, McKenzie K; Fey, Paul D; Bayles, Kenneth W

2012-01-01

The most prominent murein hydrolase of Staphylococcus aureus, AtlA, is a bifunctional enzyme that undergoes proteolytic cleavage to yield two catalytically active proteins, an amidase (AM) and a glucosaminidase (GL). Although the bifunctional nature of AtlA has long been recognized, most studies have focused on the combined functions of this protein in cell wall metabolism and biofilm development. In this study, we generated mutant derivatives of the clinical S. aureus isolate, UAMS-1, in which one or both of the AM and GL domains of AtlA have been deleted. Examination of these strains revealed that each mutant exhibited growth rates comparable to the parental strain, but showed clumping phenotypes and lysis profiles that were distinct from the parental strain and each other, suggesting distinct roles in cell wall metabolism. Given the known function of autolysis in the release of genomic DNA for use as a biofilm matrix molecule, we also tested the mutants in biofilm assays and found both AM and GL necessary for biofilm development. Furthermore, the use of enzymatically inactive point mutations revealed that both AM and GL must be catalytically active for S. aureus to form a biofilm. The results of this study provide insight into the relative contributions of AM and GL in S. aureus and demonstrate the contribution of Atl-mediated lysis in biofilm development.

A Novel Hydrolase Identified by Genomic-Proteomic Analysis of Phenylurea Herbicide Mineralization by Variovorax sp. Strain SRS16▿†

PubMed Central

Bers, Karolien; Leroy, Baptiste; Breugelmans, Philip; Albers, Pieter; Lavigne, Rob; Sørensen, Sebastian R.; Aamand, Jens; De Mot, René; Wattiez, Ruddy; Springael, Dirk

2011-01-01

The soil bacterial isolate Variovorax sp. strain SRS16 mineralizes the phenylurea herbicide linuron. The proposed pathway initiates with hydrolysis of linuron to 3,4-dichloroaniline (DCA) and N,O-dimethylhydroxylamine, followed by conversion of DCA to Krebs cycle intermediates. Differential proteomic analysis showed a linuron-dependent upregulation of several enzymes that fit into this pathway, including an amidase (LibA), a multicomponent chloroaniline dioxygenase, and enzymes associated with a modified chlorocatechol ortho-cleavage pathway. Purified LibA is a monomeric linuron hydrolase of ∼55 kDa with a Km and a Vmax for linuron of 5.8 μM and 0.16 nmol min−1, respectively. This novel member of the amidase signature family is unrelated to phenylurea-hydrolyzing enzymes from Gram-positive bacteria and lacks activity toward other tested phenylurea herbicides. Orthologues of libA are present in all other tested linuron-degrading Variovorax strains with the exception of Variovorax strains WDL1 and PBS-H4, suggesting divergent evolution of the linuron catabolic pathway in different Variovorax strains. The organization of the linuron degradation genes identified in the draft SRS16 genome sequence indicates that gene patchwork assembly is at the origin of the pathway. Transcription analysis suggests that a catabolic intermediate, rather than linuron itself, acts as effector in activation of the pathway. Our study provides the first report on the genetic organization of a bacterial pathway for complete mineralization of a phenylurea herbicide and the first report on a linuron hydrolase in Gram-negative bacteria. PMID:22003008

2-Pentadecyl-2-Oxazoline, the Oxazoline of Pea, Modulates Carrageenan-Induced Acute Inflammation

PubMed Central

Petrosino, Stefania; Campolo, Michela; Impellizzeri, Daniela; Paterniti, Irene; Allarà, Marco; Gugliandolo, Enrico; D’Amico, Ramona; Siracusa, Rosalba; Cordaro, Marika; Esposito, Emanuela; Di Marzo, Vincenzo; Cuzzocrea, Salvatore

2017-01-01

N-acylethanolamines (NAEs) involve a family of lipid molecules existent in animal and plant, with N-palmitoylethanolamide (PEA) that arouses great attention owing to its anti-inflammatory, analgesic and neuroprotective activities. Because PEA is produced on demand and exerts pleiotropic effects, the modulation of specific amidases for NAEs (and in particular NAE-hydrolyzing acid amidase NAAA, which is more selective for PEA) could be a condition to preserve its levels. Here we investigate the effect of 2-Pentadecyl-2-oxazoline (PEA-OXA) the oxazoline of PEA, on human recombinant NAAA in vitro and in an established model of Carrageenan (CAR)-induced rat paw inflammation. PEA-OXA dose-dependently significantly inhibited recombinant NAAA and, orally administered to rats (10 mg/kg), limiting histological damage, thermal hyperalgesia and the increase of infiltrating inflammatory cells after CAR injection in the rat right hindpaw, compared to ultramicronized PEA given orally at the same dose (10 mg/kg). These effects were accompanied by elevation of paw PEA levels. Moreover, PEA-OXA markedly reduced neutrophil infiltration and pro-inflammatory cytokine release and prevented CAR-induced IκB-α degradation, nuclear translocation of NF-κB p65, the increase of inducible nitric oxide synthase, cyclooxygenase-2, intercellular adhesion molecule-1, and mast cell activation. Experiments in PPAR-α knockout mice showed that the anti-inflammatory effects of PEA-OXA were not dependent on the presence of PPAR-α receptors. In conclusion, NAAA modulators as PEA-OXA could help to maximize the tissue availability of PEA by increasing its levels and anti-inflammatory effects. PMID:28611664

Correlations among assays of porcine semen quality following cryopreservation.

PubMed

Hammitt, D G; Martin, P A

1989-09-01

Correlations between in vitro tests of semen quality, used to predict the in vivo fertilizing potential of sperm, indicate that the tests may substitute for each other in predicting fertilizing potential. Lack of correlation between tests suggest that both tests should be used to estimate the fertilizing potential. The purpose of this study was to establish correlations between several in vitro tests of porcine semen quality following freezing. Tests of motility with and without caffeine, spermatozoa with normal apical ridges, sephadex filtration with and without caffeine and acrosin activity were all correlated with each other. Correlations among these tests ranged from 0.45 to 0.83 (P<0.05). Assays for glutamic oxalacetic transaninase (GOT) were not consistently correlated with other tests. None of these tests of semen quality were correlated with the sperm penetration assay except for the test of motility without caffeine, which was correlated with the number of penetrations per hamster oocyte (r = 0.71, P<0.05).

[Cloning of new acylamidase gene from Rhodococcus erythropolis and its expression in Escherichia coli].

PubMed

Lavrov, K V; Ianenko, A S

2013-10-01

The gene for new Rhodococcus erythropolis TA37 acylamidase, which possesses unique substrate specificity, has been cloned and expressed in E. coli. Substrates for this enzyme are not only simple amides, such as acetamide and propionamide, but also N-substituted amides, such as 4'-nitroacetanilide. The 1431-bp gene was expressed in E. coli BL21 (DE3) cells on pET16b plasmid under the control of a promoter of the φ 10 gene from the T7 phage. The molecular mass of recombinant acylamidase in E. coli was 55 kDa, which corresponded to that of native acylamidase from Rhodococcus erythropolis TA37. Recombinant acylamidase was able to hydrolize N-substituted amides. A search of a nucleotide database and multiple alignment revealed that acylamidase belonged to the Amidase protein family PF01425, but its nucleotide and amino acid sequences differed significantly from those of the described amidases.

LysK CHAP endopeptidase domain is required for lysis of live staphylococcal cells.

USDA-ARS?s Scientific Manuscript database

LysK is a staphylococcal bacteriophage endolysin composed of three domains, an N-terminal cysteine, histidine-dependent amidohydrolases/peptidases (CHAP) endopeptidase domain (cleaves between D-alanine of the stem peptide and glycine of the cross-bridge peptide) a mid-protein amidase 2 domain (N-ace...

Transferred interbacterial antagonism genes augment eukaryotic innate immune function.

PubMed

Chou, Seemay; Daugherty, Matthew D; Peterson, S Brook; Biboy, Jacob; Yang, Youyun; Jutras, Brandon L; Fritz-Laylin, Lillian K; Ferrin, Michael A; Harding, Brittany N; Jacobs-Wagner, Christine; Yang, X Frank; Vollmer, Waldemar; Malik, Harmit S; Mougous, Joseph D

2015-02-05

Horizontal gene transfer allows organisms to rapidly acquire adaptive traits. Although documented instances of horizontal gene transfer from bacteria to eukaryotes remain rare, bacteria represent a rich source of new functions potentially available for co-option. One benefit that genes of bacterial origin could provide to eukaryotes is the capacity to produce antibacterials, which have evolved in prokaryotes as the result of eons of interbacterial competition. The type VI secretion amidase effector (Tae) proteins are potent bacteriocidal enzymes that degrade the cell wall when delivered into competing bacterial cells by the type VI secretion system. Here we show that tae genes have been transferred to eukaryotes on at least six occasions, and that the resulting domesticated amidase effector (dae) genes have been preserved for hundreds of millions of years through purifying selection. We show that the dae genes acquired eukaryotic secretion signals, are expressed within recipient organisms, and encode active antibacterial toxins that possess substrate specificity matching extant Tae proteins of the same lineage. Finally, we show that a dae gene in the deer tick Ixodes scapularis limits proliferation of Borrelia burgdorferi, the aetiologic agent of Lyme disease. Our work demonstrates that a family of horizontally acquired toxins honed to mediate interbacterial antagonism confers previously undescribed antibacterial capacity to eukaryotes. We speculate that the selective pressure imposed by competition between bacteria has produced a reservoir of genes encoding diverse antimicrobial functions that are tailored for co-option by eukaryotic innate immune systems.

Shared strategies for β-lactam catabolism in the soil microbiome.

PubMed

Crofts, Terence S; Wang, Bin; Spivak, Aaron; Gianoulis, Tara A; Forsberg, Kevin J; Gibson, Molly K; Johnsky, Lauren A; Broomall, Stacey M; Rosenzweig, C Nicole; Skowronski, Evan W; Gibbons, Henry S; Sommer, Morten O A; Dantas, Gautam

2018-06-01

The soil microbiome can produce, resist, or degrade antibiotics and even catabolize them. While resistance genes are widely distributed in the soil, there is a dearth of knowledge concerning antibiotic catabolism. Here we describe a pathway for penicillin catabolism in four isolates. Genomic and transcriptomic sequencing revealed β-lactamase, amidase, and phenylacetic acid catabolon upregulation. Knocking out part of the phenylacetic acid catabolon or an apparent penicillin utilization operon (put) resulted in loss of penicillin catabolism in one isolate. A hydrolase from the put operon was found to degrade in vitro benzylpenicilloic acid, the β-lactamase penicillin product. To test the generality of this strategy, an Escherichia coli strain was engineered to co-express a β-lactamase and a penicillin amidase or the put operon, enabling it to grow using penicillin or benzylpenicilloic acid, respectively. Elucidation of additional pathways may allow bioremediation of antibiotic-contaminated soils and discovery of antibiotic-remodeling enzymes with industrial utility.

Structural and biochemical studies on Vibrio cholerae Hsp31 reveals a novel dimeric form and Glutathione-independent Glyoxalase activity

PubMed Central

Dey, Sanjay

2017-01-01

Vibrio cholerae experiences a highly hostile environment at human intestine which triggers the induction of various heat shock genes. The hchA gene product of V. cholerae O395, referred to a hypothetical intracellular protease/amidase VcHsp31, is one such stress-inducible homodimeric protein. Our current study demonstrates that VcHsp31 is endowed with molecular chaperone, amidopeptidase and robust methylglyoxalase activities. Through site directed mutagenesis coupled with biochemical assays on VcHsp31, we have confirmed the role of residues in the vicinity of the active site towards amidopeptidase and methylglyoxalase activities. VcHsp31 suppresses the aggregation of insulin in vitro in a dose dependent manner. Through crystal structures of VcHsp31 and its mutants, grown at various temperatures, we demonstrate that VcHsp31 acquires two (Type-I and Type-II) dimeric forms. Type-I dimer is similar to EcHsp31 where two VcHsp31 monomers associate in eclipsed manner through several intersubunit hydrogen bonds involving their P-domains. Type-II dimer is a novel dimeric organization, where some of the intersubunit hydrogen bonds are abrogated and each monomer swings out in the opposite directions centering at their P-domains, like twisting of wet cloth. Normal mode analysis (NMA) of Type-I dimer shows similar movement of the individual monomers. Upon swinging, a dimeric surface of ~400Å2, mostly hydrophobic in nature, is uncovered which might bind partially unfolded protein substrates. We propose that, in solution, VcHsp31 remains as an equilibrium mixture of both the dimers. With increase in temperature, transformation to Type-II form having more exposed hydrophobic surface, occurs progressively accounting for the temperature dependent increase of chaperone activity of VcHsp31. PMID:28235098

PGRP-LB homolog acts as a negative modulator of immunity in maintaining the gut-microbe symbiosis of red palm weevil, Rhynchophorus ferrugineus Olivier.

PubMed

Dawadi, Bishnu; Wang, Xinghong; Xiao, Rong; Muhammad, Abrar; Hou, Youming; Shi, Zhanghong

2018-09-01

Many notorious insect pests live in the symbiotic associations with gut microbiota. However, the mechanisms underlying how they host their gut microbiota are unknown. Most gut bacteria can release peptidoglycan (PGN) which is an important antigen to activate the immune response. Therefore, how to keep the appropriate gut immune intensity to host commensals while to efficiently remove enteropathogens is vital for insect health. This study is aimed at elucidating the roles of an amidase PGRP, Rf PGRP-LB, in maintaining the gut-microbe symbiosis of Red palm weevil (RPW), Rhynchophorus ferrugineus Olivier. RfPGRP-LB is a secreted protein containing a typical PGRP domain. The existence of five conservative amino acid residues, being required for amidase activity, showed that RfPGRP-LB is a catalytic protein. Expression analysis revealed abundance of RfPGRP-LB transcripts in gut was dramatically higher than those in other tissues. RfPGRP-LB could be significantly induced against the infection of Escherichia coli. In vitro assays revealed that rRfPGRP-LB impaired the growth of E. coli and agglutinated bacteria cells obviously, suggesting RfPGRP-LB is a pathogen recognition receptor and bactericidal molecule. RfPGRP-LB knockdown reduced the persistence of E. coli in gut and load of indigenous gut microbiota significantly. Furthermore, the community structure of indigenous gut microbiota was also intensively altered by RfPGRP-LB silence. Higher levels of the antimicrobial peptide, attacin, were detected in guts of RfPGRP-LB silenced larvae than controls. Collectively, RfPGRP-LB plays multiple roles in modulating the homeostasis of RPW gut microbiota not only by acting as a negative regulator of mucosal immunity through PGN degradation but also as a bactericidal effector to prevent overgrowth of commensals and persistence of noncommensals. Copyright © 2018 Elsevier Ltd. All rights reserved.

Nitrile Metabolizing Yeasts

NASA Astrophysics Data System (ADS)

Bhalla, Tek Chand; Sharma, Monica; Sharma, Nitya Nand

Nitriles and amides are widely distributed in the biotic and abiotic components of our ecosystem. Nitrile form an important group of organic compounds which find their applications in the synthesis of a large number of compounds used as/in pharmaceutical, cosmetics, plastics, dyes, etc>. Nitriles are mainly hydro-lyzed to corresponding amide/acid in organic chemistry. Industrial and agricultural activities have also lead to release of nitriles and amides into the environment and some of them pose threat to human health. Biocatalysis and biotransformations are increasingly replacing chemical routes of synthesis in organic chemistry as a part of ‘green chemistry’. Nitrile metabolizing organisms or enzymes thus has assumed greater significance in all these years to convert nitriles to amides/ acids. The nitrile metabolizing enzymes are widely present in bacteria, fungi and yeasts. Yeasts metabolize nitriles through nitrilase and/or nitrile hydratase and amidase enzymes. Only few yeasts have been reported to possess aldoxime dehydratase. More than sixty nitrile metabolizing yeast strains have been hither to isolated from cyanide treatment bioreactor, fermented foods and soil. Most of the yeasts contain nitrile hydratase-amidase system for metabolizing nitriles. Transformations of nitriles to amides/acids have been carried out with free and immobilized yeast cells. The nitrilases of Torulopsis candida>and Exophiala oligosperma>R1 are enantioselec-tive and regiospecific respectively. Geotrichum>sp. JR1 grows in the presence of 2M acetonitrile and may have potential for application in bioremediation of nitrile contaminated soil/water. The nitrilase of E. oligosperma>R1 being active at low pH (3-6) has shown promise for the hydroxy acids. Immobilized yeast cells hydrolyze some additional nitriles in comparison to free cells. It is expected that more focus in future will be on purification, characterization, cloning, expression and immobilization of nitrile metabolizing

[Human drug metabolizing enzymes. II. Conjugation enzymes].

PubMed

Vereczkey, L; Jemnitz, K; Gregus, Z

1998-09-01

In this review we focus on human conjugation enzymes (UDP-glucuronyltransferases, methyl-trasferases, N-acetyl-transferases, O-acetyl-transferases, Amidases/carboxyesterases, sulfotransferases, Glutation-S-transferases and the enzymes involved in the conjugation with amino acids) that participate in the metabolism of xenobiotics. Although conjugation reactions in most of the cases result in detoxication, more and more publications prove that the reactions catalysed by these enzymes very often lead to activated molecules that may attack macromolecules (proteins, RNAs, DNAs), resulting in toxicity (liver, neuro-, embryotoxicity, allergy, carcinogenecity). We have summarised the data available on these enzymes concerning their catalytic profile and specificity, inhibition, induction properties, their possible role in the generation of toxic compounds, their importance in clinical practice and drug development.

Influence of chronic oral intake of cannabis extract on oxidative and hydrolytic metabolism of xenobiotics in rat.

PubMed

Khanna, P; Gupta, M B; Gupta, G P; Sanwal, G G; Ali, B

1991-01-01

Dietary intake of petroleum ether extract of cannabis leaves by rats in doses of 158, 250 and 500 mg/kg in the first, second and third week, respectively, caused selective induction of hepatic microsomal carboxylesterases/amidases without affecting the renal hydrolytic activity. Acetanilide N-deacetylase, p-nitrophenylacetate (NPA) esterase and acetylsalicylic acid (ASA) esterase I and II (active at pH 5.5 and 7.4) were stimulated 125, 64, 82 and 60%, respectively, whereas the activities of procaine esterase and acetylaminofluorene (AAF) N-deacetylase remained unaltered. The hydrolysis of acetylcholine was also unchanged. Upon withdrawal of treatment microsomal hydrolytic activity receded to basal levels within 7 days. Curiously though, the two-fold induction of thiacetazone N-deacetylase (118%), a cytosolic hydrolase, remained largely undiminished (62%). An appraisal of the hepatic cytochrome P450 mediated oxidative metabolism revealed approximately three-fold induction of aromatic hydrocarbon hydroxylase (AHH) metabolizing benzo(a)pyrene whereas the N-demethylation of aminopyrene was unaffected. These activities were restored to normal when resin administration was discontinued.

Saliva Microbiota Carry Caries-Specific Functional Gene Signatures

PubMed Central

Chang, Xingzhi; Yuan, Xiao; Tu, Qichao; Yuan, Tong; Deng, Ye; Hemme, Christopher L.; Van Nostrand, Joy; Cui, Xinping; He, Zhili; Chen, Zhenggang; Guo, Dawei; Yu, Jiangbo; Zhang, Yue; Zhou, Jizhong; Xu, Jian

2014-01-01

Human saliva microbiota is phylogenetically divergent among host individuals yet their roles in health and disease are poorly appreciated. We employed a microbial functional gene microarray, HuMiChip 1.0, to reconstruct the global functional profiles of human saliva microbiota from ten healthy and ten caries-active adults. Saliva microbiota in the pilot population featured a vast diversity of functional genes. No significant distinction in gene number or diversity indices was observed between healthy and caries-active microbiota. However, co-presence network analysis of functional genes revealed that caries-active microbiota was more divergent in non-core genes than healthy microbiota, despite both groups exhibited a similar degree of conservation at their respective core genes. Furthermore, functional gene structure of saliva microbiota could potentially distinguish caries-active patients from healthy hosts. Microbial functions such as Diaminopimelate epimerase, Prephenate dehydrogenase, Pyruvate-formate lyase and N-acetylmuramoyl-L-alanine amidase were significantly linked to caries. Therefore, saliva microbiota carried disease-associated functional signatures, which could be potentially exploited for caries diagnosis. PMID:24533043

Saliva microbiota carry caries-specific functional gene signatures.

PubMed

Yang, Fang; Ning, Kang; Chang, Xingzhi; Yuan, Xiao; Tu, Qichao; Yuan, Tong; Deng, Ye; Hemme, Christopher L; Van Nostrand, Joy; Cui, Xinping; He, Zhili; Chen, Zhenggang; Guo, Dawei; Yu, Jiangbo; Zhang, Yue; Zhou, Jizhong; Xu, Jian

2014-01-01

Human saliva microbiota is phylogenetically divergent among host individuals yet their roles in health and disease are poorly appreciated. We employed a microbial functional gene microarray, HuMiChip 1.0, to reconstruct the global functional profiles of human saliva microbiota from ten healthy and ten caries-active adults. Saliva microbiota in the pilot population featured a vast diversity of functional genes. No significant distinction in gene number or diversity indices was observed between healthy and caries-active microbiota. However, co-presence network analysis of functional genes revealed that caries-active microbiota was more divergent in non-core genes than healthy microbiota, despite both groups exhibited a similar degree of conservation at their respective core genes. Furthermore, functional gene structure of saliva microbiota could potentially distinguish caries-active patients from healthy hosts. Microbial functions such as Diaminopimelate epimerase, Prephenate dehydrogenase, Pyruvate-formate lyase and N-acetylmuramoyl-L-alanine amidase were significantly linked to caries. Therefore, saliva microbiota carried disease-associated functional signatures, which could be potentially exploited for caries diagnosis.

Purification and Characterization of Allophanate Hydrolase (AtzF) from Pseudomonas sp. Strain ADP

PubMed Central

Shapir, Nir; Sadowsky, Michael J.; Wackett, Lawrence P.

2005-01-01

AtzF, allophanate hydrolase, is a recently discovered member of the amidase signature family that catalyzes the terminal reaction during metabolism of s-triazine ring compounds by bacteria. In the present study, the atzF gene from Pseudomonas sp. strain ADP was cloned and expressed as a His-tagged protein, and the protein was purified and characterized. AtzF had a deduced subunit molecular mass of 66,223, based on the gene sequence, and an estimated holoenzyme molecular mass of 260,000. The active protein did not contain detectable metals or organic cofactors. Purified AtzF hydrolyzed allophanate with a kcat/Km of 1.1 × 104 s−1 M−1, and 2 mol of ammonia was released per mol allophanate. The substrate range of AtzF was very narrow. Urea, biuret, hydroxyurea, methylcarbamate, and other structurally analogous compounds were not substrates for AtzF. Only malonamate, which strongly inhibited allophanate hydrolysis, was an alternative substrate, with a greatly reduced kcat/Km of 21 s−1 M−1. Data suggested that the AtzF catalytic cycle proceeds through a covalent substrate-enzyme intermediate. AtzF reacts with malonamate and hydroxylamine to generate malonohydroxamate, potentially derived from hydroxylamine capture of an enzyme-tethered acyl group. Three putative catalytically important residues, one lysine and two serines, were altered by site-directed mutagenesis, each with complete loss of enzyme activity. The identity of a putative serine nucleophile was probed using phenyl phosphorodiamidate that was shown to be a time-dependent inhibitor of AtzF. Inhibition was due to phosphoroamidation of Ser189 as shown by liquid chromatography/matrix-assisted laser desorption ionization mass spectrometry. The modified residue corresponds in sequence alignments to the nucleophilic serine previously identified in other members of the amidase signature family. Thus, AtzF affects the cleavage of three carbon-to-nitrogen bonds via a mechanism similar to that of enzymes

Structural insights into enzymatic degradation of oxidized polyvinyl alcohol.

PubMed

Yang, Yu; Ko, Tzu-Ping; Liu, Long; Li, Jianghua; Huang, Chun-Hsiang; Chan, Hsiu-Chien; Ren, Feifei; Jia, Dongxu; Wang, Andrew H-J; Guo, Rey-Ting; Chen, Jian; Du, Guocheng

2014-09-05

The ever-increasing production and use of polyvinyl alcohol (PVA) threaten our environment. Yet PVA can be assimilated by microbes in two steps: oxidation and cleavage. Here we report novel α/β-hydrolase structures of oxidized PVA hydrolase (OPH) from two known PVA-degrading organisms, Sphingopyxis sp. 113P3 and Pseudomonas sp. VM15C, including complexes with substrate analogues, acetylacetone and caprylate. The active site is covered by a lid-like β-ribbon. Unlike other esterase and amidase, OPH is unique in cleaving the CC bond of β-diketone, although it has a catalytic triad similar to that of most α/β-hydrolases. Analysis of the crystal structures suggests a double-oxyanion-hole mechanism, previously only found in thiolase cleaving β-ketoacyl-CoA. Three mutations in the lid region showed enhanced activity, with potential in industrial applications. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Thermophilic archaeal enzymes and applications in biocatalysis.

PubMed

Littlechild, Jennifer A

2011-01-01

Thermophilic enzymes have advantages for their use in commercial applications and particularly for the production of chiral compounds to produce optically pure pharmaceuticals. They can be used as biocatalysts in the application of 'green chemistry'. The thermophilic archaea contain enzymes that have already been used in commercial applications such as the L-aminoacylase from Thermococcus litoralis for the resolution of amino acids and amino acid analogues. This enzyme differs from bacterial L-aminoacylases and has similarities to carboxypeptidases from other archaeal species. An amidase/γ-lactamase from Sulfolobus solfataricus has been used for the production of optically pure γ-lactam, the building block for antiviral carbocyclic nucleotides. This enzyme has similarities to the bacterial signature amidase family. An alcohol dehydrogenase from Aeropyrum pernix has been used for the production of optically pure alcohols and is related to the zinc-containing eukaryotic alcohol dehydrogenases. A transaminase and a dehalogenase from Sulfolobus species have also been studied. The archaeal transaminase is found in a pathway for serine synthesis which is found only in eukaryotes and not in bacteria. It can be used for the asymmetric synthesis of homochiral amines of high enantioselective purity. The L-2-haloacid dehalogenase has applications both in biocatalysis and in bioremediation. All of these enzymes have increased thermostability over their mesophilic counterparts.

In vitro characterization of PlySK1249, a novel phage lysin, and assessment of its antibacterial activity in a mouse model of Streptococcus agalactiae bacteremia.

PubMed

Oechslin, Frank; Daraspe, Jean; Giddey, Marlyse; Moreillon, Philippe; Resch, Grégory

2013-12-01

Beta-hemolytic Streptococcus agalactiae is the leading cause of bacteremia and invasive infections. These diseases are treated with β-lactams or macrolides, but the emergence of less susceptible and even fully resistant strains is a cause for concern. New bacteriophage lysins could be promising alternatives against such organisms. They hydrolyze the bacterial peptidoglycan at the end of the phage cycle, in order to release the phage progeny. By using a bioinformatic approach to screen several beta-hemolytic streptococci, a gene coding for a lysin was identified on a prophage carried by Streptococcus dysgalactiae subsp. equisimilis SK1249. The gene product, named PlySK1249, harbored an original three-domain structure with a central cell wall-binding domain surrounded by an N-terminal amidase and a C-terminal CHAP domain. Purified PlySK1249 was highly lytic and bactericidal for S. dysgalactiae (2-log10 CFU/ml decrease within 15 min). Moreover, it also efficiently killed S. agalactiae (1.5-log10 CFU/ml decrease within 15 min) but not several streptococcal commensal species. We further investigated the activity of PlySK1249 in a mouse model of S. agalactiae bacteremia. Eighty percent of the animals (n = 10) challenged intraperitoneally with 10(6) CFU of S. agalactiae died within 72 h, whereas repeated injections of PlySK1249 (45 mg/kg 3 times within 24 h) significantly protected the mice (P < 0.01). Thus, PlySK1249, which was isolated from S. dysgalactiae, demonstrated high cross-lytic activity against S. agalactiae both in vitro and in vivo. These encouraging results indicated that PlySK1249 might represent a good candidate to be developed as a new enzybiotic for the treatment of systemic S. agalactiae infections.

MnSOD expression inhibited by electromagnetic pulse radiation in the rat testis.

PubMed

Zeng, LiHua; Ji, XiTuan; Zhang, YanJun; Miao, Xia; Zou, ChangXu; Lang, HaiYang; Zhang, Jie; Li, YuRong; Wang, XiaoWu; Qi, HongXing; Ren, DongQin; Guo, GuoZhen

2011-12-01

Male Sprague Dawley rats were exposed to EMP irradiation of 100 kV/m peak-to-peak e-field intensity and different numbers of pulses. Rat sperm samples were prepared for analysis of sperm qualities; Testes were assessed by transmission electron microscopy and serum hormone concentrations were examined by radioimmunoassay; Enzymatic activities of Total-superoxide dismutase(T-SOD) and manganese-superoxide dismutase (MnSOD), the mRNA levels of MnSOD and cuprozinc-superoxide dismutase (CuZnSOD), and the density of malondialdehyde (MDA) were also determined. EMP irradiation did not affect spermatozoon morphology, micronucleus formation rate, sperm number or viability, but the acrosin reaction rate decreased at 24 h and 48 h and recovered by 72 h after irradiation as compared to the controls. The ultrastructure of rat testis displayed more serious damage at 24 h than at other time points (6 h, 12 h, 48 h). Serum levels of luteotrophic hormone (LH) and testosterone (T) were elevated in irradiated rats as compared to controls. After irradiation, enzymatic activities of T-SOD and MnSOD were reduced by 24 h, consistent with the changes observed in MnSOD mRNA expression; MDA content increased at 6 h in turn. These studies have quantified the morphological damage and dysfunction in the rat reproductive system induced by EMP. The mechanism of EMP induced damage may be associated with the inhibition of MnSOD expression.

Metabolism of growth hormone releasing peptides.

PubMed

Thomas, Andreas; Delahaut, Philippe; Krug, Oliver; Schänzer, Wilhelm; Thevis, Mario

2012-12-04

New, potentially performance enhancing compounds have frequently been introduced to licit and illicit markets and rapidly distributed via worldwide operating Internet platforms. Developing fast analytical strategies to follow these new trends is one the most challenging issues for modern doping control analysis. Even if reference compounds for the active drugs are readily obtained, their unknown metabolism complicates effective testing strategies. Recently, a new class of small C-terminally amidated peptides comprising four to seven amino acid residues received considerable attention of sports drug testing authorities due to their ability to stimulate growth hormone release from the pituitary. The most promising candidates are the growth hormone releasing peptide (GHRP)-1, -2, -4, -5, -6, hexarelin, alexamorelin, and ipamorelin. With the exemption of GHRP-2, the entity of these peptides represents nonapproved pharmaceuticals; however, via Internet providers, all compounds are readily available. To date, only limited information on the metabolism of these substances is available and merely one metabolite for GHRP-2 is established. Therefore, a comprehensive in vivo (po and iv administration in rats) and in vitro (with human serum and recombinant amidase) study was performed in order to generate information on urinary metabolites potentially useful for routine doping controls. The urine samples from the in vivo experiments were purified by mixed-mode cation-exchange solid-phase extraction and analyzed by ultrahigh-performance liquid chromatography (UHPLC) separation followed by high-resolution/high-accuracy mass spectrometry. Combining the high resolution power of a benchtop Orbitrap mass analyzer for the first metabolite screening and the speed of a quadrupole/time-of-flight (Q-TOF) instrument for identification, urinary metabolites were screened by means of a sensitive full scan analysis and subsequently confirmed by high-accuracy product ion scan experiments. Two

Pharmacological characterization of hydrolysis-resistant analogs of oleoylethanolamide with potent anorexiant properties.

PubMed

Astarita, Giuseppe; Di Giacomo, Barbara; Gaetani, Silvana; Oveisi, Fariba; Compton, Timothy R; Rivara, Silvia; Tarzia, Giorgio; Mor, Marco; Piomelli, Daniele

2006-08-01

Oleoylethanolamide (OEA) is an endogenous lipid mediator that reduces food intake, promotes lipolysis, and decreases body weight gain in rodents by activating peroxisome proliferator-activated receptor-alpha (PPAR-alpha). The biological effects of OEA are terminated by two intracellular lipid hydrolase enzymes, fatty-acid amide hydrolase and N-acylethanolamine-hydrolyzing acid amidase. In the present study, we describe OEA analogs that resist enzymatic hydrolysis, activate PPAR-alpha with high potency in vitro, and persistently reduce feeding when administered in vivo either parenterally or orally. The most potent of these compounds, (Z)-(R)-9-octadecenamide,N-(2-hydroxyethyl,1-methyl) (KDS-5104), stimulates transcriptional activity of PPAR-alpha with a half-maximal effective concentration (EC50) of 100 +/- 21 nM (n = 11). Parenteral administration of KDS-5104 in rats produces persistent dose-dependent prolongation of feeding latency and postmeal interval (half-maximal effective dose, ED50 = 2.4 +/- 1.8 mg kg(-1) i.p.; n = 18), as well as increased and protracted tissue exposure compared with OEA. Oral administration of the compound also results in a significant tissue exposure and reduction of food intake in free-feeding rats. These results suggest that the endogenous high-affinity PPAR-alpha agonist OEA may provide a scaffold for the discovery of novel orally active PPAR-alpha ligands.

Purification and Partial Characterization of a Novel Bacteriocin Synthesized by Lactobacillus paracasei HD1-7 Isolated from Chinese Sauerkraut Juice.

PubMed

Ge, Jingping; Sun, Yanyang; Xin, Xing; Wang, Ying; Ping, Wenxiang

2016-01-14

Bacteriocins have antimicrobial activities against food-spoiling bacteria and food-borne pathogens. Paracin 1.7, a bacteriocin synthesized by Lactobacillus paracasei HD1-7 isolated from Chinese sauerkraut juice, was studied. Following partial purification with ammonium sulfate precipitation, CM Sepharose Fast Flow, and Sephadex G-10 chromatography, the molecular weight of Paracin 1.7 was about 10 kDa based on Tricine-SDS-PAGE results. A 2.87 fold purified bacteriocin was produced, reaching a final yield of 39.93% and the specific activity of 1.56 × 10(3) AU/mg. The N-terminal amino acid sequence of Paracin 1.7 was VSNTFFA, and the LC/LTQ results revealed that the N-terminal amino acid sequence was similar to that of ABC-type oligopeptide transport system protein and N-acetylmuramoyl-L-alanine amidase. Paracin 1.7 was sensitive to protease K, had antimicrobial activities at a broad pH range (3.0-8.0), and was heat resistant (121 °C for 20 min). Paracin 1.7 from Lactobacillus paracasei HD1-7 is a novel bacteriocin that has potential applications in food preservation.

Purification and Partial Characterization of a Novel Bacteriocin Synthesized by Lactobacillus paracasei HD1-7 Isolated from Chinese Sauerkraut Juice

PubMed Central

Ge, Jingping; Sun, Yanyang; Xin, Xing; Wang, Ying; Ping, Wenxiang

2016-01-01

Bacteriocins have antimicrobial activities against food-spoiling bacteria and food-borne pathogens. Paracin 1.7, a bacteriocin synthesized by Lactobacillus paracasei HD1-7 isolated from Chinese sauerkraut juice, was studied. Following partial purification with ammonium sulfate precipitation, CM Sepharose Fast Flow, and Sephadex G-10 chromatography, the molecular weight of Paracin 1.7 was about 10 kDa based on Tricine-SDS-PAGE results. A 2.87 fold purified bacteriocin was produced, reaching a final yield of 39.93% and the specific activity of 1.56 × 103 AU/mg. The N-terminal amino acid sequence of Paracin 1.7 was VSNTFFA, and the LC/LTQ results revealed that the N-terminal amino acid sequence was similar to that of ABC-type oligopeptide transport system protein and N-acetylmuramoyl-L-alanine amidase. Paracin 1.7 was sensitive to protease K, had antimicrobial activities at a broad pH range (3.0–8.0), and was heat resistant (121 °C for 20 min). Paracin 1.7 from Lactobacillus paracasei HD1-7 is a novel bacteriocin that has potential applications in food preservation. PMID:26763314

A Thermophilic Phage Endolysin Fusion to a Clostridium perfringens-Specific Cell Wall Binding Domain Creates an Anti-Clostridium Antimicrobial with Improved Thermostability.

PubMed

Swift, Steven M; Seal, Bruce S; Garrish, Johnna K; Oakley, Brian B; Hiett, Kelli; Yeh, Hung-Yueh; Woolsey, Rebekah; Schegg, Kathleen M; Line, John Eric; Donovan, David M

2015-06-12

Clostridium perfringens is the third leading cause of human foodborne bacterial disease and is the presumptive etiologic agent of necrotic enteritis among chickens. Treatment of poultry with antibiotics is becoming less acceptable. Endolysin enzymes are potential replacements for antibiotics. Many enzymes are added to animal feed during production and are subjected to high-heat stress during feed processing. To produce a thermostabile endolysin for treating poultry, an E. coli codon-optimized gene was synthesized that fused the N-acetylmuramoyl-L-alanine amidase domain from the endolysin of the thermophilic bacteriophage ɸGVE2 to the cell-wall binding domain (CWB) from the endolysin of the C. perfringens-specific bacteriophage ɸCP26F. The resulting protein, PlyGVE2CpCWB, lysed C. perfringens in liquid and solid cultures. PlyGVE2CpCWB was most active at pH 8, had peak activity at 10 mM NaCl, 40% activity at 150 mM NaCl and was still 16% active at 600 mM NaCl. The protein was able to withstand temperatures up to 50° C and still lyse C. perfringens. Herein, we report the construction and characterization of a thermostable chimeric endolysin that could potentially be utilized as a feed additive to control the bacterium during poultry production.

The SPOR Domain, a Widely Conserved Peptidoglycan Binding Domain That Targets Proteins to the Site of Cell Division.

PubMed

Yahashiri, Atsushi; Jorgenson, Matthew A; Weiss, David S

2017-07-15

Sporulation-related repeat (SPOR) domains are small peptidoglycan (PG) binding domains found in thousands of bacterial proteins. The name "SPOR domain" stems from the fact that several early examples came from proteins involved in sporulation, but SPOR domain proteins are quite diverse and contribute to a variety of processes that involve remodeling of the PG sacculus, especially with respect to cell division. SPOR domains target proteins to the division site by binding to regions of PG devoid of stem peptides ("denuded" glycans), which in turn are enriched in septal PG by the intense, localized activity of cell wall amidases involved in daughter cell separation. This targeting mechanism sets SPOR domain proteins apart from most other septal ring proteins, which localize via protein-protein interactions. In addition to SPOR domains, bacteria contain several other PG-binding domains that can exploit features of the cell wall to target proteins to specific subcellular sites. Copyright © 2017 American Society for Microbiology.

Occurrence of enzymes involved in biosynthesis of indole-3-acetic acid from indole-3-acetonitrile in plant-associated bacteria, Agrobacterium and Rhizobium.

PubMed Central

Kobayashi, M; Suzuki, T; Fujita, T; Masuda, M; Shimizu, S

1995-01-01

The occurrence of a hitherto unknown pathway involving the action of two enzymes, a nitrile hydratase and an amidase for the biosynthesis of indole-3-acetic acid was discovered in phytopathogenic bacteria Agrobacterium tumefaciens and in leguminous bacteria Rhizobium. The nitrile hydratase acting on indole-3-acetonitrile was purified to homogeneity through only two steps from the cell-free extract of A. tumefaciens. The molecular mass of the purified enzyme estimated by HPLC was about 102 kDa, and the enzyme consisted of four subunits identical in molecular mass. The enzyme exhibited a broad absorption spectrum in the visible range with absorption maxima at 408 nm and 705 nm, and it contained cobalt and iron. The enzyme stoichiometrically catalyzed the hydration of indole-3-acetonitrile into indole-3-acetamide with a specific activity of 13.7 mol per min per mg and a Km of 7.9 microM. Images Fig. 1 PMID:11607511

Isolation and structures of glycoprotein-derived free oligosaccharides from the unfertilized eggs of Scyliorhinus caniculus. Characterization of the sequences galactose(alpha 1-4)galactose(beta 1-3)-N-acetylglucosamine and N-acetylneuraminic acid(alpha 2-6)galactose(beta 1-3)-N-acetylglucosamine.

PubMed

Plancke, Y; Delplace, F; Wieruszeski, J M; Maes, E; Strecker, G

1996-01-15

As previously reported [Ishii, K., Iwasaki, M., Inoue, S., Kenny, P. T. M., Komura, H. & Inoue, Y. (1989) J. Biol. Chem. 264, 1623-1630; Inoue, S., Iwasaki, M., Ishii, K., Kitajima, K. & Inoue, Y. (1989) J. Biol. Chem. 264, 18520-185261, the unfertilized eggs of two different species of fresh-water fish, Plecoglossus altivelis and Tribodolon hakonensis, contain relatively large amounts of free sialooligosaccharides. These oligosaccharides were found to derive from glycophosphoproteins, owing to the activity of a peptide - N4-(N-acetyl-beta-D-glucosaminyl)asparagine amidase [Iwasaki, M., Seko, A., Kitajima, K., Inoue, Y. & Inoue, S. (1992) J. Biol. Chem. 267, 24287-24296; Seko, A., Kitajima, K., Inoue, Y. & Inoue, S. (1991) J. Biol. Chem. 266, 22110-22114]. Here we describe a new type of free oligosaccharides, isolated from unfertilized eggs of Scyliorhinus caniculus. From the structural analysis, based upon 1H-NMR spectroscopy, the following glycan units are proposed.[Formula: see text

α-Ketoglutaramate: An overlooked metabolite of glutamine and a biomarker for hepatic encephalopathy and inborn errors of the urea cycle

PubMed Central

Cooper, Arthur J. L.; Kuhara, Tomiko

2013-01-01

Glutamine metabolism is generally regarded as proceeding via glutaminase-catalyzed hydrolysis to glutamate and ammonia, followed by conversion of glutamate to α-ketoglutarate catalyzed by glutamate dehydrogenase or by a glutamate-linked aminotransferase (transaminase). However, another pathway exists for the conversion of glutamine to α-ketoglutarate that is often overlooked, but is widely distributed in nature. This pathway, referred to as the glutaminase II pathway, consists of a glutamine transaminase coupled to ω-amidase. Transamination of glutamine results in formation of the corresponding α-keto acid, namely, α-ketoglutaramate (KGM). KGM is hydrolyzed by ω-amidase to α-ketoglutarate and ammonia. The net glutaminase II reaction is: L-Glutamine + α-keto acid + H2O → α-ketoglutarate + L-amino acid + ammonia. In this mini-review the biochemical importance of the glutaminase II pathway is summarized, with emphasis on the key component KGM. Forty years ago it was noted that the concentration of KGM is increased in the cerebrospinal fluid (CSF) of patients with hepatic encephalopathy (HE) and that the level of KGM in the CSF correlates well with the degree of encephalopathy. In more recent work, we have shown that KGM is markedly elevated in the urine of patients with inborn errors of the urea cycle. It is suggested that KGM may be a useful biomarker for many hyperammonemic diseases including hepatic encephalopathy, inborn errors of the urea cycle, citrin deficiency and lysinuric protein intolerance. PMID:24234505

Red wine consumption may affect sperm biology: the effects of different concentrations of the phytoestrogen myricetin on human male gamete function.

PubMed

Aquila, Saveria; Santoro, Marta; De Amicis, Francesca; Guido, Carmela; Bonofiglio, Daniela; Lanzino, Marilena; Cesario, Maria Grazia; Perrotta, Ida; Sisci, Diego; Morelli, Catia

2013-02-01

Myricetin is a natural flavonoid, particularly enriched in red wines, whose occurrence is widespread among plants. Despite extensive research, the beneficial effects of Myricetin on human health are still controversial. Here, we tested the estrogen-like effect of the phytoestrogen Myricetin on human ejaculated sperm biology. To this aim, human normozoospermic samples were exposed to increasing concentrations (10 nM, 100 nM, and 1 µM) of Myricetin. Motility, viability, capacitation-associated biochemical changes (i.e., cholesterol efflux and tyrosine phosphorylation), acrosin activity, as well as glucose utilization and fatty-acid oxidation (i.e., glucose and lipid metabolism) were all significantly increased by low doses of Myricetin. Importantly, both estrogen receptors α and β (ERs) and phosphatidylinositol-3-OH kinase (PI3K)/AKT signaling are activated in the presence of Myricetin since these were both abrogated by specific inhibitors of each pathway. Our results show how Myricetin, through ERs and PI3K/AKT signalings, potentiates sperm function. This effect is dose-dependent at low concentrations of Myricetin (up to 100 nM), whereas higher amounts do not seem to improve any further sperm motility, viability, or other tested features, and, in some cases, they reduced or even abrogated the efficacy exerted by lower doses. Further studies are needed to elucidate if high levels of Myricetin, which could be attained even with moderate wine consumption, could synergize with endogenous estrogens in the female reproductive tract, interfering with the physiological sperm fertilization process. Copyright © 2012 Wiley Periodicals, Inc.

Degradation of the metal-cyano complex tetracyanonickelate (II) by Fusarium oxysporum N-10.

PubMed

Yanase, H; Sakamoto, A; Okamoto, K; Kita, K; Sato, Y

2000-03-01

A fungus with the ability to utilize a metalcyano compound, tetracyanonickelate (II) ¿K2[Ni (CN)4]; TCN¿, as its sole source of nitrogen was isolated from soil and identified as Fusarium oxysporum N-10. Both intact mycelia and cell-free extract of the strain catalyzed hydrolysis of TCN to formate and ammonia and produced formamide as an intermediate, thereby indicating that a hydratase and an amidase sequentially participated in the degradation of TCN. The enzyme catalyzing the hydration of TCN was purified approximately ten-fold from the cell-free extract of strain N-10 with a yield of 29%. The molecular mass of the active enzyme was estimated to be 160 kDa. The enzyme appears to exist as a homotetramer, each subunit having a molecular mass of 40 kDa. The enzyme also catalyzed the hydration of KCN, with a cyanide-hydrating activity 2 x 10(4) times greater than for TCN. The kinetic parameters for TCN and KCN indicated that hydratase isolated from F. oxysporum was a cyanide hydratase able to utilize a broad range of cyano compounds and nitriles as substrates.

Determination of multidrug resistance mechanisms in Clostridium perfringens type A isolates using RNA sequencing and 2D-electrophoresis.

PubMed

Ma, Yu-Hua; Ye, Gui-Sheng

2018-06-11

In this study, we screened differentially expressed genes in a multidrug-resistant isolate strain of Clostridium perfringens by RNA sequencing. We also separated and identified differentially expressed proteins (DEPs) in the isolate strain by two-dimensional electrophoresis (2-DE) and mass spectrometry (MS). The RNA sequencing results showed that, compared with the control strain, 1128 genes were differentially expressed in the isolate strain, and these included 227 up-regulated genes and 901 down-regulated genes. Bioinformatics analysis identified the following genes and gene categories that are potentially involved in multidrug resistance (MDR) in the isolate strain: drug transport, drug response, hydrolase activity, transmembrane transporter, transferase activity, amidase transmembrane transporter, efflux transmembrane transporter, bacterial chemotaxis, ABC transporter, and others. The results of the 2-DE showed that 70 proteins were differentially expressed in the isolate strain, 45 of which were up-regulated and 25 down-regulated. Twenty-seven DEPs were identified by MS and these included the following protein categories: ribosome, antimicrobial peptide resistance, and ABC transporter, all of which may be involved in MDR in the isolate strain of C. perfringens. The results provide reference data for further investigations on the drug resistant molecular mechanisms of C. perfringens.

Enzymatic preparation of optically pure (+)-2-azabicyclo[2.2.1]hept-5-en-3-one by (-)-γ-lactamase from Bradyrhizobium japonicum USDA 6.

PubMed

Zhu, Shaozhou; Ren, Lu; Yu, Songzhu; Gong, Cuiyu; Song, Dawei; Zheng, Guojun

2014-10-15

Whole cells of Bradyrhizobium japonicum USDA 6 showed both (+)-γ-lactamase activity and (-)-γ-lactamase activity. Insight into the genome of B. japonicum USDA 6 revealed two potential γ-lactamases: a type I (+)-γ-lactamase and a (-)-γ-lactamase, making it the first strain to contain two totally different enantioselective lactamases. Both recombinant enzymes could easily be used to prepare either optically pure (+)-γ-lactam ((+)-2-azabicyclo[2.2.1]hept-5-en-3-one) or optically pure (-)-γ-lactam ((-)-2-azabicyclo[2.2.1]hept-5-en-3-one), which are versatile synthetic building blocks for the synthesis of various carbocyclic nucleosides and carbocyclic sugar analogues. Bioinformatic analysis showed that the type I (+)-γ-lactamase belongs to the amidase signature family, with 504 amino acids; the (-)-γ-lactamase, which consists of 274 amino acids, belongs to the hydrolase family. Here, we report that B. japonicum USDA contains a (-)-γ-lactamase in addition to a (+)-γ-lactamase, and it is the (-)-γ-lactamase from this strain that is examined in detail in this Letter. Enzymatic synthesis of optically pure (+)-γ-lactam with nearly 50% isolated yield and >99% ee was achieved. Copyright © 2014 Elsevier Ltd. All rights reserved.

Cloning and expression of a conjugated bile acid hydrolase gene from Lactobacillus plantarum by using a direct plate assay.

PubMed

Christiaens, H; Leer, R J; Pouwels, P H; Verstraete, W

1992-12-01

The conjugated bile acid hydrolase gene from the silage isolate Lactobacillus plantarum 80 was cloned and expressed in Escherichia coli MC1061. For the screening of this hydrolase gene within the gene bank, a direct plate assay developed by Dashkevicz and Feighner (M. P. Dashkevicz and S. D. Feighner, Appl. Environ. Microbiol. 53:331-336, 1989) was adapted to the growth requirements of E. coli. Because of hydrolysis and medium acidification, hydrolase-active colonies were surrounded with big halos of precipitated, free bile acids. This phenomenon was also obtained when the gene was cloned into a multicopy shuttle vector and subsequently reintroduced into the parental Lactobacillus strain. The cbh gene and surrounding regions were characterized by nucleotide sequence analysis. The deduced amino acid sequence was shown to have 52% similarity with a penicillin V amidase from Bacillus sphaericus. Preliminary characterization of the gene product showed that it is a cholylglycine hydrolase (EC 3.5.1.24) with only slight activity against taurine conjugates. The optimum pH was between 4.7 and 5.5. Optimum temperature ranged from 30 to 45 degrees C. Southern blot analysis indicated that the cloned gene has similarity with genomic DNA of bile acid hydrolase-active Lactobacillus spp. of intestinal origin.

Glutamine metabolism and cycling in Neurospora crassa.

PubMed Central

Mora, J

1990-01-01

Evidence for the existence of a glutamine cycle in Neurospora crassa is reviewed. Through this cycle glutamine is converted into glutamate by glutamate synthase and catabolized by the glutamine transaminase-omega-amidase pathway, the products of which (2-oxoglutarate and ammonium) are the substrates for glutamate dehydrogenase-NADPH, which synthesizes glutamate. In the final step ammonium is assimilated into glutamine by the action of a glutamine synthetase (GS), which is formed by two distinct polypeptides, one catalytically very active (GS beta), and the other (GS alpha) less active but endowed with the capacity to modulate the activity of GS alpha. Glutamate synthase uses the amide nitrogen of glutamine to synthesize glutamate; glutamate dehydrogenase uses ammonium, and both are required to maintain the level of glutamate. The energy expended in the synthesis of glutamine drives the cycle. The glutamine cycle is not futile, because it is necessary to drive an effective carbon flow to support growth; in addition, it facilitates the allocation of nitrogen or carbon according to cellular demands. The glutamine cycle which dissipates energy links catabolism and anabolism and, in doing so, buffers variations in the nutrient supply and drives energy generation and carbon flow for optimal cell function. PMID:2145504

Molecular cloning and characterization of a short peptidoglycan recognition protein from silkworm Bombyx mori.

PubMed

Yang, P-J; Zhan, M-Y; Ye, C; Yu, X-Q; Rao, X-J

2017-12-01

Peptidoglycan is the major bacterial component recognized by the insect immune system. Peptidoglycan recognition proteins (PGRPs) are a family of pattern-recognition receptors that recognize peptidoglycans and modulate innate immune responses. Some PGRPs retain N-acetylmuramoyl-L-alanine amidase (Enzyme Commission number: 3.5.1.28) activity to hydrolyse bacterial peptidoglycans. Others have lost the enzymatic activity and work only as immune receptors. They are all important modulators for innate immunity. Here, we report the cloning and functional analysis of PGRP-S4, a short-form PGRP from the domesticated silkworm, Bombyx mori. The PGRP-S4 gene encodes a protein of 199 amino acids with a signal peptide and a PGRP domain. PGRP-S4 was expressed in the fat body, haemocytes and midgut. Its expression level was significantly induced by bacterial challenges in the midgut. The recombinant PGRP-S4 bound bacteria and different peptidoglycans. In addition, it inhibited bacterial growth and hydrolysed an Escherichia coli peptidoglycan in the presence of Zn 2+ . Scanning electron microscopy showed that PGRP-S4 disrupted the bacterial cell surface. PGRP-S4 further increased prophenoloxidase activation caused by peptidoglycans. Taken together, our data suggest that B. mori PGRP-S4 has multiple functions in immunity. © 2017 The Royal Entomological Society.

Antiacanthain A: New proteases isolated from Bromelia antiacantha Bertol. (Bromeliaceae).

PubMed

Vallés, Diego; Cantera, Ana M B

2018-07-01

Crude extract (CE) from pulp of Bromelia antiacantha Bertol. mature fruit, contains at least 3 cysteine proteases with proteolytic activity. By single step cation exchange chromatography (Hi-trap SP-HP) of partially purified CE, the protease with the lowest pI, Antiacanthain A (AntA), was isolated. It showed maximum activity at pH9, and 75% of remaining activity was maintained over a wide pH range (pH6-10). The AntA activity exhibits a constant increase up to 70°C. Maintains almost 100% of its activity at 45 at pH6 and 9. A 60% of AntA was active by titration with specific inhibitor, E64. Amidasic activity was studied with pyroglutamyl-phenyl-leucyl-paranitroaniline (PFLNA) substrate having higher AntA catalytic efficiency of (k cat /K m =470s -1 M -1 ) relative to stem bromelain (k cat /K m =305s -1 M -1 ). Esterase activity using p-nitrophenyl esters of N-α-CBZ-l-Lysine (z-L-LysONp) showed a 10-fold higher catalytic efficiency for AntA (k cat /K m =6376s -1 M -1 ) relative to stem bromelain (k cat /K m =688s -1 M -1 ). Incubation with 8M Urea did not affect AntA activity and remained unchanged for 18h, with 6M GndHCl resulted in a 41% decrease in activity after 30min incubation, maintained this activity 18h. AntA exhibits high sequence identity with proteases of the Bromeliaceae family. Copyright © 2018 Elsevier B.V. All rights reserved.

Acrylamide biodegradation ability and plant growth-promoting properties of Variovorax boronicumulans CGMCC 4969.

PubMed

Liu, Zhong-Hua; Cao, Yu-Min; Zhou, Qian-Wen; Guo, Kun; Ge, Feng; Hou, Jun-Yi; Hu, Si-Yi; Yuan, Sheng; Dai, Yi-Jun

2013-11-01

Species of the genus Variovorax are often isolated from nitrile or amide-containing organic compound-contaminated soil. However, there have been few biological characterizations of Variovorax and their contaminant-degrading enzymes. Previously, we reported a new soil isolate, Variovorax boronicumulans CGMCC 4969, and its nitrile hydratase that transforms the neonicotinoid insecticide thiacloprid into an amide metabolite. In this study, we showed that CGMCC 4969 is able to degrade acrylamide, a neurotoxicant and carcinogen in animals, during cell growth in a mineral salt medium as well as in its resting state. Resting cells rapidly hydrolyzed 600 mg/L acrylamide to acrylic acid with a half-life of 2.5 min. In in vitro tests, CGMCC 4969 showed plant growth-promoting properties; it produced a siderophore, ammonia, hydrogen cyanide, and the phytohormone salicylic acid. Interestingly, in soil inoculated with this strain, 200 mg/L acrylamide was completely degraded in 4 days. Gene cloning and overexpression in the Escherichia coli strain Rosetta (DE3) pLysS resulted in the production of an aliphatic amidase of 345 amino acids that hydrolyzed acrylamide into acrylic acid. The amidase contained a conserved catalytic triad, Glu59, Lys 134, and Cys166, and an "MRHGDISSS" amino acid sequence at the N-terminal region. Variovorax boronicumulans CGMCC 4969, which is able to use acrylamide for cell growth and rapidly degrade acrylamide in soil, shows promising plant growth-promoting properties. As such, it has the potential to be developed into an effective Bioaugmentation strategy to promote growth of field crops in acrylamide-contaminated soil.

Sperm quality and fertility of boar seminal doses after 2 days of storage: does the type of extender really matter?

PubMed

Pinart, Elisabeth; Yeste, Marc; Prieto-Martínez, Noelia; Reixach, Josep; Bonet, Sergi

2015-06-01

The present approach was designed to evaluate the extender effects on sperm quality and fertility of short-term refrigerated seminal doses from Landrace boars lodged in husbandry-controlled conditions. For this purpose, we analyzed the sperm quality of seminal doses diluted in short-term (Beltsville Thawing Solution) and extra-long-term (Duragen) extenders from Days 0 to 2 of storage at 17 °C during an 8-month period. Pregnancy rates and litter size were evaluated from double inseminations within an interval of 12 hours (36 and 48 hours of refrigeration) of multiparous females using seminal doses diluted in each extender type. Sperm quality was assessed from the analyses of sperm motility and kinetics, sperm viability, expressed as plasma and acrosome membrane integrity, membrane lipid disorder, intracellular calcium levels, and acrosin activity. Results indicated significant differences between the extenders in the sperm quality of seminal doses. Therefore, the seminal doses diluted in Duragen had higher percentages of progressive motile spermatozoa and membrane-intact spermatozoa than those diluted in Beltsville Thawing Solution throughout all the experimental months. Nevertheless, despite these differences in preserving the sperm quality, pregnancy rates (>90%) and litter sizes (>10 piglets born per litter) were similar between the extenders. Our results had great relevance from a practical point of view because they reported lack of an extender effect on the reproductive performance of seminal doses during short-tem storage. Copyright © 2015 Elsevier Inc. All rights reserved.

Sperm function and assisted reproduction technology

PubMed Central

MAAß, GESA; BÖDEKER, ROLF‐HASSO; SCHEIBELHUT, CHRISTINE; STALF, THOMAS; MEHNERT, CLAAS; SCHUPPE, HANS‐CHRISTIAN; JUNG, ANDREAS; SCHILL, WOLF‐BERNHARD

2005-01-01

The evaluation of different functional sperm parameters has become a tool in andrological diagnosis. These assays determine the sperm's capability to fertilize an oocyte. It also appears that sperm functions and semen parameters are interrelated and interdependent. Therefore, the question arose whether a given laboratory test or a battery of tests can predict the outcome in in vitro fertilization (IVF). One‐hundred and sixty‐one patients who underwent an IVF treatment were selected from a database of 4178 patients who had been examined for male infertility 3 months before or after IVF. Sperm concentration, motility, acrosin activity, acrosome reaction, sperm morphology, maternal age, number of transferred embryos, embryo score, fertilization rate and pregnancy rate were determined. In addition, logistic regression models to describe fertilization rate and pregnancy were developed. All the parameters in the models were dichotomized and intra‐ and interindividual variability of the parameters were assessed. Although the sperm parameters showed good correlations with IVF when correlated separately, the only essential parameter in the multivariate model was morphology. The enormous intra‐ and interindividual variability of the values was striking. In conclusion, our data indicate that the andrological status at the end of the respective treatment does not necessarily represent the status at the time of IVF. Despite a relatively low correlation coefficient in the logistic regression model, it appears that among the parameters tested, the most reliable parameter to predict fertilization is normal sperm morphology. (Reprod Med Biol 2005; 4: 7–30) PMID:29699207

Molecular cloning and functional characterization of a short peptidoglycan recognition protein (HcPGRPS1) from the freshwater mussel, Hyriopsis cumingi.

PubMed

Yang, Ziyan; Li, Junhua; Li, Ying; Wu, Hongjuan; Wang, Xiaoyan

2013-12-01

Peptidoglycan recognition proteins (PGRPs), which are evolutionarily conserved from invertebrates to vertebrates, function as pattern-recognition and effector molecules in innate immunity. In the present study, a short-form PGRP, designated as HcPGRPS1 was identified from freshwater mussel Hyriopsis cumingi. The deduced amino acid sequence of HcPGRPS1 is composed of 235 residues which contains a conserved PGRP domain at the C-terminus. Sequence analysis showed that HcPGRPS1 shared high identities with other known PGRPs. The mRNA of HcPGRPS1 is constitutively expressed in a wide range of all tested tissues, with highest expression level in hepatopancreas, and its expression in tissues (gonad, nephridium, gill and foot) was up-regulated significantly after LPS or PGN stimulation (P<0.05). The recombinant protein of HcPGRPS1 exhibited binding activity and peptidoglycan-lytic amidase activity toward Lys-PGN from Staphylococcus aureus and DAP-PGN from Bacillus subtilis. Furthermore, recombinant HcPGRPS1 displayed strong antibacterial activity to both Gram-negative bacteria Escherichia coli, Aeromonas hydrophila, Aeromonas sobria and Gram-positive bacteria S. aureus in the presence of Zn(2+). These results suggested that HcPGRPS1 plays a multifunctional role in the defense and protection mechanisms of mussel innate immunity against infections. Copyright © 2013 Elsevier Ltd. All rights reserved.

Bacteriophages of the family siphoviridae contain amidase enzymes that lyse Clostridium perfringens

USDA-ARS?s Scientific Manuscript database

*Agtech-Danisco, current address In chickens Clostridium perfringens (Cp) is the etiologic agent of necrotic enteritis and causes gas gangrene along with being the third leading cause of bacterial food-borne gastroenteritis in humans. While the disease in poultry can be controlled by antibiotics, th...

BACTERIOPHAGES OF THE FAMILY SIPHOVIRIDAE CONTAIN AMIDASE ENZYMES THAT LYSE CLOSTRIDIUM PERFRINGENS

USDA-ARS?s Scientific Manuscript database

In chickens Clostridium perfringens (Cp) is the etiologic agent of necrotic enteritis and causes gas gangrene along with being the third leading cause of bacterial food-borne gastroenteritis in humans. While the disease in poultry can be controlled by antibiotics, there is increasing pressure to ban...

Crystal Structures of Fatty Acid Amide Hydrolase Bound to the Carbamate Inhibitor URB597: Discovery of a Deacylating Water Molecule and Insight into Enzyme Inactivation

PubMed Central

Mileni, Mauro; Kamtekar, Satwik; Wood, David C.; Benson, Timothy E.; Cravatt, Benjamin F.; Stevens, Raymond C.

2010-01-01

The endocannabinoid system regulates a wide range of physiological processes including pain, inflammation, and cognitive/emotional states. URB597 is one of the best characterized covalent inhibitors of the endocannabinoid-degrading enzyme fatty acid amide hydrolase (FAAH). Here, we report the structure of the FAAH-URB597 complex at 2.3 Å resolution. The structure provides insights into mechanistic details of enzyme inactivation and experimental evidence of a previously uncharacterized active site water molecule that likely is involved in substrate deacylation. This water molecule is part of an extensive hydrogen-bonding network, and is coordinated indirectly to residues lining the cytosolic port of the enzyme. In order to corroborate our hypothesis concerning the role of this water molecule in FAAH’s catalytic mechanism, we determined the structure of FAAH conjugated to a urea-based inhibitor, PF-3845, to a higher resolution (2.4 Å) than previously reported. The higher resolution structure confirms the presence of the water molecule in a virtually identical location in the active site. Examination of the structures of serine hydrolases that are non-homologous to FAAH, such as elastase, trypsin, or chymotrypsin, shows a similarly positioned hydrolytic water molecule and suggest a functional convergence between the amidase signature enzymes and serine proteases. PMID:20493882

Crystal structure of fatty acid amide hydrolase bound to the carbamate inhibitor URB597: discovery of a deacylating water molecule and insight into enzyme inactivation.

PubMed

Mileni, Mauro; Kamtekar, Satwik; Wood, David C; Benson, Timothy E; Cravatt, Benjamin F; Stevens, Raymond C

2010-07-23

The endocannabinoid system regulates a wide range of physiological processes including pain, inflammation, and cognitive/emotional states. URB597 is one of the best characterized covalent inhibitors of the endocannabinoid-degrading enzyme fatty acid amide hydrolase (FAAH). Here, we report the structure of the FAAH-URB597 complex at 2.3 A resolution. The structure provides insights into mechanistic details of enzyme inactivation and experimental evidence of a previously uncharacterized active site water molecule that likely is involved in substrate deacylation. This water molecule is part of an extensive hydrogen-bonding network and is coordinated indirectly to residues lining the cytosolic port of the enzyme. In order to corroborate our hypothesis concerning the role of this water molecule in FAAH's catalytic mechanism, we determined the structure of FAAH conjugated to a urea-based inhibitor, PF-3845, to a higher resolution (2.4 A) than previously reported. The higher-resolution structure confirms the presence of the water molecule in a virtually identical location in the active site. Examination of the structures of serine hydrolases that are non-homologous to FAAH, such as elastase, trypsin, or chymotrypsin, shows a similarly positioned hydrolytic water molecule and suggests a functional convergence between the amidase signature enzymes and serine proteases. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Characterization of Foodborne Strains of Staphylococcus aureus by Shotgun Proteomics: Functional Networks, Virulence Factors and Species-Specific Peptide Biomarkers

PubMed Central

Carrera, Mónica; Böhme, Karola; Gallardo, José M.; Barros-Velázquez, Jorge; Cañas, Benito; Calo-Mata, Pilar

2017-01-01

In the present work, we applied a shotgun proteomics approach for the fast and easy characterization of 20 different foodborne strains of Staphylococcus aureus (S. aureus), one of the most recognized foodborne pathogenic bacteria. A total of 644 non-redundant proteins were identified and analyzed via an easy and rapid protein sample preparation procedure. The results allowed the differentiation of several proteome datasets from the different strains (common, accessory, and unique datasets), which were used to determine relevant functional pathways and differentiate the strains into different Euclidean hierarchical clusters. Moreover, a predicted protein-protein interaction network of the foodborne S. aureus strains was created. The whole confidence network contains 77 nodes and 769 interactions. Most of the identified proteins were surface-associated proteins that were related to pathways and networks of energy, lipid metabolism and virulence. Twenty-seven virulence factors were identified, and most of them corresponded to autolysins, N-acetylmuramoyl-L-alanine amidases, phenol-soluble modulins, extracellular fibrinogen-binding proteins and virulence factor EsxA. Potential species-specific peptide biomarkers were screened. Twenty-one species-specific peptide biomarkers, belonging to eight different proteins (nickel-ABC transporter, N-acetylmuramoyl-L-alanine amidase, autolysin, clumping factor A, gram-positive signal peptide YSIRK, cysteine protease/staphopain, transcriptional regulator MarR, and transcriptional regulator Sar-A), were proposed to identify S. aureus. These results constitute the first major dataset of peptides and proteins of foodborne S. aureus strains. This repository may be useful for further studies, for the development of new therapeutic treatments for S. aureus food intoxications and for microbial source-tracking in foodstuffs. PMID:29312172

Physiological aspects of fungi isolated from root nodules of faba bean (Vicia faba L.).

PubMed

Omar, S A; Abd-Alla, M H

2000-03-01

The present study was made to isolate and assess some physiological characteristics of root nodule-colonizing fungi. During this study, 17 fungal species were isolated from root nodule samples taken from faba bean plants (Vicia faba L.) collected from different sites at Assiut area (Egypt). The growth of faba bean plants in pots was significantly promoted by soil inoculation with most fungi. Growth was checked in pots with inocula of Cladosporium cladosporioides, Fusarium moniliforme, F: oxysporium, F solani, Macrophominia phaseolina and Rhizoctonia solani which were added separately. All growth-promoting fungi were capable of producing cellulase, pectin lyase, polygalacturonase, protease, urease, amidase, acid phosphatase, alkaline phosphatase and arylsulfatase in growth medium supplemented with the corresponding substrates. Four fungal species, Aspergillus awamori, A. flavus, Penicillium chrysogenum and Trichoderma koningii showed the highest rates of enzyme formation. The effect of the addition of six trace elements to the growth media at 30 micromol/ml on enzyme production revealed some dependency on species, enzyme and metal ion. Cd2+, Hg2+ and Zn2+ generally inhibited enzyme activity. Cu(1+), Fe3+ and Al3+ showed a stimulatory effect. Fungicides (afugan and tilt) and herbicides (brominal and fusilade) at 50 ppm generally promoted enzyme activity, but insecticides (kelthane and fenvalerate) caused some inhibition to enzyme activities. Salinization of the growth media with NaCl strongly inhibited the enzymatic activity of all fungi at concentrations between 0.5 and 1.5%.

Development and validation of an automated enzyme assay for paracetamol (acetaminophen).

PubMed

Morris, H C; Overton, P D; Ramsay, J R; Campbell, R S; Hammond, P M; Atkinson, T; Price, C P

1990-02-28

A rapid, enzymatic assay for serum or plasma paracetamol has been developed with the potential for adaptation to a wide range of clinical analysers. The method involves the action of an amidase enzyme to produce 4-aminophenol from paracetamol, which in turn reacts with 8-hydroxyquinoline in the presence of manganese ions to form a blue dye. Two stable reagents are used and excellent precision is achieved over the drug concentration range 0-2.5 mmol/l. The method, which is complete within 6 min, has been validated using a Monarch centrifugal analyser and shows no significant interference from endogenous serum compounds, drugs or paracetamol metabolites.

Expression, purification, and characterization of a bifunctional 99-kDa peptidoglycan hydrolase from Pediococcus acidilactici ATCC 8042.

PubMed

García-Cano, Israel; Campos-Gómez, Manuel; Contreras-Cruz, Mariana; Serrano-Maldonado, Carlos Eduardo; González-Canto, Augusto; Peña-Montes, Carolina; Rodríguez-Sanoja, Romina; Sánchez, Sergio; Farrés, Amelia

2015-10-01

Pediococcus acidilactici ATCC 8042 is a lactic acid bacteria that inhibits pathogenic microorganisms such as Staphylococcus aureus through the production of two proteins with lytic activity, one of 110 kDa and the other of 99 kDa. The 99-kDa one has high homology to a putative peptidoglycan hydrolase (PGH) enzyme reported in the genome of P. acidilactici 7_4, where two different lytic domains have been identified but not characterized. The aim of this work was the biochemical characterization of the recombinant enzyme of 99 kDa. The enzyme was cloned and expressed successfully and retains its activity against Micrococcus lysodeikticus. It has a higher N-acetylglucosaminidase activity, but the N-acetylmuramoyl-L-alanine amidase can also be detected spectrophotometrically. The protein was then purified using gel filtration chromatography. Antibacterial activity showed an optimal pH of 6.0 and was stable between 5.0 and 7.0. The optimal temperature for activity was 60 °C, and all activity was lost after 1 h of incubation at 70 °C. The number of strains susceptible to the recombinant 99-kDa enzyme was lower than that susceptible to the mixture of the 110- and 99-kDa PGHs of P. acidilactici, a result that suggests synergy between these two enzymes. This is the first PGH from LAB that has been shown to possess two lytic sites. The results of this study will aid in the design of new antibacterial agents from natural origin that can combat foodborne disease and improve hygienic practices in the industrial sector.

Low levels of PRSS37 protein in sperm are associated with many cases of unexplained male infertility.

PubMed

Liu, Jianbing; Shen, Chunling; Fan, Weimin; Chen, Yan; Zhang, Aijun; Feng, Yun; Li, Zheng; Kuang, Ying; Wang, Zhugang

2016-11-01

PRSS37, a putative trypsin-like serine protease, is highly conserved during mammalian evolution as revealed by multiple sequence alignment. Mice deficient for Prss37 gene exhibit male infertility, but their mating behavior, spermatogenesis, sperm morphology, and motility remain unaffected, similar to a situation called unexplained male infertility (UMI) in men (human being). Here, we demonstrated that PRSS37 is restrictively expressed in human testis, where it is mainly located in the elongating and elongated spermatids during spermiogenesis as shown by immunohistochemical analysis of normal human testicular sections. In mature sperm, PRSS37 appears in the acrosome region and diminishes during acrosome reaction. Further examination reveals that PRSS37 contents in sperm from patients with UMI are dramatically lower than those in sperm from men with proven fertility or from sperm donors. Sperm with low PRSS37 contents exhibit abnormal activation of the proacrosin/acrosin system and premature proteolysis of ADAM2, which may impair the functional competence of human sperm in vivo However, the in vitro fertilization outcomes of sperm with low PRSS37 contents are not affected. Together, these data implicate an important role of PRSS37 for male fertility. PRSS37 can be used as a potential molecular biomarker for evaluating sperm fertilization capability in vivo but not in vitro. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Purification and characterization of the enzymes involved in nicotinamide adenine dinucleotide degradation by Penicillium brevicompactum NRC 829.

PubMed

Ali, Thanaa Hamed; El-Ghonemy, Dina Helmy

2016-06-01

The present study was conducted to investigate a new pathway for the degradation of nicotinamide adenine dinucleotide (NAD) by Penicillium brevicompactum NRC 829 extracts. Enzymes involved in the hydrolysis of NAD, i.e. alkaline phosphatase, aminohydrolase and glycohydrolase were determined. Alkaline phosphatase was found to catalyse the sequential hydrolysis of two phosphate moieties of NAD molecule to nicotinamide riboside plus adenosine. Adenosine was then deaminated by aminohydrolase to inosine and ammonia. While glycohydrolase catalyzed the hydrolysis of the nicotinamide-ribosidic bond of NAD+ to produce nicotinamide and ADP-ribose in equimolar amounts, enzyme purification through a 3-step purification procedure revealed the existence of two peaks of alkaline phosphatases, and one peak contained deaminase and glycohydrolase activities. NAD deaminase was purified to homogeneity as estimated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis with an apparent molecular mass of 91 kDa. Characterization and determination of some of NAD aminohydrolase kinetic properties were conducted due to its biological role in the regulation of cellular NAD level. The results also revealed that NAD did not exert its feedback control on nicotinamide amidase produced by P. brevicompactum.

Characterization, Genome Sequence, and Analysis of Escherichia Phage CICC 80001, a Bacteriophage Infecting an Efficient L-Aspartic Acid Producing Escherichia coli.

PubMed

Xu, Youqiang; Ma, Yuyue; Yao, Su; Jiang, Zengyan; Pei, Jiangsen; Cheng, Chi

2016-03-01

Escherichia phage CICC 80001 was isolated from the bacteriophage contaminated medium of an Escherichia coli strain HY-05C (CICC 11022S) which could produce L-aspartic acid. The phage had a head diameter of 45-50 nm and a tail of about 10 nm. The one-step growth curve showed a latent period of 10 min and a rise period of about 20 min. The average burst size was about 198 phage particles per infected cell. Tests were conducted on the plaques, multiplicity of infection, and host range. The genome of CICC 80001 was sequenced with a length of 38,810 bp, and annotated. The key proteins leading to host-cell lysis were phylogenetically analyzed. One protein belonged to class II holin, and the other two belonged to the endopeptidase family and N-acetylmuramoyl-L-alanine amidase family, respectively. The genome showed the sequence identity of 82.7% with that of Enterobacteria phage T7, and carried ten unique open reading frames. The bacteriophage resistant E. coli strain designated CICC 11021S was breeding and its L-aspartase activity was 84.4% of that of CICC 11022S.

Characterization of the Lytic Capability of a LysK-Like Endolysin, Lys-phiSA012, Derived from a Polyvalent Staphylococcus aureus Bacteriophage

PubMed Central

Nakamura, Tomohiro; Furusawa, Takaaki; Ohno, Hazuki; Takahashi, Hiromichi; Kitana, Junya; Usui, Masaru; Higuchi, Hidetoshi; Tamura, Yutaka

2018-01-01

Antibiotic-resistant bacteria (ARB) have spread widely and rapidly, with their increased occurrence corresponding with the increased use of antibiotics. Infections caused by Staphylococcus aureus have a considerable negative impact on human and livestock health. Bacteriophages and their peptidoglycan hydrolytic enzymes (endolysins) have received significant attention as novel approaches against ARB, including S. aureus. In the present study, we purified an endolysin, Lys-phiSA012, which harbors a cysteine/histidine-dependent amidohydrolase/peptidase (CHAP) domain, an amidase domain, and a SH3b cell wall binding domain, derived from a polyvalent S. aureus bacteriophage which we reported previously. We demonstrate that Lys-phiSA012 exhibits high lytic activity towards staphylococcal strains, including methicillin-resistant S. aureus (MRSA). Analysis of deletion mutants showed that only mutants possessing the CHAP and SH3b domains could lyse S. aureus, indicating that lytic activity of the CHAP domain depended on the SH3b domain. The presence of at least 1 mM Ca2+ and 100 µM Zn2+ enhanced the lytic activity of Lys-phiSA012 in a turbidity reduction assay. Furthermore, a minimum inhibitory concentration (MIC) assay showed that the addition of Lys-phiSA012 decreased the MIC of oxacillin. Our results suggest that endolysins are a promising approach for replacing current antimicrobial agents and may contribute to the proper use of antibiotics, leading to the reduction of ARB. PMID:29495305

Method for metabolizing carbazole in petroleum

DOEpatents

Kayser, Kevin J.; Kilbane, II, John J.

2005-09-13

A method for selective cleavage of C--N bonds genes that encode for at least one enzyme suitable for conversion of carbazole to 2-aminobiphenyl-2,3-diol are combined with a gene encoding an amidase suitable for selectively cleaving a C--N bond in 2-aminobiphenyl-2,3-diol, forming an operon that encodes for cleavage of both C--N bonds of said carbazole. The operon is inserted into a host culture which, in turn, is contacted with the carbazole, resulting in selective cleavage of both C--N bonds of the carbazole. Also disclosed is a new microorganism that expresses a carbazole degradation trait constitutively and a method for degrading carbazole employing this microorganism.

Murein Lytic Enzyme TgaA of Bifidobacterium bifidum MIMBb75 Modulates Dendritic Cell Maturation through Its Cysteine- and Histidine-Dependent Amidohydrolase/Peptidase (CHAP) Amidase Domain

PubMed Central

Zanoni, Ivan; Balzaretti, Silvia; Miriani, Matteo; Taverniti, Valentina; De Noni, Ivano; Presti, Ilaria; Stuknyte, Milda; Scarafoni, Alessio; Arioli, Stefania; Iametti, Stefania; Bonomi, Francesco; Mora, Diego; Karp, Matti; Granucci, Francesca

2014-01-01

Bifidobacteria are Gram-positive inhabitants of the human gastrointestinal tract that have evolved close interaction with their host and especially with the host's immune system. The molecular mechanisms underlying such interactions, however, are largely unidentified. In this study, we investigated the immunomodulatory potential of Bifidobacterium bifidum MIMBb75, a bacterium of human intestinal origin commercially used as a probiotic. Particularly, we focused our attention on TgaA, a protein expressed on the outer surface of MIMBb75's cells and homologous to other known bacterial immunoactive proteins. TgaA is a peptidoglycan lytic enzyme containing two active domains: lytic murein transglycosylase (LT) and cysteine- and histidine-dependent amidohydrolase/peptidase (CHAP). We ran immunological experiments stimulating dendritic cells (DCs) with the B. bifidum MIMBb75 and TgaA, with the result that both the bacterium and the protein activated DCs and triggered interleukin-2 (IL-2) production. In addition, we observed that the heterologous expression of TgaA in Bifidobacterium longum transferred to the bacterium the ability to induce IL-2. Subsequently, immunological experiments performed using two purified recombinant proteins corresponding to the single domains LT and CHAP demonstrated that the CHAP domain is the immune-reactive region of TgaA. Finally, we also showed that TgaA-dependent activation of DCs requires the protein CD14, marginally involves TRIF, and is independent of Toll-like receptor 4 (TLR4) and MyD88. In conclusion, our study suggests that the bacterial CHAP domain is a novel microbe-associated molecular pattern actively participating in the cross talk mechanisms between bifidobacteria and the host's immune system. PMID:24814791

A shifted repertoire of endocannabinoid genes in the zebrafish (Danio rerio).

PubMed

McPartland, J M; Glass, Michelle; Matias, Isabel; Norris, Ryan W; Kilpatrick, C William

2007-05-01

The zebrafish has served as a model organism for developmental biology. Sequencing its genome has expanded zebrafish research into physiology and drug-development testing. Several cannabinoid pharmaceuticals are in development, but expression of endocannabinoid receptors and enzymes remains unknown in this species. We conducted a bioinformatics analysis of the zebrafish genome using 17 human endocannabinoid genes as a reference set. Putative zebrafish orthologs were identified in filtered BLAST searches as reciprocal best hits. Orthology was confirmed by three in silico methods: phylogenetic testing, synteny analysis, and functional mapping. Zebrafish expressed orthologs of cannabinoid receptor 1, transient receptor potential channel vanilloid receptor 4, GPR55 receptor, fatty acid amide hydrolase 1, monoacylglycerol lipase, NAPE-selective phospholipase D, abhydrolase domain-containing protein 4, and diacylglycerol lipase alpha and beta; and paired paralogs of cannabinoid receptor 2, fatty acid amide hydrolase 2, peroxisome proliferator-activated receptor alpha, prostaglandin-endoperoxide synthase 2, and transient receptor potential cation channel subtype A1. Functional mapping suggested the orthologs of transient receptor potential vanilloid receptor 1 and peroxisome proliferator-activated receptor gamma lack specific amino acids critical for cannabinoid ligand binding. No orthologs of N-acylethanolamine acid amidase or protein tyrosine phosphatase, non-receptor type 22 were identified. In conclusion, the zebrafish genome expresses a shifted repertoire of endocannabinoid genes. In vitro analyses are warranted before using zebrafish for cannabinoid development testing.

Cloning, Characterization and Effect of TmPGRP-LE Gene Silencing on Survival of Tenebrio Molitor against Listeria monocytogenes Infection

PubMed Central

Tindwa, Hamisi; Patnaik, Bharat Bhusan; Kim, Dong Hyun; Mun, Seulgi; Jo, Yong Hun; Lee, Bok Luel; Lee, Yong Seok; Kim, Nam Jung; Han, Yeon Soo

2013-01-01

Peptidoglycan recognition proteins (PGRPs) are a family of innate immune molecules that recognize bacterial peptidoglycan. PGRP-LE, a member of the PGRP family, selectively binds to diaminopimelic acid (DAP)-type peptidoglycan to activate both the immune deficiency (Imd) and proPhenoloxidase (proPO) pathways in insects. A PGRP-LE-dependent induction of autophagy to control Listeria monocytogenes has also been reported. We identified and partially characterized a novel PGRP-LE homologue, from Tenebrio molitor and analyzed its functional role in the survival of the insect against infection by a DAP-type PGN containing intracellular pathogen, L. monocytogenes. The cDNA is comprised of an open reading frame (ORF) of 990 bp and encodes a polypeptide of 329 residues. TmPGRP-LE contains one PGRP domain, but lacks critical residues for amidase activity. Quantitative RT-PCR analysis showed a broad constitutive expression of the transcript at various stages of development spanning from larva to adult. RNAi mediated knockdown of the transcripts, followed by a challenge with L. monocytogenes, showed a significant reduction in survival rate of the larvae, suggesting a putative role of TmPGRP-LE in sensing and control of L. monocytogenes infection in T. molitor. These results implicate PGRP-LE as a defense protein necessary for survival of T. molitor against infection by L. monocytogenes. PMID:24240808

Cloning, characterization and effect of TmPGRP-LE gene silencing on survival of Tenebrio molitor against Listeria monocytogenes infection.

PubMed

Tindwa, Hamisi; Patnaik, Bharat Bhusan; Kim, Dong Hyun; Mun, Seulgi; Jo, Yong Hun; Lee, Bok Luel; Lee, Yong Seok; Kim, Nam Jung; Han, Yeon Soo

2013-11-14

Peptidoglycan recognition proteins (PGRPs) are a family of innate immune molecules that recognize bacterial peptidoglycan. PGRP-LE, a member of the PGRP family, selectively binds to diaminopimelic acid (DAP)-type peptidoglycan to activate both the immune deficiency (Imd) and proPhenoloxidase (proPO) pathways in insects. A PGRP-LE-dependent induction of autophagy to control Listeria monocytogenes has also been reported. We identified and partially characterized a novel PGRP-LE homologue, from Tenebrio molitor and analyzed its functional role in the survival of the insect against infection by a DAP-type PGN containing intracellular pathogen, L. monocytogenes. The cDNA is comprised of an open reading frame (ORF) of 990 bp and encodes a polypeptide of 329 residues. TmPGRP-LE contains one PGRP domain, but lacks critical residues for amidase activity. Quantitative RT-PCR analysis showed a broad constitutive expression of the transcript at various stages of development spanning from larva to adult. RNAi mediated knockdown of the transcripts, followed by a challenge with L. monocytogenes, showed a significant reduction in survival rate of the larvae, suggesting a putative role of TmPGRP-LE in sensing and control of L. monocytogenes infection in T. molitor. These results implicate PGRP-LE as a defense protein necessary for survival of T. molitor against infection by L. monocytogenes.

Elucidating the pH-Dependent Structural Transition of T7 Bacteriophage Endolysin.

PubMed

Sharma, Meenakshi; Kumar, Dinesh; Poluri, Krishna Mohan

2016-08-23

Bacteriophages are the most abundant and diverse biological entities on earth. Bacteriophage endolysins are unique peptidoglycan hydrolases and have huge potential as effective enzybiotics in various infectious models. T7 bacteriophage endolysin (T7L), also known as N-acetylmuramoyl-l-alanine amidase or T7 lysozyme, is a 17 kDa protein that lyses a range of Gram-negative bacteria by hydrolyzing the amide bond between N-acetylmuramoyl residues and the l-alanine of the peptidoglycan layer. Although the activity profiles of several of the T7 family members have been known for many years, the molecular basis for their pH-dependent differential activity is not clear. In this study, we explored the pH-induced structural, stability, and activity characteristics of T7L by applying a variety of biophysical techniques and protein nuclear magnetic resonance (NMR) spectroscopy. Our studies established a reversible structural transition of T7L below pH 6 and the formation of a partially denatured conformation at pH 3. This low-pH conformation is thermally stable and exposed its hydrophobic pockets. Further, NMR relaxation measurements and structural analysis unraveled that T7L is highly dynamic in its native state and a network of His residues are responsible for the observed pH-dependent conformational dynamics and transitions. As bacteriophage chimeric and engineered endolysins are being developed as novel therapeutics against multiple drug resistance pathogens, we believe that our results are of great help in designing these entities as broadband antimicrobial and/or antibacterial agents.

The proteolytic system of pineapple stems revisited: Purification and characterization of multiple catalytically active forms.

PubMed

Matagne, André; Bolle, Laetitia; El Mahyaoui, Rachida; Baeyens-Volant, Danielle; Azarkan, Mohamed

2017-06-01

Crude pineapple proteases extract (aka stem bromelain; EC 3.4.22.4) is an important proteolytic mixture that contains enzymes belonging to the cysteine proteases of the papain family. Numerous studies have been reported aiming at the fractionation and characterization of the many molecular species present in the extract, but more efforts are still required to obtain sufficient quantities of the various purified protease forms for detailed physicochemical, enzymatic and structural characterization. In this work, we describe an efficient strategy towards the purification of at least eight enzymatic forms. Thus, following rapid fractionation on a SP-Sepharose FF column, two sub-populations with proteolytic activity were obtained: the unbound (termed acidic) and bound (termed basic) bromelain fractions. Following reversible modification with monomethoxypolyethylene glycol (mPEG), both fractions were further separated on Q-Sepharose FF and SP-Sepharose FF, respectively. This procedure yielded highly purified molecular species, all titrating ca. 1 mol of thiol group per mole of enzyme, with distinct biochemical properties. N-terminal sequencing allowed identifying at least eight forms with proteolytic activity. The basic fraction contained previously identified species, i.e. basic bromelain forms 1 and 2, ananain forms 1 and 2, and comosain (MEROPS identifier: C01.027). Furthermore, a new proteolytic species, showing similarities with basic bomelain forms 1 and 2, was discovered and termed bromelain form 3. The two remaining species were found in the acidic bromelain fraction and were arbitrarily named acidic bromelain forms 1 and 2. Both, acidic bromelain forms 1, 2 and basic bromelain forms 1, 2 and 3 are glycosylated, while ananain forms 1 and 2, and comosain are not. The eight protease forms display different amidase activities against the various substrates tested, namely small synthetic chromogenic compounds (DL-BAPNA and Boc-Ala-Ala-Gly-pNA), fluorogenic compounds

The First Paenibacillus larvae Bacteriophage Endolysin (PlyPl23) with High Potential to Control American Foulbrood.

PubMed

Oliveira, Ana; Leite, Marta; Kluskens, Leon D; Santos, Sílvio B; Melo, Luís D R; Azeredo, Joana

2015-01-01

Endolysins, which are peptidoglycan-degrading enzymes expressed during the terminal stage of the reproduction cycle of bacteriophages, have great potential to control Gram-positive pathogens. This work describes the characterization of a novel endolysin (PlyPl23) encoded on the genome of Paenibacillus larvae phage phiIBB_Pl23 with high potential to control American foulbrood. This bacterial disease, caused by P. larvae, is widespread in North America and Europe and causes important economic losses in apiculture. The restriction to antibiotic residues in honey imposed by the EU legislation hinders its therapeutic use to combat American foulbrood and enforces the development of alternative antimicrobial methods. The new endolysin described herein has an N-acetylmuramoyl-L-alanine amidase catalytic domain and exhibits a broad-spectrum activity against common P. larvae genotypes. Moreover, the enzyme displays high antimicrobial activity in a range of pH that matches environmental conditions (pH between 5.0 and 7.0), showing its feasible application in the field. At pH 7.0, a concentration of 0.2 μM of enzyme was enough to lyse 104 CFU.mL-1 of P. larvae in no more than 2 h. The presence of sucrose and of the substances present in the larvae gut content did not affect the enzyme activity. Interestingly, an increase of activity was observed when PlyPl23 was previously incubated in royal jelly. Furthermore, in vivo safety evaluation assays demonstrated that this enzyme is not toxic to the bee larvae. The present work describes for the first time an endolysin encoded in a P. larvae phage that presents high potential to integrate a commercial product to control the problematic American foulbrood.

Formation and hydrolysis of amide bonds by lipase A from Candida antarctica; exceptional features.

PubMed

Liljeblad, Arto; Kallio, Pauli; Vainio, Marita; Niemi, Jarmo; Kanerva, Liisa T

2010-02-21

Various commercial lyophilized and immobilized preparations of lipase A from Candida antarctica (CAL-A) were studied for their ability to catalyze the hydrolysis of amide bonds in N-acylated alpha-amino acids, 3-butanamidobutanoic acid (beta-amino acid) and its ethyl ester. The activity toward amide bonds is highly untypical of lipases, despite the close mechanistic analogy to amidases which normally catalyze the corresponding reactions. Most CAL-A preparations cleaved amide bonds of various substrates with high enantioselectivity, although high variations in substrate selectivity and catalytic rates were detected. The possible role of contaminant protein species on the hydrolytic activity toward these bonds was studied by fractionation and analysis of the commercial lyophilized preparation of CAL-A (Cat#ICR-112, Codexis). In addition to minor impurities, two equally abundant proteins were detected, migrating on SDS-PAGE a few kDa apart around the calculated size of CAL-A. Based on peptide fragment analysis and sequence comparison both bands shared substantial sequence coverage with CAL-A. However, peptides at the C-terminal end constituting a motile domain described as an active-site flap were not identified in the smaller fragment. Separated gel filtration fractions of the two forms of CAL-A both catalyzed the amide bond hydrolysis of ethyl 3-butanamidobutanoate as well as the N-acylation of methyl pipecolinate. Hydrolytic activity towards N-acetylmethionine was, however, solely confined to the fractions containing the truncated form of CAL-A. These fractions were also found to contain a trace enzyme impurity identified in sequence analysis as a serine carboxypeptidase. The possible role of catalytic impurities versus the function of CAL-A in amide bond hydrolysis is further discussed in the paper.

Targeted Mutagenesis of the Mycobacterium smegmatis mca Gene, Encoding a Mycothiol-Dependent Detoxification Protein

PubMed Central

Rawat, Mamta; Uppal, Mandeep; Newton, Gerald; Steffek, Micah; Fahey, Robert C.; Av-Gay, Yossef

2004-01-01

Mycothiol (MSH), a functional analogue of glutathione (GSH) that is found exclusively in actinomycetes, reacts with electrophiles and toxins to form MSH-toxin conjugates. Mycothiol S-conjugate amidase (Mca) then catalyzes the hydrolysis of an amide bond in the S conjugates, producing a mercapturic acid of the toxin, which is excreted from the bacterium, and glucosaminyl inositol, which is recycled back to MSH. In this study, we have generated and characterized an allelic exchange mutant of the mca gene of Mycobacterium smegmatis. The mca mutant accumulates the S conjugates of the thiol-specific alkylating agent monobromobimane and the antibiotic rifamycin S. Introduction of M. tuberculosis mca epichromosomally or introduction of M. smegmatis mca integratively resulted in complementation of Mca activity and reduced levels of S conjugates. The mutation in mca renders the mutant strain more susceptible to electrophilic toxins, such as N-ethylmalemide, iodoacetamide, and chlorodinitrobenzene, and to several oxidants, such as menadione and plumbagin. Additionally we have shown that the mca mutant is also more susceptible to the antituberculous antibiotic streptomycin. Mutants disrupted in genes belonging to MSH biosynthesis are also more susceptible to streptomycin, providing further evidence that Mca detoxifies streptomycin in the mycobacterial cell in an MSH-dependent manner. PMID:15342574

Peptidoglycan Hydrolases of Escherichia coli

PubMed Central

van Heijenoort, Jean

2011-01-01

Summary: The review summarizes the abundant information on the 35 identified peptidoglycan (PG) hydrolases of Escherichia coli classified into 12 distinct families, including mainly glycosidases, peptidases, and amidases. An attempt is also made to critically assess their functions in PG maturation, turnover, elongation, septation, and recycling as well as in cell autolysis. There is at least one hydrolytic activity for each bond linking PG components, and most hydrolase genes were identified. Few hydrolases appear to be individually essential. The crystal structures and reaction mechanisms of certain hydrolases having defined functions were investigated. However, our knowledge of the biochemical properties of most hydrolases still remains fragmentary, and that of their cellular functions remains elusive. Owing to redundancy, PG hydrolases far outnumber the enzymes of PG biosynthesis. The presence of the two sets of enzymes acting on the PG bonds raises the question of their functional correlations. It is difficult to understand why E. coli keeps such a large set of PG hydrolases. The subtle differences in substrate specificities between the isoenzymes of each family certainly reflect a variety of as-yet-unidentified physiological functions. Their study will be a far more difficult challenge than that of the steps of the PG biosynthesis pathway. PMID:22126997

Cloning and analysis of peptidoglycan recognition protein-LC and immune deficiency from the diamondback moth, Plutella xylostella.

PubMed

Zhan, Ming-Yue; Yang, Pei-Jin; Rao, Xiang-Jun

2018-02-01

Peptidoglycan (PGN) exists in both Gram-negative and Gram-positive bacteria as a component of the cell wall. PGN is an important target to be recognized by the innate immune system of animals. PGN recognition proteins (PGRP) are responsible for recognizing PGNs. In Drosophila melanogaster, PGRP-LC and IMD (immune deficiency) are critical for activating the Imd pathway. Here, we report the cloning and analysis of PGRP-LC and IMD (PxPGRP-LC and PxIMD) from diamondback moth, Plutella xylostella (L.), the insect pest of cruciferous vegetables. PxPGRP-LC gene consists of six exons encoding a polypeptide of 308 amino acid residues with a transmembrane region and a PGRP domain. PxIMD cDNA encodes a polypeptide of 251 amino acid residues with a death domain. Sequence comparisons indicate that they are characteristic of Drosophila PGRP-LC and IMD homologs. PxPGRP-LC and PxIMD were expressed in various tissues and developmental stages. Their mRNA levels were affected by bacterial challenges. The PGRP domain of PxPGRP-LC lacks key residues for the amidase activity, but it can recognize two types of PGNs. Overexpression of full-length and deletion mutants in Drosophila S2 cells induced expression of some antimicrobial peptide genes. These results indicate that PxPGRP-LC and PxIMD may be involved in the immune signaling of P. xylostella. This study provides a foundation for further studies of the immune system of P. xylostella. © 2017 Wiley Periodicals, Inc.

Determination of the mitigating effect of colon-specific bioreversible codrugs of mycophenolic acid and aminosugars in an experimental colitis model in Wistar rats

PubMed Central

Chopade, Shakuntala Santosh; Dhaneshwar, Suneela Sunil

2018-01-01

AIM To design colon-targeted codrugs of mycophenolic acid (MPA) and aminosugars as a safer option to mycophenolate mofetil (MMF) in the management of inflammatory bowel disease. METHODS Codrugs were synthesized by coupling MPA with aminosugars (D-glucosamine and D-galactosamine) using EDCI coupling. The structures were confirmed by infrared radiation, nuclear magnetic resonance, mass spectroscopy and elemental analysis. The release profile of codrugs was extensively studied in aqueous buffers, upper gastrointestinal homogenates, faecal matter and caecal homogenates (in vitro) and rat blood (in vitro). Anti-colitic activity was assessed in 2,4,6-trinitrobezenesulfonic acid-induced colitis in Wistar rats by the estimation of various demarcating parameters. Statistical evaluation was performed by applying one-way and two-way ANOVA when compared with the disease control. RESULTS The prodrugs resisted activation in HCl buffer (pH 1.2) and stomach homogenates of rats with negligible hydrolysis in phosphate buffer (pH 7.4) and intestinal homogenates. Incubation with colon homogenates (in vitro) produced 76% to 89% release of MPA emphasizing colon-specific activation of codrugs and the release of MPA and aminosugars at the site of action. In the in vitro studies, the prodrug of MPA with D-glucosamine (MGLS) was selected which resulted in 68% release of MPA in blood. in vitro studies on MGLS revealed its colon-specific activation after a lag time of 8 h which could be ascribed to the hydrolytic action of N-acyl amidases found in the colon. The synthesized codrugs markedly diminished disease activity score and revived the disrupted architecture of the colon that was comparable to MMF but superior to MPA. CONCLUSION The significant attenuating effect of prodrugs and individual aminosugars on colonic inflammation proved that the rationale of the codrug approach is valid. PMID:29563754

A Catalog of Proteins Expressed in the AG Secreted Fluid during the Mature Phase of the Chinese Mitten Crabs (Eriocheir sinensis)

PubMed Central

He, Lin; Li, Qing; Liu, Lihua; Wang, Yuanli; Xie, Jing; Yang, Hongdan; Wang, Qun

2015-01-01

The accessory gland (AG) is an important component of the male reproductive system of arthropods, its secretions enhance fertility, some AG proteins bind to the spermatozoa and affect its function and properties. Here we report the first comprehensive catalog of the AG secreted fluid during the mature phase of the Chinese mitten crab (Eriocheir sinensis). AG proteins were separated by one-dimensional gel electrophoresis and analyzed by reverse phase high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). Altogether, the mass spectra of 1173 peptides were detected (1067 without decoy and contaminants) which allowed for the identification of 486 different proteins annotated upon the NCBI database (http://www.ncbi.nlm.nih.gov/) and our transcritptome dataset. The mass spectrometry proteomics data have been deposited at the ProteomeXchange with identifier PXD000700. An extensive description of the AG proteome will help provide the basis for a better understanding of a number of reproductive mechanisms, including potentially spermatophore breakdown, dynamic functional and morphological changes in sperm cells and sperm acrosin enzyme vitality. Thus, the comprehensive catalog of proteins presented here can serve as a valuable reference for future studies of sperm maturation and regulatory mechanisms involved in crustacean reproduction. PMID:26305468

A Novel Method for Relative Quantitation of N-Glycans by Isotopic Labeling Using 18O-Water

PubMed Central

Tao, Shujuan; Orlando, Ron

2014-01-01

Quantitation is an essential aspect of comprehensive glycomics study. Here, a novel isotopic-labeling method is described for N-glycan quantitation using 18O-water. The incorporation of the 18O-labeling into the reducing end of N-glycans is simply and efficiently achieved during peptide-N4-(N-acetyl-β-glucosaminyl) asparagine amidase F release. This process provides a 2-Da mass difference compared with the N-glycans released in 16O-water. A mathematical calculation method was also developed to determine the 18O/16O ratios from isotopic peaks. Application of this method to several standard glycoprotein mixtures and human serum demonstrated that this method can facilitate the relative quantitation of N-glycans over a linear dynamic range of two orders, with high accuracy and reproducibility. PMID:25365792

Bacterial cell motility of Burkholderia gut symbiont is required to colonize the insect gut.

PubMed

Lee, Jun Beom; Byeon, Jin Hee; Jang, Ho Am; Kim, Jiyeun Kate; Yoo, Jin Wook; Kikuchi, Yoshitomo; Lee, Bok Luel

2015-09-14

We generated a Burkholderia mutant, which is deficient of an N-acetylmuramyl-l-alanine amidase, AmiC, involved in peptidoglycan degradation. When non-motile ΔamiC mutant Burkholderia cells harboring chain form were orally administered to Riptortus insects, ΔamiC mutant cells were unable to establish symbiotic association. But, ΔamiC mutant complemented with amiC gene restored in vivo symbiotic association. ΔamiC mutant cultured in minimal medium restored their motility with single-celled morphology. When ΔamiC mutant cells harboring single-celled morphology were administered to the host insect, this mutant established normal symbiotic association, suggesting that bacterial motility is essential for the successful symbiosis between host insect and Burkholderia symbiont. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Distribution, industrial applications, and enzymatic synthesis of D-amino acids.

PubMed

Gao, Xiuzhen; Ma, Qinyuan; Zhu, Hailiang

2015-04-01

D-Amino acids exist widely in microbes, plants, animals, and food and can be applied in pharmaceutical, food, and cosmetics. Because of their widespread applications in industry, D-amino acids have recently received more and more attention. Enzymes including D-hydantoinase, N-acyl-D-amino acid amidohydrolase, D-amino acid amidase, D-aminopeptidase, D-peptidase, L-amino acid oxidase, D-amino acid aminotransferase, and D-amino acid dehydrogenase can be used for D-amino acids synthesis by kinetic resolution or asymmetric amination. In this review, the distribution, industrial applications, and enzymatic synthesis methods are summarized. And, among all the current enzymatic methods, D-amino acid dehydrogenase method not only produces D-amino acid by a one-step reaction but also takes environment and atom economics into consideration; therefore, it is deserved to be paid more attention.

Purification and characterization of an amidohydrolase for N4-long-chain fatty acyl derivatives of 1-beta-D-arabinofuranosylcytosine from mouse liver microsomes.

PubMed

Hori, K; Tsuruo, T; Tsukagoshi, S; Sakurai, Y

1984-03-01

N4-Long-chain fatty acyl-1-beta-D-arabinofuranosylcytosine amidohydrolase, a metabolizing enzyme for N4-acyl derivatives of 1-beta-D-arabinofuranosylcytosine with long-chain fatty acids, was purified from mouse liver microsomes. The purification was accomplished by solubilization of liver microsomes with Triton X-100, diethylaminoethyl cellulose chromatography, gel filtrations, hydroxyapatite chromatography, and concanavalin A:Sepharose chromatography. On sodium dodecyl sulfate:polyacrylamide gel electrophoresis, the purified enzyme preparation produced a single protein band with a molecular weight of 54,000. The enzyme had an optimal pH of 9.0, and the Michaelis constant for N4-palmitoyl-1-beta-D-arabinofuranosylcytosine was 67 microM. The thiols such as dithiothreitol or 2-mercaptoethanol stabilized the enzyme and stimulated its activity. p-Chloromercuribenzoate, N-ethylmaleimide, diisopropylfluorophosphate, and phenylmethylsulfonyl fluoride strongly inhibited the reaction. Bovine serum albumin markedly stimulated the enzyme activity, whereas detergents such as Triton X-100, deoxycholate, and sodium dodecyl sulfate had little effect. The enzyme did not require monovalent or divalent cations. Among the series of N4-acyl derivatives of 1-beta-D-arabinofuranosylcytosine with different chain lengths of acyl residues, the purified enzyme preferentially hydrolyzed the derivatives with long-chain fatty acids (C12 to C18), and N4-palmitoyl-1-beta-D-arabinofuranosylcytosine was the most susceptible. The purified enzyme was inactive on various N-acylamino acids, amides, oligopeptides, proteins, N-acylsphingosines (ceramides), triglyceride, lecithin, and lysolecithin. These results suggest that N4-long-chain fatty acyl-1-beta-D-arabinofuranosylcytosine amidohydrolase may be a new type of linear amidase.

Detoxification of toxins by bacillithiol in Staphylococcus aureus

PubMed Central

Newton, Gerald L.; Fahey, Robert C.

2012-01-01

Bacillithiol (BSH), an α-anomeric glycoside of l-cysteinyl-d-glucosaminyl-l-malate, is a major low-molecular-mass thiol found in bacteria such as Bacillus sp., Staphylococcus aureus and Deinococcus radiodurans. Like other low-molecular-mass thiols such as glutathione and mycothiol, BSH is likely to be involved in protection against environmental toxins including thiol-reactive antibiotics. We report here a BSH-dependent detoxification mechanism in S. aureus. When S. aureus Newman strain was treated with monobromobimane and monochlorobimane, the cellular BSH was converted to the fluorescent S-conjugate BS-bimane. A bacillithiol conjugate amidase activity acted upon the BS-bimane to produce Cys-bimane, which was then acetylated by an N-acetyltransferase to generate N-acetyl-Cys-bimane, a mercapturic acid. An S. aureus mutant lacking BSH did not produce mercapturic acid when treated with monobromobimane and monochlorobimane, confirming the involvement of bacillithiol. Furthermore, treatment of S. aureus Newman with rifamycin, the parent compound of the first-line anti-tuberculosis drug, rifampicin, indicated that this thiol-reactive antibiotic is also detoxified in a BSH-dependent manner, since mercapturic acids of rifamycin were observed in the culture medium. These data indicate that toxins and thiol-reactive antibiotics are detoxified to less potent mercapturic acids in a BSH-dependent manner and then exported out of the cell in S. aureus. PMID:22262099

Microbial nitrilases: versatile, spiral forming, industrial enzymes.

PubMed

Thuku, R N; Brady, D; Benedik, M J; Sewell, B T

2009-03-01

The nitrilases are enzymes that convert nitriles to the corresponding acid and ammonia. They are members of a superfamily, which includes amidases and occur in both prokaryotes and eukaryotes. The superfamily is characterized by having a homodimeric building block with a alpha beta beta alpha-alpha beta beta alpha sandwich fold and an active site containing four positionally conserved residues: cys, glu, glu and lys. Their high chemical specificity and frequent enantioselectivity makes them attractive biocatalysts for the production of fine chemicals and pharmaceutical intermediates. Nitrilases are also used in the treatment of toxic industrial effluent and cyanide remediation. The superfamily enzymes have been visualized as dimers, tetramers, hexamers, octamers, tetradecamers, octadecamers and variable length helices, but all nitrilase oligomers have the same basic dimer interface. Moreover, in the case of the octamers, tetradecamers, octadecamers and the helices, common principles of subunit association apply. While the range of industrially interesting reactions catalysed by this enzyme class continues to increase, research efforts are still hampered by the lack of a high resolution microbial nitrilase structure which can provide insights into their specificity, enantioselectivity and the mechanism of catalysis. This review provides an overview of the current progress in elucidation of structure and function in this enzyme class and emphasizes insights that may lead to further biotechnological applications.

Identification of NH4+-regulated genes of Herbaspirillum seropedicae by random insertional mutagenesis.

PubMed

Schwab, Stefan; Ramos, Humberto J; Souza, Emanuel M; Pedrosa, Fábio O; Yates, Marshall G; Chubatsu, Leda S; Rigo, Liu U

2007-05-01

Random mutagenesis using transposons with promoterless reporter genes has been widely used to examine differential gene expression patterns in bacteria. Using this approach, we have identified 26 genes of the endophytic nitrogen-fixing bacterium Herbaspirillum seropedicae regulated in response to ammonium content in the growth medium. These include nine genes involved in the transport of nitrogen compounds, such as the high-affinity ammonium transporter AmtB, and uptake systems for alternative nitrogen sources; nine genes coding for proteins responsible for restoring intracellular ammonium levels through enzymatic reactions, such as nitrogenase, amidase, and arginase; and a third group includes metabolic switch genes, coding for sensor kinases or transcription regulation factors, whose role in metabolism was previously unknown. Also, four genes identified were of unknown function. This paper describes their involvement in response to ammonium limitation. The results provide a preliminary profile of the metabolic response of Herbaspirillum seropedicae to ammonium stress.

Self-assembling choline mimicks with enhanced binding affinities to C-LytA protein

PubMed Central

Shi, Yang; Zhou, Hao; Zhang, Xiaoli; Wang, Jingyu; Long, Jiafu; Yang, Zhimou; Ding, Dan

2014-01-01

Streptococcus pneumoniae (pneumococcus) causes multiple illnesses in humans. Exploration of effective inhibitors with multivalent attachment sites for choline-binding modules is of great importance to reduce the pneumococcal virulence. In this work, we successfully developed two self-assembling choline mimicks, Ada-GFFYKKK' and Nap-GFFYKKK', which have the abilities to self-assemble into nanoparticles and nanofibers, respectively, yielding multivalent architectures. Additionally, the best characterized choline-binding module, C-terminal moiety of the pneumococcal cell-wall amidase LytA (C-LytA) was also produced with high purity. The self-assembling Ada-GFFYKKK' and Nap-GFFYKKK' show strong interactions with C-LytA, which possess much higher association constant values to the choline-binding modules as compared to the individual peptide Fmoc-K'. This study thus provides a self-assembly approach to yield inhibitors that are very promising for reducing the pneumococcal virulence. PMID:25315737

Diversity of Innate Immune Recognition Mechanism for Bacterial Polymeric meso-Diaminopimelic Acid-type Peptidoglycan in Insects

PubMed Central

Yu, Yang; Park, Ji-Won; Kwon, Hyun-Mi; Hwang, Hyun-Ok; Jang, In-Hwan; Masuda, Akiko; Kurokawa, Kenji; Nakayama, Hiroshi; Lee, Won-Jae; Dohmae, Naoshi; Zhang, Jinghai; Lee, Bok Luel

2010-01-01

In Drosophila, the synthesis of antimicrobial peptides in response to microbial infections is under the control of the Toll and immune deficiency (Imd) signaling pathway. The Toll signaling pathway responds mainly to the lysine-type peptidoglycan of Gram-positive bacteria and fungal β-1,3-glucan, whereas the Imd pathway responds to the meso-diaminopimelic acid (DAP)-type peptidoglycan of Gram-negative bacteria and certain Gram-positive bacilli. Recently we determined the activation mechanism of a Toll signaling pathway biochemically using a large beetle, Tenebrio molitor. However, DAP-type peptidoglycan recognition mechanism and its signaling pathway are still unclear in the fly and beetle. Here, we show that polymeric DAP-type peptidoglycan, but not its monomeric form, formed a complex with Tenebrio peptidoglycan recognition protein-SA, and this complex activated the three-step proteolytic cascade to produce processed Spätzle, a Toll receptor ligand, and induced Drosophila defensin-like antimicrobial peptide in Tenebrio larvae similarly to polymeric lysine-type peptidoglycan. Monomeric DAP-type peptidoglycan induced Drosophila diptericin-like antimicrobial peptide in Tenebrio hemocytes. In addition, both polymeric and monomeric DAP-type peptidoglycans induced expression of Tenebrio peptidoglycan recognition protein-SC2, which is DAP-type peptidoglycan-selective N-acetylmuramyl-l-alanine amidase that functions as a DAP-type peptidoglycan scavenger, appearing to function as a negative regulator of the DAP-type peptidoglycan signaling by cleaving DAP-type peptidoglycan in Tenebrio larvae. Taken together, these results demonstrate that molecular recognition mechanism for polymeric DAP-type peptidoglycan is different between Tenebrio larvae and Drosophila adults, providing biochemical evidences of biological diversity of innate immune responses in insects. PMID:20702416

Synthesis, properties, and application in peptide chemistry of a magnetically separable and reusable biocatalyst

NASA Astrophysics Data System (ADS)

Liria, Cleber W.; Ungaro, Vitor A.; Fernandes, Raphaella M.; Costa, Natália J. S.; Marana, Sandro R.; Rossi, Liane M.; Machini, M. Teresa

2014-11-01

Enzyme-catalyzed chemical processes are selective, very productive, and generate little waste. Nevertheless, they may be optimized using enzymes bound to solid supports, which are particularly important for protease-mediated reactions since proteases undergo fast autolysis in solution. Magnetic nanoparticles are suitable supports for this purpose owing to their high specific surface area and to be easily separated from reaction media. Here we describe the immobilization of bovine α-chymotrypsin (αCT) on silica-coated superparamagnetic nanoparticles (Fe3O4@silica) and the characterization of the enzyme-nanoparticle hybrid (Fe3O4@silica-αCT) in terms of protein content, properties, recovery from reaction media, application, and reuse in enzyme-catalyzed peptide synthesis. The results revealed that (i) full acid hydrolysis of the immobilized protease followed by amino acid analysis of the hydrolyzate is a reliable method to determine immobilization yield; (ii) despite showing lower amidase activity and a lower K cat/ K m value for a specific substrate than free αCT, the immobilized enzyme is chemically and thermally more stable, magnetically recoverable from reaction media, and can be consecutively reused for ten cycles to catalyze the amide bond hydrolysis and ester hydrolysis of the protected dipeptide Z-Ala-Phe-OMe. Altogether, these properties indicate the potential of Fe3O4@silica-αCT to act as an efficient, suitably stable, and reusable catalyst in amino acid, peptide, and protein chemistry as well as in proteomic studies.

LytN, a Murein Hydrolase in the Cross-wall Compartment of Staphylococcus aureus, Is Involved in Proper Bacterial Growth and Envelope Assembly*

PubMed Central

Frankel, Matthew B.; Hendrickx, Antoni P. A.; Missiakas, Dominique M.; Schneewind, Olaf

2011-01-01

Cell cycle progression for the spherical microbe Staphylococcus aureus requires the coordinated synthesis and remodeling of peptidoglycan. The majority of these rearrangements takes place at the mid-cell, in a compartment designated the cross-wall. Secreted polypeptides endowed with a YSIRK-G/S signal peptide are directly delivered to the cross-wall compartment. One such YSIRK-containing protein is the murein hydrolase LytN. lytN mutations precipitate structural damage to the cross-wall and interfere with staphylococcal growth. Overexpression of lytN also affects growth and triggers rupture of the cross-wall. The lytN phenotype can be reversed by the controlled expression of lytN but not by adding purified LytN to staphylococcal cultures. LytN harbors LysM and CHAP domains, the latter of which functions as both an N-acetylmuramoyl-l-alanine amidase and d-alanyl-glycine endopeptidase. Thus, LytN secretion into the cross-wall promotes peptidoglycan separation and completion of the staphylococcal cell cycle. PMID:21784864

Phenotypic and genotypic characterization of peptidoglycan hydrolases of Lactobacillus sakei

PubMed Central

Najjari, Afef; Amairi, Houda; Chaillou, Stéphane; Mora, Diego; Boudabous, Abdellatif; Zagorec, Monique; Ouzari, Hadda

2015-01-01

Lactobacillus sakei, a lactic acid bacterium naturally found in fresh meat and sea products, is considered to be one of the most important bacterial species involved in meat fermentation and bio-preservation. Several enzymes of Lb. sakei species contributing to microbial safeguarding and organoleptic properties of fermented-meat were studied. However, the specific autolytic mechanisms and associated enzymes involved in Lb. sakei are not well understood. The autolytic phenotype of 22 Lb. sakei strains isolated from Tunisian meat and seafood products was evaluated under starvation conditions, at pH 6.5 and 8.5, and in the presence of different carbon sources. A higher autolytic rate was observed when cells were grown in the presence of glucose and incubated at pH 6.5. Almost all strains showed high resistance to mutanolysin, indicating a minor role of muramidases in Lb. sakei cell lysis. Using Micrococcus lysodeikticus cells as a substrate in activity gels zymogram, peptidoglycan hydrolase (PGH) patterns for all strains was characterized by two lytic bands of ∼80 (B1) and ∼70 kDa (B2), except for strain BMG.167 which harbored two activity signals at a lower MW. Lytic activity was retained in high salt and in acid/basic conditions and was active toward cells of Lb. sakei, Listeria monocytogenes, Listeria ivanovii and Listeria innocua. Analysis of five putative PGH genes found in the Lb. sakei 23 K model strain genome, indicated that one gene, lsa1437, could encode a PGH (N-acetylmuramoyl-L-alanine amidase) containing B1 and B2 as isoforms. According to this hypothesis, strain BMG.167 showed an allelic version of lsa1437 gene deleted of one of the five LysM domains, leading to a reduction in the MW of lytic bands and the high autolytic rate of this strain. Characterization of autolytic phenotype of Lb. sakei should expand the knowledge of their role in fermentation processes where they represent the dominant species. PMID:26843981

Cryopreservation of adult cervid testes.

PubMed

Pothana, Lavanya; Devi, Lalitha; Goel, Sandeep

2017-02-01

Several species of cervids are currently classified as threatened or endangered due to a rapid decline in their populations. Sperm cryopreservation, in association with assisted reproductive technologies, can find application for the conservation of endangered cervids. In cases of unsuccessful sperm retrieval through other means prior to the death of the animal, adult testis is the only source of sperm. Recovery of viable sperm from adult testes depends on the effective preservation of testicular tissues through optimization of cryopreservation protocols. The present study evaluated combinations of 10% dimethyl sulfoxide (DMSO) with 0% or 80% fetal bovine serum (FBS) and 20% DMSO with 0 or 20% FBS for the cryopreservation of testicular tissues of three adult cervids using uncontrolled slow freezing protocol. The cryopreserved testis was compared to chilled tissue without cryoprotectants. Results revealed that testicular tissues of barking deer cryopreserved in 20% DMSO (D20) had all the analyzed 7 parameters (number of TNP1-, PRM2 and acrosin-expressing cells/tubule and, the number of viable, morphologically normal, acrosome intact, Annexin V-negative sperm) comparable to the chilled testis. However, testicular tissues of sambhar and hog deer cryopreserved only in D20S20 had 5 of 7 parameters comparable to the chilled testis. In conclusion, D20 is acceptable for cryopreservation of barking deer and D20S20 for sambar and hog deer testes. Copyright © 2016 Elsevier Inc. All rights reserved.

Identification of a novel gene cluster in the upstream region of the S-layer gene sbpA involved in cell wall metabolism of Lysinibacillus sphaericus CCM 2177 and characterization of the recombinantly produced autolysin and pyruvyl transferase.

PubMed

Pleschberger, Magdalena; Hildner, Florian; Rünzler, Dominik; Gelbmann, Nicola; Mayer, Harald F; Sleytr, Uwe B; Egelseer, Eva M

2013-05-01

The S-layer protein SbpA of Lysinibacillus sphaericus CCM 2177 assembles into a square (p4) lattice structure and recognizes a pyruvylated secondary cell wall polymer (SCWP) as the proper anchoring structure to the rigid cell wall layer. Sequencing of 8,004 bp in the 5'-upstream region of the S-layer gene sbpA led to five ORFs-encoding proteins involved in cell wall metabolism. After cloning and heterologous expression of ORF1 and ORF5 in Escherichia coli, the recombinant autolysin rAbpA and the recombinant pyruvyl transferase rCsaB were isolated, purified, and correct folding was confirmed by circular dichroism. Although rAbpA encoded by ORF1 showed amidase activity, it could attack whole cells of Ly. sphaericus CCM 2177 only after complete extraction of the S-layer lattice. Despite the presence of three S-layer-homology motifs on the N-terminal part, rAbpA did not show detectable affinity to peptidoglycan-containing sacculi, nor to isolated SCWP. As the molecular mass of the autolysin lies above the molecular exclusion limit of the S-layer, AbpA is obviously trapped within the rigid cell wall layer by the isoporous protein lattice. Immunogold-labeling of ultrathin-sectioned whole cells of Ly. sphaericus CCM 2177 with a polyclonal rabbit antiserum raised against rCsaB encoded by ORF5, and cell fractionation experiments demonstrated that the pyruvyl transferase was located in the cytoplasm, but not associated with cell envelope components including the plasma membrane. In enzymatic assays, rCsaB clearly showed pyruvyl transferase activity. By using RT-PCR, specific transcripts for each ORF could be detected. Cotranscription could be confirmed for ORF2 and ORF3.

Evolutionary history, structural features and biochemical diversity of the NlpC/P60 superfamily of enzymes.

PubMed

Anantharaman, Vivek; Aravind, L

2003-01-01

Peptidoglycan is hydrolyzed by a diverse set of enzymes during bacterial growth, development and cell division. The N1pC/P60 proteins define a family of cell-wall peptidases that are widely represented in various bacterial lineages. Currently characterized members are known to hydrolyze D-gamma-glutamyl-meso-diaminopimelate or N-acetylmuramate-L-alanine linkages. Detailed analysis of the N1pC/P60 peptidases showed that these proteins define a large superfamily encompassing several diverse groups of proteins. In addition to the well characterized P60-like proteins, this superfamily includes the AcmB/LytN and YaeF/YiiX families of bacterial proteins, the amidase domain of bacterial and kinetoplastid glutathionylspermidine synthases (GSPSs), and several proteins from eukaryotes, phages, poxviruses, positive-strand RNA viruses, and certain archaea. The eukaryotic members include lecithin retinol acyltransferase (LRAT), nematode developmental regulator Egl-26, and candidate tumor suppressor H-rev107. These eukaryotic proteins, along with the bacterial YaeF/poxviral G6R family, show a circular permutation of the catalytic domain. We identified three conserved residues, namely a cysteine, a histidine and a polar residue, that are involved in the catalytic activities of this superfamily. Evolutionary analysis of this superfamily shows that it comprises four major families, with diverse domain architectures in each of them. Several related, but distinct, catalytic activities, such as murein degradation, acyl transfer and amide hydrolysis, have emerged in the N1pC/P60 superfamily. The three conserved catalytic residues of this superfamily are shown to be equivalent to the catalytic triad of the papain-like thiol peptidases. The predicted structural features indicate that the N1pC/P60 enzymes contain a fold similar to the papain-like peptidases, transglutaminases and arylamine acetyltransferases.

Sex-dependent alterations in motor and anxiety-like behavior of aged bacterial peptidoglycan sensing molecule 2 knockout mice.

PubMed

Arentsen, Tim; Khalid, Roksana; Qian, Yu; Diaz Heijtz, Rochellys

2018-01-01

Peptidoglycan recognition proteins (PGRPs) are key sensing-molecules of the innate immune system that specifically detect bacterial peptidoglycan (PGN) and its derivates. PGRPs have recently emerged as potential key regulators of normal brain development and behavior. To test the hypothesis that PGRPs play a role in motor control and anxiety-like behavior in later life, we used 15-month old male and female peptidoglycan recognition protein 2 (Pglyrp2) knockout (KO) mice. Pglyrp2 is an N-acetylmuramyl-l-alanine amidase that hydrolyzes PGN between the sugar backbone and the peptide chain (which is unique among the mammalian PGRPs). Using a battery of behavioral tests, we demonstrate that Pglyrp2 KO male mice display decreased levels of anxiety-like behavior compared with wild type (WT) males. In contrast, Pglyrp2 KO female mice show reduced rearing activity and increased anxiety-like behavior compared to WT females. In the accelerated rotarod test, however, Pglyrp2 KO female mice performed better compared to WT females (i.e., they had longer latency to fall off the rotarod). Further, Pglyrp2 KO male mice exhibited decreased expression levels of synaptophysin, gephyrin, and brain-derived neurotrophic factor in the frontal cortex, but not in the amygdala. Pglyrp2 KO female mice exhibited increased expression levels of spinophilin and alpha-synuclein in the frontal cortex, while exhibiting decreased expression levels of synaptophysin, gephyrin and spinophilin in the amygdala. Our findings suggest a novel role for Pglyrp2asa key regulator of motor and anxiety-like behavior in late life. Copyright © 2017. Published by Elsevier Inc.

Contribution of Three Bile-Associated Loci, bsh, pva, and btlB, to Gastrointestinal Persistence and Bile Tolerance of Listeria monocytogenes

PubMed Central

Begley, Máire; Sleator, Roy D.; Gahan, Cormac G. M.; Hill, Colin

2005-01-01

Listeria monocytogenes must resist the deleterious actions of bile in order to infect and subsequently colonize the human gastrointestinal tract. The molecular mechanisms used by the bacterium to resist bile and the influence of bile on pathogenesis are as yet largely unexplored. This study describes the analysis of three genes—bsh, pva, and btlB—previously annotated as bile-associated loci in the sequenced L. monocytogenes EGDe genome (lmo2067, lmo0446, and lmo0754, respectively). Analysis of deletion mutants revealed a role for all three genes in resisting the acute toxicity of bile and bile salts, particularly glycoconjugated bile salts at low pH. Mutants were unaffected in the other stress responses examined (acid, salt, and detergents). Bile hydrolysis assays demonstrate that L. monocytogenes possesses only one bile salt hydrolase gene, namely, bsh. Transcriptional analyses and activity assays revealed that, although it is regulated by both PrfA and σB, the latter appears to play the greater role in modulating bsh expression. In addition to being incapable of bile hydrolysis, a sigB mutant was shown to be exquisitely sensitive to bile salts. Furthermore, increased expression of sigB was detected under anaerobic conditions and during murine infection. A gene previously annotated as a possible penicillin V amidase (pva) or bile salt hydrolase was shown to be required for resistance to penicillin V but not penicillin G but did not demonstrate a role in bile hydrolysis. Finally, animal (murine) studies revealed an important role for both bsh and btlB in the intestinal persistence of L. monocytogenes. PMID:15664931

Structures of enzyme-intermediate complexes of yeast Nit2: insights into its catalytic mechanism and different substrate specificity compared with mammalian Nit2.

PubMed

Liu, Hejun; Gao, Yongxiang; Zhang, Mengying; Qiu, Xiaoting; Cooper, Arthur J L; Niu, Liwen; Teng, Maikun

2013-08-01

The Nit (nitrilase-like) protein subfamily constitutes branch 10 of the nitrilase superfamily. Nit proteins are widely distributed in nature. Mammals possess two members of the Nit subfamily, namely Nit1 and Nit2. Based on sequence similarity, yeast Nit2 (yNit2) is a homologue of mouse Nit1, a tumour-suppressor protein whose substrate specificity is not yet known. Previous studies have shown that mammalian Nit2 (also a putative tumour suppressor) is identical to ω-amidase, an enzyme that catalyzes the hydrolysis of α-ketoglutaramate (α-KGM) and α-ketosuccinamate (α-KSM) to α-ketoglutarate (α-KG) and oxaloacetate (OA), respectively. In the present study, crystal structures of wild-type (WT) yNit2 and of WT yNit2 in complex with α-KG and with OA were determined. In addition, the crystal structure of the C169S mutant of yNit2 (yNit2-C169S) in complex with an endogenous molecule of unknown structure was also solved. Analysis of the structures revealed that α-KG and OA are covalently bound to Cys169 by the formation of a thioester bond between the sulfhydryl group of the cysteine residue and the γ-carboxyl group of α-KG or the β-carboxyl group of OA, reflecting the presumed reaction intermediates. However, an enzymatic assay suggests that α-KGM is a relatively poor substrate of yNit2. Finally, a ligand was found in the active site of yNit2-C169S that may be a natural substrate of yNit2 or an endogenous regulator of enzyme activity. These crystallographic analyses provide information on the mode of substrate/ligand binding at the active site of yNit2 and insights into the catalytic mechanism. These findings suggest that yNit2 may have broad biological roles in yeast, especially in regard to nitrogen homeostasis, and provide a framework for the elucidation of the substrate specificity and biological role of mammalian Nit1.

Mass spectrometry characterization for N-glycosylation of immunoglobulin Y from hen egg yolk.

PubMed

Sheng, Long; He, Zhenjiao; Liu, Yaping; Ma, Meihu; Cai, Zhaoxia

2018-03-01

Immunoglobulin Y (IgY) is a new therapeutic antibody that exists in hen egg yolk. It is a glycoprotein, not much is known about its N-glycan structures, site occupancy and site-specific N-glycosylation. In this study, purified protein from hen egg yolk was identified as IgY based on SDS-PAGE and MALDI-TOF/TOF MS. N-glycan was released from IgY using peptide-N4-(N-acetyl-beta-glucosaminyl) asparagine-amidase treatment, and the molecular weight of IgY was calculated using the difference between the molecular weight of IgY and deglycosylated IgY. Two potential N-Glycosylation sites (ASN 308 and ASN 409 ) were detected on IgY by nanoLC-ESI MS. Sugar chains were separated using normal phase liquid chromatography after fluorescence labeling, and 17 N-glycan structures were confirmed using ESI-MS. The sugar chain pattern contained high-mannose oligosaccharide, hybrid oligosaccharide and complex oligosaccharide. These results could lead to other important information regarding IgY glycosylation. Copyright © 2017 Elsevier B.V. All rights reserved.

The impact of N-glycosylation on conformation and stability of immunoglobulin Y from egg yolk.

PubMed

Sheng, Long; He, Zhenjiao; Chen, Jiahui; Liu, Yaofa; Ma, Meihu; Cai, Zhaoxia

2017-03-01

Immunoglobulin Y (IgY) is a new therapeutic antibody, and its applications in industry are very broad. To provide insight into the effects of N-glycosylation on IgY, its conformation and stability were studied. In this research, IgY was extracted from egg yolk and then digested by peptide-N4-(N-acetyl-beta-glucosaminyl) asparagine-amidase. SDS-PAGE and infrared absorption spectrum showed that carbohydrates were distinctly reduced after enzymolysis. The circular dichroism spectrum indicated that the IgY molecule became more flexible and disordered after removal of N-glycan. The fluorescence intensity revealed that Trp residues were buried in a more hydrophobic environment after disposal of N-glycan. Storage stability decreased with the removal of oligosaccharide chains based on size-exclusion chromatography analysis. Deglycosylated IgY exhibited less resistance to guanidine hydrochloride-induced unfolding. After deglycosylation, IgY was more sensitive to pepsin. Therefore, N-glycosylation played an important role in the maintenance of the structure and stability of IgY. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparative genomics of four closely related Clostridium perfringens bacteriophages reveals variable evolution among core genes with therapeutic potential

PubMed Central

2011-01-01

Background Because biotechnological uses of bacteriophage gene products as alternatives to conventional antibiotics will require a thorough understanding of their genomic context, we sequenced and analyzed the genomes of four closely related phages isolated from Clostridium perfringens, an important agricultural and human pathogen. Results Phage whole-genome tetra-nucleotide signatures and proteomic tree topologies correlated closely with host phylogeny. Comparisons of our phage genomes to 26 others revealed three shared COGs; of particular interest within this core genome was an endolysin (PF01520, an N-acetylmuramoyl-L-alanine amidase) and a holin (PF04531). Comparative analyses of the evolutionary history and genomic context of these common phage proteins revealed two important results: 1) strongly significant host-specific sequence variation within the endolysin, and 2) a protein domain architecture apparently unique to our phage genomes in which the endolysin is located upstream of its associated holin. Endolysin sequences from our phages were one of two very distinct genotypes distinguished by variability within the putative enzymatically-active domain. The shared or core genome was comprised of genes with multiple sequence types belonging to five pfam families, and genes belonging to 12 pfam families, including the holin genes, which were nearly identical. Conclusions Significant genomic diversity exists even among closely-related bacteriophages. Holins and endolysins represent conserved functions across divergent phage genomes and, as we demonstrate here, endolysins can have significant variability and host-specificity even among closely-related genomes. Endolysins in our phage genomes may be subject to different selective pressures than the rest of the genome. These findings may have important implications for potential biotechnological applications of phage gene products. PMID:21631945

Structural characterization of the N-glycans of gpMuc from Mucuna pruriens seeds.

PubMed

Di Patrizi, Lisa; Rosati, Floriana; Guerranti, Roberto; Pagani, Roberto; Gerwig, Gerrit J; Kamerling, Johannis P

2006-11-01

Mucuna pruriens seeds are used in some countries as a human prophylactic oral anti-snake remedy. Aqueous extracts of M. pruriens seeds possess in vivo activity against cobra and viper venoms, and protect mice against Echis carinatus venom. It was recently demonstrated that the seed immunogen generating the antibody that cross-reacts with the venom proteins is a multiform glycoprotein (gpMuc), and the immunogenic properties of gpMuc seemed to mainly reside in its glycan chains. In the present study, gpMuc was found to contain only N-glycans. Part of the N-glycans could be released with peptide-(N (4)-(N-acetyl-beta -glucosaminyl)asparagine amidase F (PNGase F-sensitive N-glycans); the PNGase F-resistant N-glycans were PNGase A-sensitive. The oligosaccharides released were analyzed by a combination of MALDI-TOF mass spectrometry, HPLC profiling of 2-aminobenzamide-labelled derivatives and (1)H NMR spectroscopy. The PNGase F-sensitive N-glycans comprised a mixture of oligomannose-type structures ranging from Man(5)GlcNAc(2) to Man(9)GlcNAc(2), and two xylosylated structures, Xyl(1)Man(3)GlcNAc(2) and Xyl(1)Man(4)GlcNAc(2). The PNGase A-sensitive N-glycans, containing (alpha 1-3)-linked fucose, were identified as Fuc(1)Xyl(1)Man(2)GlcNAc(2) and Fuc(1)Xyl(1)Man(3)GlcNAc(2). In view of the determined N-glycan ensemble, the immunoreactivity of gpMuc was ascribed to the presence of core (beta 1-2)-linked xylose- and core alpha (1-3)-linked fucose-modified N-glycan chains.

Stage-specific expression of DDX4 and c-kit at different developmental stages of the porcine testis.

PubMed

Lee, Ran; Lee, Won-Young; Park, Hyun-Jung; Ha, Woo-Tae; Woo, Jae-Seok; Chung, Hak-Jae; Lee, Ji-Heon; Hong, Kwonho; Song, Hyuk

2018-03-01

Spermatogenesis begins with spermatogonial stem cells (SSCs), which are located in the basement membrane of the adult testes. Previous studies have described specific biomarkers for undifferentiated porcine spermatogonia or SSCs; however, these markers are not sufficient to understand spermatogenesis at different developmental stages. The objective of this study was characterize the expression of DEAD-Box polypeptide 4 (DDX4, also known as VASA) and tyrosine-protein kinase kit (c-kit), as potential markers of male germ cells in the porcine testis. In porcine testis tissue at prepubertal stages (5, 30, and 60 days), DDX4 and c-kit protein expression was detected in the most undifferentiated spermatogonia, which also express protein gene product 9.5 (PGP9.5). However, in porcine testis tissues from pubertal and postpubertal stages (90, 120, and 150 days), DDX4 and c-kit were not detected in PGP9.5-positive undifferentiated spermatogonia. The DDX4 expression pattern was similar to that of c-kit in the porcine testis. In adult porcine testes, DDX4-expressing cells were located on the lumenal side, compared to synaptonemal complex protein 3-positive primary spermatocytes, but DDX-4 was not co-expressed with acrosin, a known acrosome marker. In addition, DDX4 was detected in PGP9.5-expressing porcine SSCs in culture. Based on our results, we suggest that DDX4 and c-kit are putative markers of undifferentiated spermatogonia in the prepubertal porcine testis. While in the postpubertal porcine testis, they are markers of differentiated spermatocytes. These findings may facilitate future studies of porcine spermatogenesis. Copyright © 2018 Elsevier B.V. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Tianyu; University of Chinese Academy of Sciences, Beijing 100049; Ding, Jinjing

The structure of the Tse3–Tsi3 complex associated with the bacterial type VI secretion system of P. aeruginosa has been solved and refined at 1.9 Å resolution. The structural basis of the recognition of the muramidase effector and its inactivation by its cognate immunity protein is revealed. The type VI secretion system (T6SS) is a bacterial protein-export machine that is capable of delivering virulence effectors between Gram-negative bacteria. The T6SS of Pseudomonas aeruginosa transports two lytic enzymes, Tse1 and Tse3, to degrade cell-wall peptidoglycan in the periplasm of rival bacteria that are competing for niches via amidase and muramidase activities, respectively.more » Two cognate immunity proteins, Tsi1 and Tsi3, are produced by the bacterium to inactivate the two antibacterial effectors, thereby protecting its siblings from self-intoxication. Recently, Tse1–Tsi1 has been structurally characterized. Here, the structure of the Tse3–Tsi3 complex is reported at 1.9 Å resolution. The results reveal that Tse3 contains a C-terminal catalytic domain that adopts a soluble lytic transglycosylase (SLT) fold in which three calcium-binding sites were surprisingly observed close to the catalytic Glu residue. The electrostatic properties of the substrate-binding groove are also distinctive from those of known structures with a similar fold. All of these features imply that a unique catalytic mechanism is utilized by Tse3 in cleaving glycosidic bonds. Tsi3 comprises a single domain showing a β-sandwich architecture that is reminiscent of the immunoglobulin fold. Three loops of Tsi3 insert deeply into the groove of Tse3 and completely occlude its active site, which forms the structural basis of Tse3 inactivation. This work is the first crystallographic report describing the three-dimensional structure of the Tse3–Tsi3 effector–immunity pair.« less

A sensitive LC-MS/MS method for the simultaneous determination of amoxicillin and ambroxol in human plasma with segmental monitoring.

PubMed

Dong, Xin; Ding, Li; Cao, Xiaomei; Jiang, Liyuan; Zhong, Shuisheng

2013-04-01

Amoxicillin (AMO) degrades in plasma at room temperature and readily undergoes hydrolysis by the plasma amidase. In this paper, a novel, rapid and sensitive LC-MS/MS method operated in segmental and multiple reaction monitoring has been developed for the simultaneous determination of amoxicillin and ambroxol in human plasma. The degradation of amoxicillin in plasma was well prevented by immediate addition of 20 μL glacial acetic acid to 200 μL aliquot of freshly collected plasma samples before storage at -80°C. The sensitivity of the method was improved with segmental monitoring of the analytes, and lower limits of quantitation of 0.5 ng/mL for ambroxol and 5 ng/mL for amoxicillin were obtained. The sensitivity of our method was five times better than those of the existing methods. Furthermore, the mass response saturation problem with amoxicillin was avoided by diluting the deproteinized plasma samples with water before injection into the LC-MS/MS system. The method was successfully employed in a pharmacokinetic study of the compound amoxicillin and ambroxol hydrochloride tablets. Copyright © 2012 John Wiley & Sons, Ltd.

Enzymes in Fermented Fish.

PubMed

Giyatmi; Irianto, H E

Fermented fish products are very popular particularly in Southeast Asian countries. These products have unique characteristics, especially in terms of aroma, flavor, and texture developing during fermentation process. Proteolytic enzymes have a main role in hydrolyzing protein into simpler compounds. Fermentation process of fish relies both on naturally occurring enzymes (in the muscle or the intestinal tract) as well as bacteria. Fermented fish products processed using the whole fish show a different characteristic compared to those prepared from headed and gutted fish. Endogenous enzymes like trypsin, chymotrypsin, elastase, and aminopeptidase are the most involved in the fermentation process. Muscle tissue enzymes like cathepsins, peptidases, transaminases, amidases, amino acid decarboxylases, glutamic dehydrogenases, and related enzymes may also play a role in fish fermentation. Due to the decreased bacterial number during fermentation, contribution of microbial enzymes to proteolysis may be expected prior to salting of fish. Commercial enzymes are supplemented during processing for specific purposes, such as quality improvement and process acceleration. In the case of fish sauce, efforts to accelerate fermentation process and to improve product quality have been studied by addition of enzymes such as papain, bromelain, trypsin, pepsin, and chymotrypsin. © 2017 Elsevier Inc. All rights reserved.

Reversed-phase high-performance liquid chromatographic method for the determination of peptidoglycan monomers and structurally related peptides and adamantyltripeptides.

PubMed

Krstanović, Marina; Frkanec, Ruza; Vranesić, Branka; Ljevaković, Durdica; Sporec, Vesna; Tomasić, Jelka

2002-06-25

The reversed-phase HPLC method using UV detection was developed for the determination of (a) immunostimulating peptidoglycan monomers represented by the basic structure GlcNAc-MurNAc-L-Ala-D-isoGln-meso-DAP(omegaNH(2))-D-Ala-D-Ala (PGM) and two more lipophilic derivatives, Boc-Tyr-PGM and (Ada-1-yl)-CH(2)-CO-PGM, (b) two diastereomeric immunostimulating adamantyltripeptides L- and D-(adamant-2-yl)-Gly-L-Ala-D-isoGln and (c) peptides obtained by the enzyme hydrolyses of peptidoglycans and related peptides. The enzymes used, N-acetylmuramyl-L-alanine amidase and an L,D-aminopeptidase are present in mammalian sera and are involved in the metabolism of peptidoglycans and related peptides. Appropriate solvent systems were chosen with regard to structure and lipophilicity of each compound. As well, different gradient systems within the same solvent system had to be applied in order to achieve satisfactory separation and retention time. HPLC separation was developed with the aim to use this method for the study of the stability of the tested compounds, the purity during preparation and isolation and for following the enzyme hydrolyses.

Combination of a recombinant bacterium with organonitrile-degrading and biofilm-forming capability and a positively charged carrier for organonitriles removal.

PubMed

Li, Chunyan; Sun, Yueling; Yue, Zhenlei; Huang, Mingyan; Wang, Jinming; Chen, Xi; An, Xuejiao; Zang, Hailian; Li, Dapeng; Hou, Ning

2018-04-10

The immobilization of organonitrile-degrading bacteria via the addition of biofilm-forming bacteria represents a promising technology for the treatment of organonitrile-containing wastewater, but biofilm-forming bacteria simply mixed with degrading bacteria may reduce the biodegradation efficiency. Nitrile hydratase and amidase genes, which play critical roles in organonitriles degradation, were cloned and transformed into the biofilm-forming bacterium Bacillus subtilis N4 to construct a recombinant bacterium B. subtilis N4/pHTnha-ami. Modified polyethylene carriers with positive charge was applied to promote bacterial adherence and biofilm formation. The immobilized B. subtilis N4/pHTnha-ami was resistant to organonitriles loading shocks and could remove organic cyanide ion with a initial concentration of 392.6 mg/L for 24 h in a moving bed biofilm reactor. The imputed quorum-sensing signal and the high-throughput sequencing analysis of the biofilm indicated that B. subtilis N4/pHTnha-ami was successfully immobilized and became dominant. The successful application of the immobilized recombinant bacterium offers a novel strategy for the biodegradation of recalcitrant compounds. Copyright © 2018 Elsevier B.V. All rights reserved.

Complete genome sequence of 285P, a novel T7-like polyvalent E. coli bacteriophage.

PubMed

Xu, Bin; Ma, Xiangyu; Xiong, Hongyan; Li, Yafei

2014-06-01

Bacteriophages are considered potential biological agents for the control of infectious diseases and environmental disinfection. Here, we describe a novel T7-like polyvalent Escherichia coli bacteriophage, designated "285P," which can lyse several strains of E. coli. The genome, which consists of 39,270 base pairs with a G+C content of 48.73 %, was sequenced and annotated. Forty-three potential open reading frames were identified using bioinformatics tools. Based on whole-genome sequence comparison, phage 285P was identified as a novel strain of subgroup T7. It showed strongest sequence similarity to Kluyvera phage Kvp1. The phylogenetic analyses of both non-structural proteins (endonuclease gp3, amidase gp3.5, DNA primase/helicase gp4, DNA polymerase gp5, and exonuclease gp6) and structural protein (tail fiber protein gp17) led to the identification of 285P as T7-like phage. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analyses verified the annotation of the structural proteins (major capsid protein gp10a, tail protein gp12, and tail fiber protein gp17).

Recombinant Expression of a Genome-encoded N-acetylmuramoyl-L-alanine Amidase that Synergistically Lyses Listeria monocytogenes Biofilms with a Protease

USDA-ARS?s Scientific Manuscript database

Listeria monocytogenes plays a significant role in human food-borne disease caused by eating food contaminated with the bacterium and although incidence is low it is a leading cause of life-threatening, bacterial food-borne disease in humans. L. monocytogenes serotypes 1/2a and 4b can form mixed-cu...

EVALUATION OF ACTIVATED BIOFILTRATION AND ACTIVATED BIOFILTRATION/ACTIVATED SLUDGE TECHNOLOGIES

EPA Science Inventory

The paper presents the results of a review and investigation of the activated biofilter (ABF) and activated biofilter/activated sludge (ABF/AS) technologies and a review of operating records of several municipal plants in the U.S. using these technologies. The overall objective o...

A Personal Retrospective: Elevating Anandamide (AEA) by Targeting Fatty Acid Amide Hydrolase (FAAH) and the Fatty Acid Binding Proteins (FABPs).

PubMed

Deutsch, Dale G

2016-01-01

This perspective was adapted from a Career Achievement Award talk given at the International Cannabinoid Research Society Symposium in Bukovina, Poland on June 27, 2016. As a biochemist working in the neurosciences, I was always fascinated with neurotransmitter inactivation. In 1993 we identified an enzyme activity that breaks down anandamide. We called the enzyme anandamide amidase, now called FAAH. We and other laboratories developed FAAH inhibitors that were useful reagents that also proved to have beneficial physiological effects and until recently, new generations of inhibitors were in clinical trials. Nearly all neurotransmitters are water soluble and as such, require a transmembrane protein transporter to pass through the lipid membrane for inactivation inside the cell. However, using model systems, we and others have shown that this is unnecessary for anandamide, an uncharged hydrophobic molecule that readily diffuses across the cellular membrane. Interestingly, its uptake is driven by the concentration gradient resulting from its breakdown mainly by FAAH localized in the endoplasmic reticulum. We identified the FABPs as intracellular carriers that "solubilize" anandamide, transporting anandamide to FAAH. Compounds that bind to FABPs block AEA breakdown, raising its level. The cannabinoids (THC and CBD) also were discovered to bind FABPs and this may be one of the mechanisms by which CBD works in childhood epilepsy, raising anandamide levels. Targeting FABPs may be advantageous since they have some tissue specificity and do not require reactive serine hydrolase inhibitors, as does FAAH, with potential for off-target reactions. At the International Cannabis Research Society Symposium in 1992, Raphe Mechoulam revealed that his laboratory isolated an endogenous lipid molecule that binds to the CB1 receptor (cannabinoid receptor type 1) and this became the milestone paper published in December of that year describing anandamide (AEA, Devane et al., 1992). As to

A Personal Retrospective: Elevating Anandamide (AEA) by Targeting Fatty Acid Amide Hydrolase (FAAH) and the Fatty Acid Binding Proteins (FABPs)

PubMed Central

Deutsch, Dale G.

2016-01-01

This perspective was adapted from a Career Achievement Award talk given at the International Cannabinoid Research Society Symposium in Bukovina, Poland on June 27, 2016. As a biochemist working in the neurosciences, I was always fascinated with neurotransmitter inactivation. In 1993 we identified an enzyme activity that breaks down anandamide. We called the enzyme anandamide amidase, now called FAAH. We and other laboratories developed FAAH inhibitors that were useful reagents that also proved to have beneficial physiological effects and until recently, new generations of inhibitors were in clinical trials. Nearly all neurotransmitters are water soluble and as such, require a transmembrane protein transporter to pass through the lipid membrane for inactivation inside the cell. However, using model systems, we and others have shown that this is unnecessary for anandamide, an uncharged hydrophobic molecule that readily diffuses across the cellular membrane. Interestingly, its uptake is driven by the concentration gradient resulting from its breakdown mainly by FAAH localized in the endoplasmic reticulum. We identified the FABPs as intracellular carriers that “solubilize” anandamide, transporting anandamide to FAAH. Compounds that bind to FABPs block AEA breakdown, raising its level. The cannabinoids (THC and CBD) also were discovered to bind FABPs and this may be one of the mechanisms by which CBD works in childhood epilepsy, raising anandamide levels. Targeting FABPs may be advantageous since they have some tissue specificity and do not require reactive serine hydrolase inhibitors, as does FAAH, with potential for off-target reactions. At the International Cannabis Research Society Symposium in 1992, Raphe Mechoulam revealed that his laboratory isolated an endogenous lipid molecule that binds to the CB1 receptor (cannabinoid receptor type 1) and this became the milestone paper published in December of that year describing anandamide (AEA, Devane et al., 1992

Active-oxygen scavenging activity of plant extracts.

PubMed

Masaki, H; Sakaki, S; Atsumi, T; Sakurai, H

1995-01-01

To find antioxidative compounds present in plants, 65 types of plant extract were tested using the neotetrazolium method for evidence of superoxide anion-scavenging effects and 7 plant extracts were selected for further investigation. The activity of active-oxygen scavengers such as superoxide anion radicals, hydroxyl radicals, singlet oxygens and lipid peroxides in the 7 plant extracts (Aeseclus hippocastanum L., Hamamelis virginiana L. Polygonum cuspidatum Sieb., Quercus robur L., Rosemarinous officinalis L., Salvia officinalis L. and Sanguisorba officinalis L.) was examined in detail by both ESR spin-trapping and malondialdehyde generation. Furthermore, the active-oxygen scavenging activity of these plant extracts was evaluated using a murine dermal fibroblast culture system. Both Aeseclus hippocastanum L. and Hamamelis virginia L. were found to have strong active-oxygen scavenging activity of and protective activity against cell damage induced by active oxygen. Both Aeseclus hippocastanum L. and Hamamelis virginiana L. are proposed as potent plant extracts with potential application as anti-aging or anti-wrinkle material for the skin.

Recombinant expression of a putative prophage amidase cloned from the genome of Listeria monocytogenes that lyses the bacterium and its biofilm

USDA-ARS?s Scientific Manuscript database

Listeria monocytogenes is a Gram-positive, non-sporeforming, catalase-positive rod that is a major bacterial food-borne disease agent, causing listeriosis. Listeria can be associated with uncooked meats including poultry, uncooked vegetables, soft cheeses and unpasteurized milk. The bacterium can be...

Expression of a Clostridium perfringens genome-encoded putative N-acetylmuramoyl-L-alanine amidase as a potential antimicrobial to control the bacterium

USDA-ARS?s Scientific Manuscript database

Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a substantial role in non-foodborne human, animal and avian diseases as well as human foodborne disease. Previously discovered C. perfringens bacteriophage lytic enzyme amino acid sequences were utilized to iden...

Total enzymatic synthesis of cholecystokinin CCK-5.

PubMed

Xiang, H; Xiang, G Y; Lu, Z M; Guo, L; Eckstein, H

2004-08-01

This paper describes the enzymatic synthesis of the C-terminal fragment H-Gly-Trp-Met-Asp-Phe-NH2 of cholecystokinin. Immobilized enzymes were used for the formation of all peptide bonds except thermolysin. Beginning the synthesis with phenylacetyl (PhAc) glycine carboxamidomethyl ester (OCam) and H-Trp-OMe by using immobilized papain as biocatalyst in buffered ethyl acetate, the dipeptide methyl ester was then coupled directly with Met-OEt.HCl by alpha-chymotrypsin/Celite 545 in a solvent free system. For the 3+2 coupling PhAc-Gly-Trp-Met-OEt had to be converted into its OCam ester. The other fragment H-Asp(OMe)-Phe-NH2 resulted from the coupling of Cbo-Asp(OMe)-OH with H-Phe-NH2.HCl and thermolysin as catalyst, followed by catalytic hydrogenation. Finally PhAc-Gly-Trp-Met-Asp-Phe-NH2 was obtained in a smooth reaction from PhAc-Gly-Trp-Met-OCam and H-Asp(OMe)-Phe-NH2 with alpha-chymotrypsin/Celite 545 in acetonitrile, followed by basic hydrolysis of the beta-methyl ester. The PhAc-group is removed with penicillin G amidase and CCK-5 is obtained in an overall isolated yield of 19.6%.

Use of enzyme inhibitors to evaluate the conversion pathways of ester and amide prodrugs: a case study example with the prodrug ceftobiprole medocaril.

PubMed

Eichenbaum, Gary; Skibbe, Jennifer; Parkinson, Andrew; Johnson, Mark D; Baumgardner, Dawn; Ogilvie, Brian; Usuki, Etsuko; Tonelli, Fred; Holsapple, Jeff; Schmitt-Hoffmann, Anne

2012-03-01

An approach was developed that uses enzyme inhibitors to support the assessment of the pathways that are responsible for the conversion of intravenously administered ester and amide prodrugs in different biological matrices. The methodology was applied to ceftobiprole medocaril (BAL5788), the prodrug of the cephalosporin antibiotic, ceftobiprole. The prodrug was incubated in plasma, postmitochondrial supernatant fractions from human liver (impaired and nonimpaired), kidney, and intestine as well as erythrocytes, in the presence and absence of different enzyme inhibitors (acetylcholinesterase, pseudocholinesterase, retinyl palmitoyl hydrolase, serine esterases, amidases, and cholinesterase). Hydrolysis was rapid, extensive, and not dependent on the presence of β-nicotinamide-adenine dinucleotide phosphate (reduced form) in all matrices tested, suggesting the involvement of carboxylesterases but not P450 enzymes. Hydrolysis in healthy human plasma was rapid and complete and only partially inhibited in the presence of paraoxonase inhibitors or in liver from hepatic impaired patients, suggesting involvement of nonparaoxonase pathways. The results demonstrate the utility of this approach in confirming the presence of multiple conversion pathways of intravenously administered prodrugs and in the case of BAL5788 demonstrated that this prodrug is unlikely to be affected by genetic polymorphisms, drug interactions, or other environmental factors that might inhibit or induce the enzymes involved in its conversion. Copyright © 2011 Wiley Periodicals, Inc.

Automated activity-aware prompting for activity initiation.

PubMed

Holder, Lawrence B; Cook, Diane J

2013-01-01

Performing daily activities without assistance is important to maintaining an independent functional lifestyle. As a result, automated activity prompting systems can potentially extend the period of time that adults can age in place. In this paper we introduce AP, an algorithm to automate activity prompting based on smart home technology. AP learns prompt rules based on the time when activities are typically performed as well as the relationship between activities that normally occur in a sequence. We evaluate the AP algorithm based on smart home datasets and demonstrate its ability to operate within a physical smart environment.

Large-scale physical activity data reveal worldwide activity inequality

PubMed Central

Althoff, Tim; Sosič, Rok; Hicks, Jennifer L.; King, Abby C.; Delp, Scott L.; Leskovec, Jure

2018-01-01

Understanding the basic principles that govern physical activity is needed to curb the global pandemic of physical inactivity1–7 and the 5.3 million deaths per year associated with in-activity2. Our knowledge, however, remains limited owing to the lack of large-scale measurements of physical activity patterns across free-living populations worldwide1, 6. Here, we leverage the wide usage of smartphones with built-in accelerometry to measure physical activity at planetary scale. We study a dataset consisting of 68 million days of physical activity for 717,527 people, giving us a window into activity in 111 countries across the globe. We find inequality in how activity is distributed within countries and that this inequality is a better predictor of obesity prevalence in the population than average activity volume. Reduced activity in females contributes to a large portion of the observed activity inequality. Aspects of the built environment, such as the walkability of a city, were associated with less gender gap in activity and activity inequality. In more walkable cities, activity is greater throughout the day and throughout the week, across age, gender, and body mass index (BMI) groups, with the greatest increases in activity for females. Our findings have implications for global public health policy and urban planning and highlight the role of activity inequality and the built environment for improving physical activity and health. PMID:28693034

Is Enhanced Physical Activity Possible Using Active Videogames?

PubMed

Baranowski, Tom; Baranowski, Janice; O'Connor, Teresia; Lu, Amy Shirong; Thompson, Debbe

2012-06-01

Our research indicated that 10-12-year-old children receiving two active Wii ™ (Nintendo ® ; Nintendo of America, Inc., Redmond, WA) console videogames were no more physically active than children receiving two inactive videogames. Research is needed on how active videogames may increase physical activity.

Is active travel associated with greater physical activity? The contribution of commuting and non-commuting active travel to total physical activity in adults.

PubMed

Sahlqvist, Shannon; Song, Yena; Ogilvie, David

2012-09-01

To complement findings that active travel reduces the risk of morbidity and mortality from chronic diseases, an understanding of the mechanisms through which active travel may lead to improved health is required. The aim of this study is to examine the descriptive epidemiology of all active travel and its associations with recreational and total physical activity in a sample of adults in the UK. In April 2010, data were collected from 3516 adults as part of the baseline survey for the iConnect study in the UK. Travel and recreational physical activity were assessed using detailed seven-day recall instruments. Linear regression analyses, controlling for demographic characteristics, examined associations between active travel, defined as any walking and cycling for transport, and recreational and total physical activity. 65% of respondents (mean age 50.5 years) reported some form of active travel, accumulating an average of 195 min/week (standard deviation=188.6). There were no differences in the recreational physical activity levels of respondents by travel mode category. Adults who used active travel did however report significantly higher total physical activity than those who did not. Substantial physical activity can be accumulated through active travel which also contributes to greater total physical activity. Copyright © 2012 Elsevier Inc. All rights reserved.

Is Enhanced Physical Activity Possible Using Active Videogames?

PubMed Central

Baranowski, Janice; O'Connor, Teresia; Lu, Amy Shirong; Thompson, Debbe

2012-01-01

Abstract Our research indicated that 10–12-year-old children receiving two active Wii™ (Nintendo®; Nintendo of America, Inc., Redmond, WA) console videogames were no more physically active than children receiving two inactive videogames. Research is needed on how active videogames may increase physical activity. PMID:24416640

CD18 activation epitopes induced by leukocyte activation.

PubMed

Beals, C R; Edwards, A C; Gottschalk, R J; Kuijpers, T W; Staunton, D E

2001-12-01

The cell surface adhesion molecule LFA-1 coordinates leukocyte trafficking and is a costimulatory molecule for T cell activation. We developed a panel of mAbs that recognize activation epitopes on the CD18 subunit, and show that stimulation of T lymphocytes appears to be accompanied by a conformational change in a subpopulation of LFA-1 that does not require ligand binding. Activation epitope up-regulation requires divalent cations, is sensitive to cellular signal transduction events, and correlates with cell adhesion. In addition, the stimulated appearance of these activation epitopes is absent in cell lines from patients with leukocyte adhesion deficiency-1/variant that has previously been shown to be defective in LFA-1 activation. Thus, these activation epitope Abs can be used to dissect signal transmission to CD18. Evidence suggests that these CD18 activation epitopes are induced early in cellular activation and are independent of actin rearrangement necessary for avid adhesion. We have also determined that function-blocking CD18 Abs inhibit the induction of activation epitopes. One activation epitope Ab binds to a site on CD18 distinct from that of the blocking Abs, indicating that the blocking Abs suppress a conformational change in LFA-1. We also find that these neoepitopes are present on rLFA-1 with high affinity for ICAM-1 and their binding is modulated in parallel with the affinity of LFA-1 for ICAM-1. Collectively, these neoepitope Abs identify a subpopulation of LFA-1 most likely with high affinity for ICAM-1 and necessary for LFA-1 function.

Active nematic gels as active relaxing solids

NASA Astrophysics Data System (ADS)

Turzi, Stefano S.

2017-11-01

I propose a continuum theory for active nematic gels, defined as fluids or suspensions of orientable rodlike objects endowed with active dynamics, that is based on symmetry arguments and compatibility with thermodynamics. The starting point is our recent theory that models (passive) nematic liquid crystals as relaxing nematic elastomers. The interplay between viscoelastic response and active dynamics of the microscopic constituents is naturally taken into account. By contrast with standard theories, activity is not introduced as an additional term of the stress tensor, but it is added as an external remodeling force that competes with the passive relaxation dynamics and drags the system out of equilibrium. In a simple one-dimensional channel geometry, we show that the interaction between nonuniform nematic order and activity results in either a spontaneous flow of particles or a self-organization into subchannels flowing in opposite directions.

Gamma Band Activity in the Reticular Activating System

PubMed Central

Urbano, Francisco J.; Kezunovic, Nebojsa; Hyde, James; Simon, Christen; Beck, Paige; Garcia-Rill, Edgar

2012-01-01

This review considers recent evidence showing that cells in three regions of the reticular activating system (RAS) exhibit gamma band activity, and describes the mechanisms behind such manifestation. Specifically, we discuss how cells in the mesopontine pedunculopontine nucleus (PPN), intralaminar parafascicular nucleus (Pf), and pontine subcoeruleus nucleus dorsalis (SubCD) all fire in the beta/gamma band range when maximally activated, but no higher. The mechanisms behind this ceiling effect have been recently elucidated. We describe recent findings showing that every cell in the PPN have high-threshold, voltage-dependent P/Q-type calcium channels that are essential, while N-type calcium channels are permissive, to gamma band activity. Every cell in the Pf also showed that P/Q-type and N-type calcium channels are responsible for this activity. On the other hand, every SubCD cell exhibited sodium-dependent subthreshold oscillations. A novel mechanism for sleep–wake control based on well-known transmitter interactions, electrical coupling, and gamma band activity is described. The data presented here on inherent gamma band activity demonstrates the global nature of sleep–wake oscillation that is orchestrated by brainstem–thalamic mechanism, and questions the undue importance given to the hypothalamus for regulation of sleep–wakefulness. The discovery of gamma band activity in the RAS follows recent reports of such activity in other subcortical regions like the hippocampus and cerebellum. We hypothesize that, rather than participating in the temporal binding of sensory events as seen in the cortex, gamma band activity manifested in the RAS may help stabilize coherence related to arousal, providing a stable activation state during waking and paradoxical sleep. Most of our thoughts and actions are driven by pre-conscious processes. We speculate that continuous sensory input will induce gamma band activity in the RAS that could participate in the processes of

Potent antitumor activity of a urokinase-activated engineered anthrax toxin

NASA Astrophysics Data System (ADS)

Liu, Shihui; Aaronson, Hannah; Mitola, David J.; Leppla, Stephen H.; Bugge, Thomas H.

2003-01-01

The acquisition of cell-surface urokinase plasminogen activator activity is a hallmark of malignancy. We generated an engineered anthrax toxin that is activated by cell-surface urokinase in vivo and displays limited toxicity to normal tissue but broad and potent tumoricidal activity. Native anthrax toxin protective antigen, when administered with a chimeric anthrax toxin lethal factor, Pseudomonas exotoxin fusion protein, was extremely toxic to mice, causing rapid and fatal organ damage. Replacing the furin activation sequence in anthrax toxin protective antigen with an artificial peptide sequence efficiently activated by urokinase greatly attenuated toxicity to mice. In addition, the mutation conferred cell-surface urokinase-dependent toxin activation in vivo, as determined by using a panel of plasminogen, plasminogen activator, plasminogen activator receptor, and plasminogen activator inhibitor-deficient mice. Surprisingly, toxin activation critically depended on both urokinase plasminogen activator receptor and plasminogen in vivo, showing that both proteins are essential cofactors for the generation of cell-surface urokinase. The engineered toxin displayed potent tumor cell cytotoxicity to a spectrum of transplanted tumors of diverse origin and could eradicate established solid tumors. This tumoricidal activity depended strictly on tumor cell-surface plasminogen activation. The data show that a simple change of protease activation specificity converts anthrax toxin from a highly lethal to a potent tumoricidal agent.

Evaluating Active U: an internet-mediated physical activity program

PubMed Central

Buis, Lorraine R; Poulton, Timothy A; Holleman, Robert G; Sen, Ananda; Resnick, Paul J; Goodrich, David E; Palma-Davis, LaVaughn; Richardson, Caroline R

2009-01-01

Background Engaging in regular physical activity can be challenging, particularly during the winter months. To promote physical activity at the University of Michigan during the winter months, an eight-week Internet-mediated program (Active U) was developed providing participants with an online physical activity log, goal setting, motivational emails, and optional team participation and competition. Methods This study is a program evaluation of Active U. Approximately 47,000 faculty, staff, and graduate students were invited to participate in the online Active U intervention in the winter of 2007. Participants were assigned a physical activity goal and were asked to record each physical activity episode into the activity log for eight weeks. Statistics for program reach, effectiveness, adoption, and implementation were calculated using the Re-Aim framework. Multilevel regression analyses were used to assess the decline in rates of data entry and goal attainment during the program, to assess the likelihood of joining a team by demographic characteristics, to test the association between various predictors and the number of weeks an individual met his or her goal, and to analyze server load. Results Overall, 7,483 individuals registered with the Active U website (≈16% of eligible), and 79% participated in the program by logging valid data at least once. Staff members, older participants, and those with a BMI < 25 were more likely to meet their weekly physical activity goals, and average rate of meeting goals was higher among participants who joined a competitive team compared to those who participated individually (IRR = 1.28, P < .001). Conclusion Internet-mediated physical activity interventions that focus on physical activity logging and goal setting while incorporating team competition may help a significant percentage of the target population maintain their physical activity during the winter months. PMID:19744311

Insights into substrate specificity of NlpC/P60 cell wall hydrolases containing bacterial SH3 domains

DOE PAGES

Xu, Qingping; Mengin-Lecreulx, Dominique; Liu, Xueqian W.; ...

2015-09-15

Bacterial SH3 (SH3b) domains are commonly fused with papain-like Nlp/P60 cell wall hydrolase domains. To understand how the modular architecture of SH3b and NlpC/P60 affects the activity of the catalytic domain, three putative NlpC/P60 cell wall hydrolases were biochemically and structurally characterized. In addition, these enzymes all have γ-d-Glu-A 2pm (A 2pm is diaminopimelic acid) cysteine amidase (ordl-endopeptidase) activities but with different substrate specificities. One enzyme is a cell wall lysin that cleaves peptidoglycan (PG), while the other two are cell wall recycling enzymes that only cleave stem peptides with an N-terminall-Ala. Their crystal structures revealed a highly conserved structuremore » consisting of two SH3b domains and a C-terminal NlpC/P60 catalytic domain, despite very low sequence identity. Interestingly, loops from the first SH3b domain dock into the ends of the active site groove of the catalytic domain, remodel the substrate binding site, and modulate substrate specificity. Two amino acid differences at the domain interface alter the substrate binding specificity in favor of stem peptides in recycling enzymes, whereas the SH3b domain may extend the peptidoglycan binding surface in the cell wall lysins. Remarkably, the cell wall lysin can be converted into a recycling enzyme with a single mutation.Peptidoglycan is a meshlike polymer that envelops the bacterial plasma membrane and bestows structural integrity. Cell wall lysins and recycling enzymes are part of a set of lytic enzymes that target covalent bonds connecting the amino acid and amino sugar building blocks of the PG network. These hydrolases are involved in processes such as cell growth and division, autolysis, invasion, and PG turnover and recycling. To avoid cleavage of unintended substrates, these enzymes have very selective substrate specificities. Our biochemical and structural analysis of three modular NlpC/P60 hydrolases, one lysin, and two recycling enzymes

Insights into substrate specificity of NlpC/P60 cell wall hydrolases containing bacterial SH3 domains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Qingping; Mengin-Lecreulx, Dominique; Liu, Xueqian W.

Bacterial SH3 (SH3b) domains are commonly fused with papain-like Nlp/P60 cell wall hydrolase domains. To understand how the modular architecture of SH3b and NlpC/P60 affects the activity of the catalytic domain, three putative NlpC/P60 cell wall hydrolases were biochemically and structurally characterized. In addition, these enzymes all have γ-d-Glu-A 2pm (A 2pm is diaminopimelic acid) cysteine amidase (ordl-endopeptidase) activities but with different substrate specificities. One enzyme is a cell wall lysin that cleaves peptidoglycan (PG), while the other two are cell wall recycling enzymes that only cleave stem peptides with an N-terminall-Ala. Their crystal structures revealed a highly conserved structuremore » consisting of two SH3b domains and a C-terminal NlpC/P60 catalytic domain, despite very low sequence identity. Interestingly, loops from the first SH3b domain dock into the ends of the active site groove of the catalytic domain, remodel the substrate binding site, and modulate substrate specificity. Two amino acid differences at the domain interface alter the substrate binding specificity in favor of stem peptides in recycling enzymes, whereas the SH3b domain may extend the peptidoglycan binding surface in the cell wall lysins. Remarkably, the cell wall lysin can be converted into a recycling enzyme with a single mutation.Peptidoglycan is a meshlike polymer that envelops the bacterial plasma membrane and bestows structural integrity. Cell wall lysins and recycling enzymes are part of a set of lytic enzymes that target covalent bonds connecting the amino acid and amino sugar building blocks of the PG network. These hydrolases are involved in processes such as cell growth and division, autolysis, invasion, and PG turnover and recycling. To avoid cleavage of unintended substrates, these enzymes have very selective substrate specificities. Our biochemical and structural analysis of three modular NlpC/P60 hydrolases, one lysin, and two recycling enzymes

Insights into Substrate Specificity of NlpC/P60 Cell Wall Hydrolases Containing Bacterial SH3 Domains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Qingping; Mengin-Lecreulx, Dominique; Liu, Xueqian W.

ABSTRACT Bacterial SH3 (SH3b) domains are commonly fused with papain-like Nlp/P60 cell wall hydrolase domains. To understand how the modular architecture of SH3b and NlpC/P60 affects the activity of the catalytic domain, three putative NlpC/P60 cell wall hydrolases were biochemically and structurally characterized. These enzymes all have γ-d-Glu-A 2pm (A 2pm is diaminopimelic acid) cysteine amidase (ordl-endopeptidase) activities but with different substrate specificities. One enzyme is a cell wall lysin that cleaves peptidoglycan (PG), while the other two are cell wall recycling enzymes that only cleave stem peptides with an N-terminall-Ala. Their crystal structures revealed a highly conserved structure consistingmore » of two SH3b domains and a C-terminal NlpC/P60 catalytic domain, despite very low sequence identity. Interestingly, loops from the first SH3b domain dock into the ends of the active site groove of the catalytic domain, remodel the substrate binding site, and modulate substrate specificity. Two amino acid differences at the domain interface alter the substrate binding specificity in favor of stem peptides in recycling enzymes, whereas the SH3b domain may extend the peptidoglycan binding surface in the cell wall lysins. Remarkably, the cell wall lysin can be converted into a recycling enzyme with a single mutation. IMPORTANCEPeptidoglycan is a meshlike polymer that envelops the bacterial plasma membrane and bestows structural integrity. Cell wall lysins and recycling enzymes are part of a set of lytic enzymes that target covalent bonds connecting the amino acid and amino sugar building blocks of the PG network. These hydrolases are involved in processes such as cell growth and division, autolysis, invasion, and PG turnover and recycling. To avoid cleavage of unintended substrates, these enzymes have very selective substrate specificities. Our biochemical and structural analysis of three modular NlpC/P60 hydrolases, one lysin, and two recycling

Youth Physical Activity Resources Use and Activity Measured by Accelerometry

PubMed Central

Maslow, Andréa L.; Colabianchi, Natalie

2014-01-01

Objectives To examine whether utilization of physical activity resources (eg, parks) was associated with daily physical activity measured by accelerometry. Methods 111 adolescents completed a travel diary with concurrent accelerometry. The main exposure was self-reported utilization of a physical activity resource (none/1+ resources). The main outcomes were total minutes spent in daily 1) moderate-vigorous physical activity and 2) vigorous physical activity. Results Utilizing a physical activity resource was significantly associated with total minutes in moderate-vigorous physical activity. African-Americans and males had significantly greater moderate-vigorous physical activity. Conclusions Results from this study support the development and use of physical activity resources. PMID:21204684

Youth physical activity resource use and activity measured by accelerometry.

PubMed

Maslow, Andréa L; Colabianchi, Natalie

2011-01-01

To examine whether use of physical activity resources (eg, parks) was associated with daily physical activity measured by accelerometry. One hundred eleven adolescents completed a travel diary with concurrent accelerometry. The main exposure was self-reported use of a physical activity resource (none /1+ resources). The main outcomes were total minutes spent in daily (1) moderate-vigorous physical activity and (2) vigorous physical activity. Using a physical activity resource was significantly associated with total minutes in moderate-vigorous physical activity. African Americans and males had significantly greater moderate-vigorous physical activity. Results from this study support the development and use of physical activity resources.

Prothrombin activation on the activated platelet surface optimizes expression of procoagulant activity

PubMed Central

Wood, Jeremy P.; Silveira, Jay R.; Maille, Nicole M.; Haynes, Laura M.

2011-01-01

Effective hemostasis relies on the timely formation of α-thrombin via prothrombinase, a Ca2+-dependent complex of factors Va and Xa assembled on the activated platelet surface, which cleaves prothrombin at Arg271 and Arg320. Whereas initial cleavage at Arg271 generates the inactive intermediate prethrombin-2, initial cleavage at Arg320 generates the enzymatically active intermediate meizothrombin. To determine which of these intermediates is formed when prothrombin is processed on the activated platelet surface, the cleavage of prothrombin, and prothrombin mutants lacking either one of the cleavage sites, was monitored on the surface of either thrombin- or collagen-activated platelets. Regardless of the agonist used, prothrombin was initially cleaved at Arg271 generating prethrombin-2, with α-thrombin formation quickly after via cleavage at Arg320. The pathway used was independent of the source of factor Va (plasma- or platelet-derived) and was unaffected by soluble components of the platelet releasate. When both cleavage sites are presented within the same substrate molecule, Arg271 effectively competes against Arg320 (with an apparent IC50 = 0.3μM), such that more than 90% to 95% of the initial cleavage occurs at Arg271. We hypothesize that use of the prethrombin-2 pathway serves to optimize the procoagulant activity expressed by activated platelets, by limiting the anticoagulant functions of the alternate intermediate, meizothrombin. PMID:21131592

Prothrombin activation on the activated platelet surface optimizes expression of procoagulant activity.

PubMed

Wood, Jeremy P; Silveira, Jay R; Maille, Nicole M; Haynes, Laura M; Tracy, Paula B

2011-02-03

Effective hemostasis relies on the timely formation of α-thrombin via prothrombinase, a Ca(2+)-dependent complex of factors Va and Xa assembled on the activated platelet surface, which cleaves prothrombin at Arg271 and Arg320. Whereas initial cleavage at Arg271 generates the inactive intermediate prethrombin-2, initial cleavage at Arg320 generates the enzymatically active intermediate meizothrombin. To determine which of these intermediates is formed when prothrombin is processed on the activated platelet surface, the cleavage of prothrombin, and prothrombin mutants lacking either one of the cleavage sites, was monitored on the surface of either thrombin- or collagen-activated platelets. Regardless of the agonist used, prothrombin was initially cleaved at Arg271 generating prethrombin-2, with α-thrombin formation quickly after via cleavage at Arg320. The pathway used was independent of the source of factor Va (plasma- or platelet-derived) and was unaffected by soluble components of the platelet releasate. When both cleavage sites are presented within the same substrate molecule, Arg271 effectively competes against Arg320 (with an apparent IC(50) = 0.3μM), such that more than 90% to 95% of the initial cleavage occurs at Arg271. We hypothesize that use of the prethrombin-2 pathway serves to optimize the procoagulant activity expressed by activated platelets, by limiting the anticoagulant functions of the alternate intermediate, meizothrombin.

Stated Uptake of Physical Activity Rewards Programmes Among Active and Insufficiently Active Full-Time Employees.

PubMed

Ozdemir, Semra; Bilger, Marcel; Finkelstein, Eric A

2017-10-01

Employers are increasingly relying on rewards programmes in an effort to promote greater levels of activity among employees; however, if enrolment in these programmes is dominated by active employees, then they are unlikely to be a good use of resources. This study uses a stated-preference survey to better understand who participates in rewards-based physical activity programmes, and to quantify stated uptake by active and insufficiently active employees. The survey was fielded to a national sample of 950 full-time employees in Singapore between 2012 and 2013. Participants were asked to choose between hypothetical rewards programmes that varied along key dimensions and whether or not they would join their preferred programme if given the opportunity. A mixed logit model was used to analyse the data and estimate predicted uptake for specific programmes. We then simulated employer payments based on predictions for the percentage of each type of employee likely to meet the activity goal. Stated uptake ranged from 31 to 67% of employees, depending on programme features. For each programme, approximately two-thirds of those likely to enrol were insufficiently active. Results showed that insufficiently active employees, who represent the majority, are attracted to rewards-based physical activity programmes, and at approximately the same rate as active employees, even when enrolment fees are required. This suggests that a programme with generous rewards and a modest enrolment fee may have strong employee support and be within the range of what employers may be willing to spend.

Activated coconut shell charcoal carbon using chemical-physical activation

NASA Astrophysics Data System (ADS)

Budi, Esmar; Umiatin, Nasbey, Hadi; Bintoro, Ridho Akbar; Wulandari, Futri; Erlina

2016-02-01

The use of activated carbon from natural material such as coconut shell charcoal as metal absorbance of the wastewater is a new trend. The activation of coconut shell charcoal carbon by using chemical-physical activation has been investigated. Coconut shell was pyrolized in kiln at temperature about 75 - 150 °C for about 6 hours in producing charcoal. The charcoal as the sample was shieved into milimeter sized granule particle and chemically activated by immersing in various concentration of HCl, H3PO4, KOH and NaOH solutions. The samples then was physically activated using horizontal furnace at 400°C for 1 hours in argon gas environment with flow rate of 200 kg/m3. The surface morphology and carbon content of activated carbon were characterized by using SEM/EDS. The result shows that the pores of activated carbon are openned wider as the chemical activator concentration is increased due to an excessive chemical attack. However, the pores tend to be closed as further increasing in chemical activator concentration due to carbon collapsing.

Activity flow over resting-state networks shapes cognitive task activations.

PubMed

Cole, Michael W; Ito, Takuya; Bassett, Danielle S; Schultz, Douglas H

2016-12-01

Resting-state functional connectivity (FC) has helped reveal the intrinsic network organization of the human brain, yet its relevance to cognitive task activations has been unclear. Uncertainty remains despite evidence that resting-state FC patterns are highly similar to cognitive task activation patterns. Identifying the distributed processes that shape localized cognitive task activations may help reveal why resting-state FC is so strongly related to cognitive task activations. We found that estimating task-evoked activity flow (the spread of activation amplitudes) over resting-state FC networks allowed prediction of cognitive task activations in a large-scale neural network model. Applying this insight to empirical functional MRI data, we found that cognitive task activations can be predicted in held-out brain regions (and held-out individuals) via estimated activity flow over resting-state FC networks. This suggests that task-evoked activity flow over intrinsic networks is a large-scale mechanism explaining the relevance of resting-state FC to cognitive task activations.

Activity flow over resting-state networks shapes cognitive task activations

PubMed Central

Cole, Michael W.; Ito, Takuya; Bassett, Danielle S.; Schultz, Douglas H.

2016-01-01

Resting-state functional connectivity (FC) has helped reveal the intrinsic network organization of the human brain, yet its relevance to cognitive task activations has been unclear. Uncertainty remains despite evidence that resting-state FC patterns are highly similar to cognitive task activation patterns. Identifying the distributed processes that shape localized cognitive task activations may help reveal why resting-state FC is so strongly related to cognitive task activations. We found that estimating task-evoked activity flow (the spread of activation amplitudes) over resting-state FC networks allows prediction of cognitive task activations in a large-scale neural network model. Applying this insight to empirical functional MRI data, we found that cognitive task activations can be predicted in held-out brain regions (and held-out individuals) via estimated activity flow over resting-state FC networks. This suggests that task-evoked activity flow over intrinsic networks is a large-scale mechanism explaining the relevance of resting-state FC to cognitive task activations. PMID:27723746

Modeling Patterns of Activities using Activity Curves

PubMed Central

Dawadi, Prafulla N.; Cook, Diane J.; Schmitter-Edgecombe, Maureen

2016-01-01

Pervasive computing offers an unprecedented opportunity to unobtrusively monitor behavior and use the large amount of collected data to perform analysis of activity-based behavioral patterns. In this paper, we introduce the notion of an activity curve, which represents an abstraction of an individual’s normal daily routine based on automatically-recognized activities. We propose methods to detect changes in behavioral routines by comparing activity curves and use these changes to analyze the possibility of changes in cognitive or physical health. We demonstrate our model and evaluate our change detection approach using a longitudinal smart home sensor dataset collected from 18 smart homes with older adult residents. Finally, we demonstrate how big data-based pervasive analytics such as activity curve-based change detection can be used to perform functional health assessment. Our evaluation indicates that correlations do exist between behavior and health changes and that these changes can be automatically detected using smart homes, machine learning, and big data-based pervasive analytics. PMID:27346990

Modeling Patterns of Activities using Activity Curves.

PubMed

Dawadi, Prafulla N; Cook, Diane J; Schmitter-Edgecombe, Maureen

2016-06-01

Pervasive computing offers an unprecedented opportunity to unobtrusively monitor behavior and use the large amount of collected data to perform analysis of activity-based behavioral patterns. In this paper, we introduce the notion of an activity curve , which represents an abstraction of an individual's normal daily routine based on automatically-recognized activities. We propose methods to detect changes in behavioral routines by comparing activity curves and use these changes to analyze the possibility of changes in cognitive or physical health. We demonstrate our model and evaluate our change detection approach using a longitudinal smart home sensor dataset collected from 18 smart homes with older adult residents. Finally, we demonstrate how big data-based pervasive analytics such as activity curve-based change detection can be used to perform functional health assessment. Our evaluation indicates that correlations do exist between behavior and health changes and that these changes can be automatically detected using smart homes, machine learning, and big data-based pervasive analytics.

Parents’ Activity-Related Parenting Practices Predict Girls’ Physical Activity

PubMed Central

DAVISON, KIRSTEN KRAHNSTOEVER; CUTTING, TANJA M.; BIRCH, LEANN L.

2008-01-01

Purpose Using a sample of 180 9-yr-old girls and their parents, this study examined (a) parents’ activity-related parenting strategies and similarities and differences in such strategies for mothers and fathers, and (b) links between activity-related parenting strategies and girls’ physical activity patterns. Methods Measures of girls’ physical activity included the Children’s Physical Activity scale, participation in organized sports, and physical fitness. We developed a questionnaire to assess ways in which parents promote physical activity among their children. Results Exploratory and confirmatory factor analyses identified two factors for each parent including logistic support of girls’ activity (i.e., enrolling girls in sports and driving them to events) and parents’ explicit modeling (i.e., the extent to which parents used their own behavior to encourage their daughters to be active). Mothers reported significantly higher levels of logistic support than fathers, whereas fathers reported higher levels of explicit modeling than mothers. Although mothers and fathers tended to report different methods of support, both methods were associated with higher physical activity among girls. Finally, girls reported significantly higher levels of physical activity when at least one parent reported high levels of overall support in comparison to no parents; no significant differences were identified for support from one versus two parents. Conclusion Results from this study indicate the positive contribution that parents can have on activity practices of their young daughters. PMID:12972881

Incorporation of Socio-scientific Content into Active Learning Activities

NASA Astrophysics Data System (ADS)

King, D. B.; Lewis, J. E.; Anderson, K.; Latch, D.; Sutheimer, S.; Webster, G.; Moog, R.

2014-12-01

Active learning has gained increasing support as an effective pedagogical technique to improve student learning. One way to promote active learning in the classroom is the use of in-class activities in place of lecturing. As part of an NSF-funded project, a set of in-class activities have been created that use climate change topics to teach chemistry content. These activities use the Process Oriented Guided Inquiry Learning (POGIL) methodology. In this pedagogical approach a set of models and a series of critical thinking questions are used to guide students through the introduction to or application of course content. Students complete the activities in their groups, with the faculty member as a facilitator of learning. Through assigned group roles and intentionally designed activity structure, process skills, such as teamwork, communication, and information processing, are developed during completion of the activity. Each of these climate change activities contains a socio-scientific component, e.g., social, ethical and economic data. In one activity, greenhouse gases are used to explain the concept of dipole moment. Data about natural and anthropogenic production rates, global warming potential and atmospheric lifetimes for a list of greenhouse gases are presented. The students are asked to identify which greenhouse gas they would regulate, with a corresponding explanation for their choice. They are also asked to identify the disadvantages of regulating the gas they chose in the previous question. In another activity, where carbon sequestration is used to demonstrate the utility of a phase diagram, students use economic and environmental data to choose the best location for sequestration. Too often discussions about climate change (both in and outside the classroom) consist of purely emotional responses. These activities force students to use data to support their arguments and hypothesize about what other data could be used in the corresponding discussion to

Activity computer program for calculating ion irradiation activation

NASA Astrophysics Data System (ADS)

Palmer, Ben; Connolly, Brian; Read, Mark

2017-07-01

A computer program, Activity, was developed to predict the activity and gamma lines of materials irradiated with an ion beam. It uses the TENDL (Koning and Rochman, 2012) [1] proton reaction cross section database, the Stopping and Range of Ions in Matter (SRIM) (Biersack et al., 2010) code, a Nuclear Data Services (NDS) radioactive decay database (Sonzogni, 2006) [2] and an ENDF gamma decay database (Herman and Chadwick, 2006) [3]. An extended version of Bateman's equation is used to calculate the activity at time t, and this equation is solved analytically, with the option to also solve by numeric inverse Laplace Transform as a failsafe. The program outputs the expected activity and gamma lines of the activated material.

Sirtuin activators.

PubMed

Alcaín, Francisco J; Villalba, José M

2009-04-01

Sirtuin 1-7 (SIRT1-7) are deacetylases that are dependent on NAD(+) for their activity. SIRT1 down-regulates p53 activity, increasing lifespan, cell survival, and neuroprotection; it also deacetylates peroxisome proliferator-activated receptor-gamma and its coactivator 1alpha, promoting fat mobilization, increasing mitochondrial size and number, and positively regulating insulin secretion. Sirtuins link nutrient availability and energy metabolism. Calorie restriction, which increases lifespan and is beneficial in age-related disorders, activates sirtuin. Major efforts are thus focused to developing sirtuin activators. After discussing the potential involvement of sirtuins in pathophysiological processes, this review looks at new, synthetic sirtuin activators. To date, resveratrol is the most potent natural compound able to activate SIRT1, mimicking the positive effect of calorie restriction. Resveratrol might help in the treatment or prevention of obesity and in preventing the aging-related decline in heart function and neuronal loss. As resveratrol has low bioavailability and interacts with multiple molecular targets, the development of new molecules with better bioavailability and targeting sirtuin at lower concentrations is a promising field of the medicinal chemistry. New SIRT1 activators that are up to 1000 times more effective than resveratrol have recently been identified. These improve the response to insulin and increase the number and activity of mitochondria in obese mice. Human trials with a formulation of resveratrol with improved bioavailability and with a synthetic SIRT1 activator are in progress.

Older people and 'active ageing': Subjective aspects of ageing actively.

PubMed

Stenner, Paul; McFarquhar, Tara; Bowling, Ann

2011-04-01

Following a critical overview of the active ageing concept, a thematic decomposition of 42 transcribed interviews with British people aged 72 years and over indicates that active ageing is understood in relation to physical, cognitive, psychological and social factors, but that these co-exist in complex combinations. The notion of activity in active ageing is grasped in relation to an active/passive distinction which emphasizes the enhancement or diminishment of concrete powers of activity. A 'challenge and response' framework is suggested for future research on active ageing.

Active-R filter

DOEpatents

Soderstrand, Michael A.

1976-01-01

An operational amplifier-type active filter in which the only capacitor in the circuit is the compensating capacitance of the operational amplifiers, the various feedback and coupling elements being essentially solely resistive.

Is enhanced physical activity possible using active videogames?

USDA-ARS?s Scientific Manuscript database

Our research indicated that 10– to 12-year-old children receiving two active Wii (TM)(Nintendo (R); Nintendo of America, Inc., Redmond, WA) console videogames were no more physically active than children receiving two inactive videogames. Research is needed on how active videogames may increase phys...

101 Environmental Education Activities. Booklet 4--Science Activities.

ERIC Educational Resources Information Center

Whitney, Helen, Comp.

Fourth in the series "101 Environmental Education Activities" by the Upper Mississippi River ECO-Center, the booklet contains 39 environment-based science activities directed to students in primary, intermediate, and junior high classes. Organization of the activities usually includes grade level, objectives, procedures, and materials,…

Lipid-induced NOX2 activation inhibits autophagic flux by impairing lysosomal enzyme activity[S

PubMed Central

Jaishy, Bharat; Zhang, Quanjiang; Chung, Heaseung S.; Riehle, Christian; Soto, Jamie; Jenkins, Stephen; Abel, Patrick; Cowart, L. Ashley; Van Eyk, Jennifer E.; Abel, E. Dale

2015-01-01

Autophagy is a catabolic process involved in maintaining energy and organelle homeostasis. The relationship between obesity and the regulation of autophagy is cell type specific. Despite adverse consequences of obesity on cardiac structure and function, the contribution of altered cardiac autophagy in response to fatty acid overload is incompletely understood. Here, we report the suppression of autophagosome clearance and the activation of NADPH oxidase (Nox)2 in both high fat-fed murine hearts and palmitate-treated H9C2 cardiomyocytes (CMs). Defective autophagosome clearance is secondary to superoxide-dependent impairment of lysosomal acidification and enzyme activity in palmitate-treated CMs. Inhibition of Nox2 prevented superoxide overproduction, restored lysosome acidification and enzyme activity, and reduced autophagosome accumulation in palmitate-treated CMs. Palmitate-induced Nox2 activation was dependent on the activation of classical protein kinase Cs (PKCs), specifically PKCβII. These findings reveal a novel mechanism linking lipotoxicity with a PKCβ-Nox2-mediated impairment in pH-dependent lysosomal enzyme activity that diminishes autophagic turnover in CMs. PMID:25529920

Plant cysteine proteases that evoke itch activate protease-activated receptors

PubMed Central

Reddy, V.B.; Lerner, E.A.

2013-01-01

Background Bromelain, ficin and papain are cysteine proteases from plants that produce itch upon injection into skin. Their mechanism of action has not been considered previously. Objectives To determine the mechanism by which these proteases function. Methods The ability of these proteases to activate protease-activated receptors was determined by ratiometric calcium imaging. Results We show here that bromelain, ficin and papain activate protease-activated receptors 2 and 4. Conclusions Bromelain, ficin and papain function as signalling molecules and activate protease-activated receptors. Activation of these receptors is the likely mechanism by which these proteases evoke itch. PMID:20491769

Global optogenetic activation of inhibitory interneurons during epileptiform activity.

PubMed

Ledri, Marco; Madsen, Marita Grønning; Nikitidou, Litsa; Kirik, Deniz; Kokaia, Merab

2014-02-26

Optogenetic techniques provide powerful tools for bidirectional control of neuronal activity and investigating alterations occurring in excitability disorders, such as epilepsy. In particular, the possibility to specifically activate by light-determined interneuron populations expressing channelrhodopsin-2 provides an unprecedented opportunity of exploring their contribution to physiological and pathological network activity. There are several subclasses of interneurons in cortical areas with different functional connectivity to the principal neurons (e.g., targeting their perisomatic or dendritic compartments). Therefore, one could optogenetically activate specific or a mixed population of interneurons and dissect their selective or concerted inhibitory action on principal cells. We chose to explore a conceptually novel strategy involving simultaneous activation of mixed populations of interneurons by optogenetics and study their impact on ongoing epileptiform activity in mouse acute hippocampal slices. Here we demonstrate that such approach results in a brief initial action potential discharge in CA3 pyramidal neurons, followed by prolonged suppression of ongoing epileptiform activity during light exposure. Such sequence of events was caused by massive light-induced release of GABA from ChR2-expressing interneurons. The inhibition of epileptiform activity was less pronounced if only parvalbumin- or somatostatin-expressing interneurons were activated by light. Our data suggest that global optogenetic activation of mixed interneuron populations is a more effective approach for development of novel therapeutic strategies for epilepsy, but the initial action potential generation in principal neurons needs to be taken in consideration.

Activation of peroxisome proliferator-activated receptors (PPARs) by their ligands and protein kinase A activators

PubMed Central

Lazennec, Gwendal; Canaple, Laurence; Saugy, Damien; Wahli, Walter

2000-01-01

The nuclear peroxisome proliferator-activated receptors (PPARs) α, β and γ activate the transcription of multiple genes involved in lipid metabolism. Several natural and synthetic ligands have been identified for each PPAR isotype but little is known about the phosphorylation state of these receptors. We show here that activators of protein kinase A (PKA) can enhance mouse PPAR activity in the absence and the presence of exogenous ligands in transient transfection experiments. The activation function 1 (AF-1) of PPARs was dispensable for transcriptional enhancement, whereas the activation function 2 (AF-2) was required for this effect. We also show that several domains of PPAR can be phosphorylated by PKA in vitro. Moreover, gel experiments suggest that PKA stabilizes binding of the liganded PPAR to DNA. PKA inhibitors decreased not only the kinase dependent induction of PPARs but also their ligand-dependent induction, suggesting that the ligands may also mobilize the PKA pathway to lead to maximal transcriptional induction by PPARs. Moreover, comparing PPARα KO with PPARα wild-type mice, we show that the expression of the ACO gene can be regulated by PKA-activated PPARα in liver. These data demonstrate that the PKA pathway is an important modulator of PPAR activity and we propose a model associating this pathway in the control of fatty acid β-oxidation under conditions of fasting, stress and exercise. PMID:11117527

Parental Activity as a Determinant of Activity Level and Patterns of Activity in Obese Children.

ERIC Educational Resources Information Center

Kalakanis, Lisa E.; Goldfield, Gary S.; Paluch, Rocco A.; Epstein, Leonard H.

2001-01-01

Investigated the level and pattern of moderate-to-vigorous physical activity (MVPA) in obese children, examining predictors of their activity. Children and their parents wore accelerometers for several days and provided demographic data. Parental activity levels significantly and independently predicted and improved the prediction of children's…

Impact of Activity Behaviors on Physical Activity Identity and Self-Efficacy.

ERIC Educational Resources Information Center

Miller, Kim H.; Ogletree, Robert J.; Welshimer, Kathleen

2002-01-01

Examined the relationship of physical activity level and length of time of adherence to physical activity with physical activity identity (PAI) and physical activity self-efficacy (PASE). Surveys of 409 adult university employees indicated that vigorous activity related to higher PAI and PASE scores, and activity level contributed significantly…

In vivo imaging of protease activity by Probody therapeutic activation

PubMed Central

Wong, Kenneth R.; Menendez, Elizabeth; Craik, Charles S.; Kavanaugh, W. Michael; Vasiljeva, Olga

2017-01-01

Probody™ therapeutics are recombinant, proteolytically-activated antibody prodrugs, engineered to remain inert until activated locally by tumor-associated proteases. Probody therapeutics exploit the fundamental dysregulation of extracellular protease activity that exists in tumors relative to healthy tissue. Leveraging the ability of a Probody therapeutic to bind its target at the site of disease after proteolytic cleavage, we developed a novel method for profiling protease activity in living animals. Using NIR optical imaging, we demonstrated that a non-labeled anti-EGFR Probody therapeutic can become activated and compete for binding to tumor cells in vivo with a labeled anti-EGFR monoclonal antibody. Furthermore, by inhibiting matriptase activity in vivo with a blocking-matriptase antibody, we show that the ability of the Probody therapeutic to bind EGFR in vivo was dependent on protease activity. These results demonstrate that in vivo imaging of Probody therapeutic activation can be used for screening and characterization of protease activity in living animals, and provide a method that avoids some of the limitations of prior methods. This approach can improve our understanding of the activity of proteases in disease models and help to develop efficient strategies for cancer diagnosis and treatment. PMID:26546838

Physical Activity and Health: The Benefits of Physical Activity

MedlinePlus

... State and Local Programs Related Topics Diabetes Nutrition Physical Activity and Health Recommend on Facebook Tweet Share Compartir ... Your Chances of Living Longer The Benefits of Physical Activity Regular physical activity is one of the most ...

Assessing physical activity using wearable monitors: measures of physical activity.

PubMed

Butte, Nancy F; Ekelund, Ulf; Westerterp, Klaas R

2012-01-01

Physical activity may be defined broadly as "all bodily actions produced by the contraction of skeletal muscle that increase energy expenditure above basal level." Physical activity is a complex construct that can be classified into major categories qualitatively, quantitatively, or contextually. The quantitative assessment of physical activity using wearable monitors is grounded in the measurement of energy expenditure. Six main categories of wearable monitors are currently available to investigators: pedometers, load transducers/foot-contact monitors, accelerometers, HR monitors, combined accelerometer and HR monitors, and multiple sensor systems. Currently available monitors are capable of measuring total physical activity as well as components of physical activity that play important roles in human health. The selection of wearable monitors for measuring physical activity will depend on the physical activity component of interest, study objectives, characteristics of the target population, and study feasibility in terms of cost and logistics. Future development of sensors and analytical techniques for assessing physical activity should focus on the dynamic ranges of sensors, comparability for sensor output across manufacturers, and the application of advanced modeling techniques to predict energy expenditure and classify physical activities. New approaches for qualitatively classifying physical activity should be validated using direct observation or recording. New sensors and methods for quantitatively assessing physical activity should be validated in laboratory and free-living populations using criterion methods of calorimetry or doubly labeled water.

Is a Perceived Activity-Friendly Environment Associated with More Physical Activity and Fewer Screen-Based Activities in Adolescents?

PubMed

Kopcakova, Jaroslava; Dankulincova Veselska, Zuzana; Madarasova Geckova, Andrea; Bucksch, Jens; Nalecz, Hanna; Sigmundova, Dagmar; van Dijk, Jitse P; Reijneveld, Sijmen A

2017-01-03

Background: The aim of this study is to explore if perception of an activity-friendly environment is associated with more physical activity and fewer screen-based activities among adolescents. Methods: We collected self-reported data in 2014 via the Health Behavior in School-aged Children cross-sectional study from four European countries ( n = 13,800, mean age = 14.4, 49.4% boys). We explored the association of perceived environment (e.g., "There are other children nearby home to go out and play with") with physical activity and screen-based activities using a binary logistic regression model adjusted for age, gender, family affluence and country. Results: An environment perceived as activity-friendly was associated with higher odds that adolescents meet recommendations for physical activity (odds ratio (OR) for one standard deviation (SD) change = 1.11, 95% confidence interval (CI) 1.05-1.18) and lower odds for excessive screen-based activities (OR for 1 SD better = 0.93, 95% CI 0.88-0.98). Conclusions: Investment into an activity-friendly environment may support the promotion of active life styles in adolescence.

Antioxidant activity of taxifolin: an activity-structure relationship.

PubMed

Topal, Fevzi; Nar, Meryem; Gocer, Hulya; Kalin, Pınar; Kocyigit, Umit M; Gülçin, İlhami; Alwasel, Saleh H

2016-08-01

Taxifolin is a kind of flavanonol, whose biological ability. The objectives of this study were to investigate the antioxidants and antiradical activities of taxifolin by using different in vitro bioanalytical antioxidant methods including DMPD√(+), ABTS√(+), [Formula: see text], and DPPH√-scavenging effects, the total antioxidant influence, reducing capabilities, and Fe(2+)-chelating activities. Taxifolin demonstrated 81.02% inhibition of linoleic acid emulsion peroxidation at 30 µg/mL concentration. At the same concentration, standard antioxidants including trolox, α-tocopherol, BHT, and BHA exhibited inhibitions of linoleic acid emulsion as 88.57, 73.88, 94.29, and 90.12%, respectively. Also, taxifolin exhibited effective DMPD√(+), ABTS√(+), [Formula: see text], and DPPH√-scavenging effects, reducing capabilities, and Fe(2+)-chelating effects. The results obtained from this study clearly showed that taxifolin had marked antioxidant, reducing ability, radical scavenging and metal-chelating activities. Also, this study exhibits a scientific shore for the significant antioxidant activity of taxifolin and its structure-activity insight.

Dutch children and parents' views on active and non-active video gaming.

PubMed

De Vet, Emely; Simons, Monique; Wesselman, Maarten

2014-06-01

Active video games that require whole body movement to play the game may be an innovative health promotion tool to substitute sedentary pastime with more active time and may therefore contribute to children's health. To inform strategies aimed at reducing sedentary behavior by replacing non-active by active gaming, opinions about active and non-active video games are explored among 8- to 12-year-old children and their parents. Six qualitative, semi-structured focus groups were held with 8- to 12-year-old children (n = 46) and four with their parents (n = 19) at three different primary schools in The Netherlands. The focus groups with children discussed game preferences, gaming context and perceived game-related parenting. The focus groups with parents addressed considerations in purchasing video games, perceived positive and negative consequences of gaming, and game-related parenting. Both children and their parents were very positive about active video games and preferred active games over non-active games. Active video games were considered more social than non-active video games, and active games were played more often together with friends and family than non-active video games. Parenting practices did not differ for active and non-active video games, although some parents were less strict regarding active games. Two conditions for practical implementation were met: children enjoyed active video games, and parents were willing to buy active video games. Active video games were preferred to non-active video games, illustrating that using active video games is a promising health promotion tool to reduce sedentary pastime in youth.

Active Brownian particles with velocity-alignment and active fluctuations

NASA Astrophysics Data System (ADS)

Großmann, R.; Schimansky-Geier, L.; Romanczuk, P.

2012-07-01

We consider a model of active Brownian particles (ABPs) with velocity alignment in two spatial dimensions with passive and active fluctuations. Here, active fluctuations refers to purely non-equilibrium stochastic forces correlated with the heading of an individual active particle. In the simplest case studied here, they are assumed to be independent stochastic forces parallel (speed noise) and perpendicular (angular noise) to the velocity of the particle. On the other hand, passive fluctuations are defined by a noise vector independent of the direction of motion of a particle, and may account, for example, for thermal fluctuations. We derive a macroscopic description of the ABP gas with velocity-alignment interaction. Here, we start from the individual-based description in terms of stochastic differential equations (Langevin equations) and derive equations of motion for the coarse-grained kinetic variables (density, velocity and temperature) via a moment expansion of the corresponding probability density function. We focus here on the different impact of active and passive fluctuations on onset of collective motion and show how active fluctuations in the active Brownian dynamics can change the phase-transition behaviour of the system. In particular, we show that active angular fluctuations lead to an earlier breakdown of collective motion and to the emergence of a new bistable regime in the mean-field case.

Identification of fibrinogen-binding proteins of Aspergillus fumigatus using proteomic approach.

PubMed

Upadhyay, Santosh Kumar; Gautam, Poonam; Pandit, Hrishikesh; Singh, Yogendra; Basir, Seemi Farhat; Madan, Taruna

2012-03-01

Aspergillus fumigatus, the main etiological agent for various forms of human aspergillosis, gets access to the respiratory system of human host by inhalation of airborne conidia. These conidia possibly adhere to extracellular matrix (ECM) proteins. Among the ECM proteins involved in adherence, fibrinogen is thought to be crucial. Here, we studied whether A. fumigatus three-week culture filtrate (3wcf) proteins promote binding of A. fumigatus to ECM proteins and promote fungal growth. We observed that incubation of ECM with 3wcf proteins led to dose- and time-dependent increase in adherence of conidia to the ECM. In order to identify the catalogue of fibrinogen-binding A. fumigatus proteins, we carried out fibrinogen affinity blotting using two-dimensional gel electrophoresed 3wcf proteins. A total of 15 fibrinogen-binding protein spots corresponding to 7 unique proteins were identified in 3wcf using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF-TOF). Among these, 4 proteins, namely, beta-glucosidase, alpha-mannosidase, pectate lyase A and oryzin precursor were predicted to have cell wall or extracellular localization, whereas amidase family protein and two hypothetical proteins did not display the signal sequence. This study reports seven novel fibrinogen-binding proteins of A. fumigatus, some of which could be further explored for targeting the adhesion phenomenon as antifungal strategy.

29 CFR 779.208 - Auxiliary activities which are “related activities.”

Code of Federal Regulations, 2010 CFR

2010-07-01

... 29 Labor 3 2010-07-01 2010-07-01 false Auxiliary activities which are ârelated activities.â 779...; Enterprise Coverage Related Activities § 779.208 Auxiliary activities which are “related activities.” As... activities, such as central office and warehousing activities and bookkeeping, auditing, purchasing...

Weekday and weekend sedentary time and physical activity in differentially active children.

PubMed

Fairclough, Stuart J; Boddy, Lynne M; Mackintosh, Kelly A; Valencia-Peris, Alexandra; Ramirez-Rico, Elena

2015-07-01

To investigate whether weekday-weekend differences in sedentary time and specific intensities of physical activity exist among children categorised by physical activity levels. Cross-sectional observational study. Seven-day accelerometer data were obtained from 810 English children (n=420 girls) aged 10-11 years. Daily average minday(-1) spent in moderate to vigorous physical activity were calculated for each child. Sex-specific moderate to vigorous physical activity quartile cut-off values categorised boys and girls separately into four graded groups representing the least (Q1) through to the most active (Q4) children. Sex- and activity quartile-specific multilevel linear regression analyses analysed differences in sedentary time, light physical activity, moderate physical activity, vigorous physical activity, and moderate to vigorous physical activity between weekdays and weekends. On weekdays Q2 boys spent longer in light physical activity (p<0.05), Q1 (p<0.001), Q2 boys (p<0.01) did significantly more moderate physical activity, and Q1-Q3 boys accumulated significantly more vigorous physical activity and moderate to vigorous physical activity than at weekends. There were no significant differences in weekday and weekend sedentary time or physical activity for Q4 boys. On weekdays Q2 and Q3 girls accumulated more sedentary time (p<0.05), Q1 and Q2 girls did significantly more moderate physical activity (p<0.05), and Q1-Q3 girls engaged in more vigorous physical activity (p<0.05) and more moderate to vigorous physical activity (p<0.01) than at weekends. Q4 girls' sedentary time and physical activity varied little between weekdays and weekends. The most active children maintained their sedentary time and physical activity levels at weekends, while among less active peers weekend sedentary time and physical activity at all intensities was lower. Low active children may benefit most from weekend intervention strategies. Copyright © 2014 Sports Medicine Australia

Comparison of the performance of the activPAL Professional physical activity logger to a discrete accelerometer-based activity monitor.

PubMed

Godfrey, A; Culhane, K M; Lyons, G M

2007-10-01

The aim of this study was to assess the accuracy of the 'activPAL Professional' physical activity logger by comparing its output to that of a proven discrete accelerometer-based activity monitor during extended measurements on healthy subjects while performing activities of daily living (ADL). Ten healthy adults, with unrestricted mobility, wore both the activPAL and the discrete dual accelerometer (Analog Devices ADXL202)-based activity monitor that recorded in synchronization with each other. The accelerometer derived data were then compared to that generated by the activPAL and a complete statistical and error analysis was performed using a Matlab program. This program determined trunk and thigh inclination angles to distinguish between sitting/lying, standing and stepping for the discrete accelerometer device and amount of time spent on each activity. Analysis was performed on a second-by-second basis and then categorized at 15s intervals in direct comparison with the activPAL generated data. Of the total time monitored (approximately 60 h) the detection accuracies for static and dynamic activities were approximately 98%. In a population of healthy adults, the data obtained from the activPAL Professional physical activity logger for both static and dynamic activities showed a close match to a proven discrete accelerometer data with an offset of approximately 2% between the two systems.

Integration of Active Video Games in Extracurricular Activity at Schools

ERIC Educational Resources Information Center

Lee, Jung Eun; Huang, Charles; Pope, Zachary; Gao, Zan

2015-01-01

Active video games require players to be physically active. Dance Dance Revolution (DDR) is an interactive dancing game that requires fast-foot movement coordinated with energetic music and visuals. The Wii and Xbox Kinect games have also become good active video games for the promotion of physical activity participation. These games are much more…

Retro-binding thrombin active site inhibitors: identification of an orally active inhibitor of thrombin catalytic activity.

PubMed

Iwanowicz, Edwin J; Kimball, S David; Lin, James; Lau, Wan; Han, W-C; Wang, Tammy C; Roberts, Daniel G M; Schumacher, W A; Ogletree, Martin L; Seiler, Steven M

2002-11-04

A series of retro-binding inhibitors of human alpha-thrombin was prepared to elucidate structure-activity relationships (SAR) and optimize in vivo performance. Compounds 9 and 11, orally active inhibitors of thrombin catalytic activity, were identified to be efficacious in a thrombin-induced lethality model in mice.

Mutagenic activation reduces carcinogenic activity of ortho-aminoazotoluene for mouse liver.

PubMed

Ovchinnikova, L P; Bogdanova, L A; Kaledin, V I

2013-03-01

Pentachlorophenol (aromatic amine and azo stain metabolic stimulation inhibitor) reduced the hepatocarcinogenic activity of 4-aminoazobenzene and reduced that of ortho-aminoazotoluene in suckling mice. Both 4-aminoazobenzene and ortho-aminoazotoluene exhibited mutagenic activity in Ames' test in vitro on S. typhimurium TA 98 strain with activation with liver enzymes; this mutagenic activity was similarly suppressed by adding pentachlorophenol into activation medium. Induction of xenobiotic metabolism enzymes, stimulating the mutagenic activity of ortho-aminoazotoluene, suppressed its carcinogenic effect on mouse liver. Hence, ortho-aminotoluene (the initial compound), but not its mutagenic metabolites, was the direct active hepatocarcinogen for mice.

Assessing adult leisure activities: an extension of a self-report activity questionnaire.

PubMed

Jopp, Daniela S; Hertzog, Christopher

2010-03-01

Everyday leisure activities in adulthood and old age have been investigated with respect to constructs such as successful aging, an engaged lifestyle, and prevention of age-related cognitive decline. They also relate to mental health and have clinical value, as they can inform diagnosis and interventions. In the present study, the authors enhanced the content validity of the Victoria Longitudinal Study activity questionnaire by adding items on physical and social activities and validated a shortened version of the questionnaire. The proposed leisure activity model included 11 activity categories: 3 types of social activities (i.e., activities with close social partners, group-centered public activity, religious activities), physical activities, developmental activities, experiential activities, crafts, game playing, TV watching, travel, and technology use. Confirmatory factor analyses validated the proposed factor structure in 2 independent samples. A higher order model with a general activity factor fitted the activity factor correlations with relatively little loss of fit. Convergent and discriminant validity for the activity scales were supported by patterns of their correlations with education, health, depression, cognition, and personality. In sum, the scores derived from of the augmented Victoria Longitudinal Study activity questionnaire demonstrate good reliability, and validity evidence supports their use as measures of leisure activities in young, middle-aged, and older individuals. PsycINFO Database Record (c) 2010 APA, all rights reserved.

Physical Activity Guidelines

MedlinePlus

... use this site. health.gov Physical Activity Guidelines Physical Activity Physical activity is key to improving the health of the Nation. Based on the latest science, the Physical Activity Guidelines for Americans is an essential resource for ...

Elevation of oleate-activated phospholipase D activity during thymic atrophy

PubMed Central

Lee, Youngkyun; Song, Soo-Mee; Park, Heung Soon; Kim, Sungyeol; Koh, Eun-Hee; Choi, Myung Sun; Choi, Myung-Un

2002-01-01

Various phospholipases are thought to be associated with the in vitro apoptosis of thymocytes. In the present study, the in vivo phospholipase D (PLD) activity of rat thymus was studied after whole-body X-irradiation or injection of dexamethasone (DEX). Using exogenous [14C]dipalmitoyl phosphatidylcholine (PC) as the substrate, an elevation of oleate-activated PLD activity was observed during thymic atrophy. The activity increases were sevenfold at 48 hr after 5-Gy irradiation and fourfold at 72 hr after injection of 5 mg/kg DEX. The elevation of PLD activity appeared to parallel extensive thymus shrinkage. An increased level of thymic phosphatidic acid (PA), the presumed physiological product of PLD action on PC, was also detected. By comparing the acyl chains of PA with those of other phospholipids, PA appeared to originate from PC. To assess the role of PLD during thymic atrophy, thymocytes and stromal cells were isolated. Although thymocytes themselves exhibited significant PLD activation, the major elevation in PLD activity (greater than fourfold) was found in isolated stromal cells. PLD was also activated during in vitro phagocytosis of apoptotic thymocytes by the macrophage-like cell line P388D1. This in vitro phagocytosis was significantly inhibited by PLD action blockers, such as 2,3-diphosphoglycerate and 1-butanol. These observations strongly suggest that the alteration of oleate-activated PLD activity is part of an in vivo event in the progression of thymic atrophy, including phagocytic clearance of apoptotic thymocytes. PMID:12460188

Active nematic emulsions

PubMed Central

Hardoüin, Jérôme; Sagués, Francesc

2018-01-01

The formation of emulsions from multiple immiscible fluids is governed by classical concepts such as surface tension, differential chemical affinity and viscosity, and the action of surface-active agents. Much less is known about emulsification when one of the components is active and thus inherently not constrained by the laws of thermodynamic equilibrium. We demonstrate one such realization consisting in the encapsulation of an active liquid crystal (LC)–like gel, based on microtubules and kinesin molecular motors, into a thermotropic LC. These active nematic emulsions exhibit a variety of dynamic behaviors that arise from the cross-talk between topological defects separately residing in the active and passive components. Using numerical simulations, we show a feedback mechanism by which active flows continuously drive the passive defects that, in response, resolve the otherwise degenerated trajectories of the active defects. Our experiments show that the choice of surfactant, which stabilizes the active/passive interface, allows tuning the regularity of the self-sustained dynamic events. The hybrid active-passive system demonstrated here provides new perspectives for dynamic self-assembly driven by an active material but regulated by the equilibrium properties of the passive component. PMID:29740605

Active nematic emulsions.

PubMed

Guillamat, Pau; Kos, Žiga; Hardoüin, Jérôme; Ignés-Mullol, Jordi; Ravnik, Miha; Sagués, Francesc

2018-04-01

The formation of emulsions from multiple immiscible fluids is governed by classical concepts such as surface tension, differential chemical affinity and viscosity, and the action of surface-active agents. Much less is known about emulsification when one of the components is active and thus inherently not constrained by the laws of thermodynamic equilibrium. We demonstrate one such realization consisting in the encapsulation of an active liquid crystal (LC)-like gel, based on microtubules and kinesin molecular motors, into a thermotropic LC. These active nematic emulsions exhibit a variety of dynamic behaviors that arise from the cross-talk between topological defects separately residing in the active and passive components. Using numerical simulations, we show a feedback mechanism by which active flows continuously drive the passive defects that, in response, resolve the otherwise degenerated trajectories of the active defects. Our experiments show that the choice of surfactant, which stabilizes the active/passive interface, allows tuning the regularity of the self-sustained dynamic events. The hybrid active-passive system demonstrated here provides new perspectives for dynamic self-assembly driven by an active material but regulated by the equilibrium properties of the passive component.

A Youth Compendium of Physical Activities: Activity Codes and Metabolic Intensities

PubMed Central

BUTTE, NANCY F.; WATSON, KATHLEEN B.; RIDLEY, KATE; ZAKERI, ISSA F.; MCMURRAY, ROBERT G.; PFEIFFER, KARIN A.; CROUTER, SCOTT E.; HERRMANN, STEPHEN D.; BASSETT, DAVID R.; LONG, ALEXANDER; BERHANE, ZEKARIAS; TROST, STEWART G.; AINSWORTH, BARBARA E.; BERRIGAN, DAVID; FULTON, JANET E.

2018-01-01

ABSTRACT Purpose A Youth Compendium of Physical Activities (Youth Compendium) was developed to estimate the energy costs of physical activities using data on youth only. Methods On the basis of a literature search and pooled data of energy expenditure measurements in youth, the energy costs of 196 activities were compiled in 16 activity categories to form a Youth Compendium of Physical Activities. To estimate the intensity of each activity, measured oxygen consumption (V˙O2) was divided by basal metabolic rate (Schofield age-, sex-, and mass-specific equations) to produce a youth MET (METy). A mixed linear model was developed for each activity category to impute missing values for age ranges with no observations for a specific activity. Results This Youth Compendium consists of METy values for 196 specific activities classified into 16 major categories for four age-groups, 6–9, 10–12, 13–15, and 16–18 yr. METy values in this Youth Compendium were measured (51%) or imputed (49%) from youth data. Conclusion This Youth Compendium of Physical Activities uses pediatric data exclusively, addresses the age dependency of METy, and imputes missing METy values and thus represents advancement in physical activity research and practice. This Youth Compendium will be a valuable resource for stakeholders interested in evaluating interventions, programs, and policies designed to assess and encourage physical activity in youth. PMID:28938248

Vestibular activation of sympathetic nerve activity

NASA Technical Reports Server (NTRS)

Ray, C. A.; Carter, J. R.

2003-01-01

AIM: The vestibulosympathetic reflex refers to sympathetic nerve activation by the vestibular system. Animal studies indicate that the vestibular system assists in blood pressure regulation during orthostasis. Although human studies clearly demonstrate activation of muscle sympathetic nerve activity (MSNA) during engagement of the otolith organs, the role of the vestibulosympathetic reflex in maintaining blood pressure during orthostasis is not well-established. Examination of the vestibulosympathetic reflex with other cardiovascular reflexes indicates that it is a powerful and independent reflex. Ageing, which is associated with an increased risk for orthostatic hypotension, attenuates the vestibulosympathetic reflex. The attenuated reflex is associated with a reduction in arterial pressure. CONCLUSION: These findings suggest that the vestibulosympathetic reflex assists in blood pressure regulation in humans, but future studies examining this reflex in other orthostatically intolerant populations are necessary to address this hypothesis.

Increasing Youth Physical Activity with Activity Calendars

ERIC Educational Resources Information Center

Eckler, Seth

2016-01-01

Physical educators often struggle with ways to get their students to be active beyond the school day. One strategy to accomplish this is the use of physical activity calendars (PACs). The purpose of this article is to support the use of PACs and give practical advice for creating effective PACs.

Activating AMP-activated protein kinase by an α1 selective activator compound 13 attenuates dexamethasone-induced osteoblast cell death

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guo, Shiguang; Mao, Li; Ji, Feng, E-mail: huaiaifengjidr@163.com

Excessive glucocorticoid (GC) usage may lead to non-traumatic femoral head osteonecrosis. Dexamethasone (Dex) exerts cytotoxic effect to cultured osteoblasts. Here, we investigated the potential activity of Compound 13 (C13), a novel α1 selective AMP-activated protein kinase (AMPK) activator, against the process. Our data revealed that C13 pretreatment significantly attenuated Dex-induced apoptosis and necrosis in both osteoblastic-like MC3T3-E1 cells and primary murine osteoblasts. AMPK activation mediated C13′ cytoprotective effect in osteoblasts. The AMPK inhibitor Compound C, shRNA-mediated knockdown of AMPKα1, or dominant negative mutation of AMPKα1 (T172A) almost abolished C13-induced AMPK activation and its pro-survival effect in osteoblasts. On the othermore » hand, forced AMPK activation by adding AMPK activator A-769662 or exogenous expression a constitutively-active (ca) AMPKα1 (T172D) mimicked C13's actions and inhibited Dex-induced osteoblast cell death. Meanwhile, A-769662 or ca-AMPKα1 almost nullified C13's activity in osteoblast. Further studies showed that C13 activated AMPK-dependent nicotinamide adenine dinucleotide phosphate (NADPH) pathway to inhibit Dex-induced reactive oxygen species (ROS) production in MC3T3-E1 cells and primary murine osteoblasts. Such effects by C13 were almost reversed by Compound C or AMPKα1 depletion/mutation. Together, these results suggest that C13 alleviates Dex-induced osteoblast cell death via activating AMPK signaling pathway. - Highlights: • Compound 13 (C13) attenuates dexamethasone (Dex)-induced osteoblast cell death. • C13-induced cytoprotective effect against Dex in osteoblasts requires AMPK activation. • Forced AMPK activation protects osteoblasts from Dex, nullifying C13's activities. • C13 increases NADPH activity and inhibits Dex-induced oxidative stress in osteoblasts.« less

Adolescents' Views on Active and Non-Active Videogames: A Focus Group Study.

PubMed

Simons, Monique; de Vet, Emely; Hoornstra, Sjoukje; Brug, Johannes; Seidell, Jaap; Chinapaw, Mai

2012-06-01

Active games require whole-body movement and may be an innovative tool to substitute sedentary pastime with more active time and may therefore contribute to adolescents' health. To inform strategies aimed at reducing sedentary behavior by replacing non-active with active gaming, perceptions and context of active and non-active gaming are explored. Six focus groups were conducted with adolescents 12-16 years old representing a range of education levels. A semistructured question route was used containing questions about perceptions and the context of gaming. The adolescents had positive attitudes toward active gaming, especially the social interactive aspect, which was greatly appreciated. A substantial number of adolescents enjoyed non-active games more than active ones, mainly because of better game controls and more diversity in non-active games. Active games were primarily played when there was a social gathering. Few game-related rules and restrictions at home were reported. Given the positive attitudes of adolescents and the limited restrictions for gaming at home, active videogames may potentially be used in a home setting as a tool to reduce sedentary behavior. However, to make active games as appealing as non-active games, attention should be paid to the quality, diversity, and sustainability of active games, as these aspects are currently inferior to those of traditional non-active games.

Assessing Adult Leisure Activities: An Extension of a Self-Report Activity Questionnaire

PubMed Central

Jopp, Daniela; Hertzog, Christopher

2009-01-01

Everyday leisure activities in adulthood and old age have been investigated with respect to constructs such as successful aging, an engaged lifestyle, and prevention of age-related cognitive decline. They also relate to mental health and have clinical value as they can inform diagnosis and interventions. In the present study, we enhanced the content validity of the Victoria Longitudinal Study activity questionnaire by adding items on physical and social activities, and validated a shortened version of the questionnaire. Our proposed leisure activity model included 11 activity categories: three types of social activities (i.e., activities with close social partners, group-centered public activity, religious activities), physical, developmental, and experiential activities, crafts, game playing, TV watching, travel, and technology use. Confirmatory factor analyses validated the proposed factor structure in two independent samples. A higher-order model with a general activity factor fitted the activity factor correlations with relatively little loss of fit. Convergent and discriminant validity for the activity scales were supported by patterns of their correlations with education, health, depression, cognition, and personality. In sum, the scores derived from of the augmented VLS activity questionnaire demonstrate good reliability, and validity evidence supports their use as measure of leisure activities in young, middle-aged, and older individuals. PMID:20230157

What Is an Activity? Appropriating an Activity-Centric System

NASA Astrophysics Data System (ADS)

Yarosh, Svetlana; Matthews, Tara; Moran, Thomas P.; Smith, Barton

Activity-Centric Computing (ACC) systems seek to address the fragmentation of office work across tools and documents by allowing users to organize work around the computational construct of an Activity. Defining and structuring appropriate Activities within a system poses a challenge for users that must be overcome in order to benefit from ACC support. We know little about how knowledge workers appropriate the Activity construct. To address this, we studied users’ appropriation of a production-quality ACC system, Lotus Activities, for everyday work by employees in a large corporation. We contribute to a better understanding of how users articulate their individual and collaborative work in the system by providing empirical evidence of their patterns of appropriation. We conclude by discussing how our findings can inform the design of other ACC systems for the workplace.

Active compounds, antioxidant activity and α-glucosidase inhibitory activity of different varieties of Chaenomeles fruits.

PubMed

Miao, Jing; Li, Xia; Zhao, Chengcheng; Gao, Xiaoxiao; Wang, Ying; Gao, Wenyuan

2018-05-15

Chaenomeles is an important source for food industry in China, and its planting area is expanding year by year. This study was conducted to evaluate different varieties of Chaenomeles by comparing the chemical compositions, antioxidant activity and α-glucosidase inhibitory activity of peels and fleshes from twelve varieties of Chaenomeles. In the results, peels of Chaenomeles contain more phenolics, flavonoids and triterpenes, and show better antioxidant activity and α-glucosidase inhibitory activity than their fleshes. All varieties of Chaenomeles perform different depend on cultivar and climatic conditions. Oleanolic acid, ursolic acid, protocatechuic acid, rutin, catechin, caffeic acid, syringic acid, epicatechin, hyperin, quercetin, kaempferol and chlorogenic acid are main active compounds in Chaenomeles. Zheng'an, Liufu, Zimugua1, Qijiang and Changjun get Top five scores. This is the first study on the peels and fleshes of twelve varieties of Chaenomeles, and it gives insights into variety selection in the planting and production of Chaenomeles. Copyright © 2017 Elsevier Ltd. All rights reserved.

Immotile Active Matter: Activity from Death and Reproduction.

PubMed

Kalziqi, Arben; Yanni, David; Thomas, Jacob; Ng, Siu Lung; Vivek, Skanda; Hammer, Brian K; Yunker, Peter J

2018-01-05

Unlike equilibrium atomic solids, biofilms-soft solids composed of bacterial cells-do not experience significant thermal fluctuations at the constituent level. However, living cells stochastically reproduce and die, provoking a mechanical response. We investigate the mechanical consequences of cellular death and reproduction by measuring surface-height fluctuations of biofilms containing two mutually antagonistic strains of Vibrio cholerae that kill one another on contact via the type VI secretion system. While studies of active matter typically focus on activity via constituent mobility, here, activity is mediated by reproduction and death events in otherwise immobilized cells. Biofilm surface topography is measured in the nearly homeostatic limit via white light interferometry. Although biofilms are far from equilibrium systems, measured surface-height fluctuation spectra resemble the spectra of thermal permeable membranes but with an activity-mediated effective temperature, as predicted by Risler, Peilloux, and Prost [Phys. Rev. Lett. 115, 258104 (2015)PRLTAO0031-900710.1103/PhysRevLett.115.258104]. By comparing the activity of killer strains of V. cholerae with that of genetically modified strains that cannot kill each other and validating with individual-based simulations, we demonstrate that extracted effective temperatures increase with the amount of death and reproduction and that death and reproduction can fluidize biofilms. Together, these observations demonstrate the unique physical consequences of activity mediated by death and reproduction events.

Immotile Active Matter: Activity from Death and Reproduction

NASA Astrophysics Data System (ADS)

Kalziqi, Arben; Yanni, David; Thomas, Jacob; Ng, Siu Lung; Vivek, Skanda; Hammer, Brian K.; Yunker, Peter J.

2018-01-01

Unlike equilibrium atomic solids, biofilms—soft solids composed of bacterial cells—do not experience significant thermal fluctuations at the constituent level. However, living cells stochastically reproduce and die, provoking a mechanical response. We investigate the mechanical consequences of cellular death and reproduction by measuring surface-height fluctuations of biofilms containing two mutually antagonistic strains of Vibrio cholerae that kill one another on contact via the type VI secretion system. While studies of active matter typically focus on activity via constituent mobility, here, activity is mediated by reproduction and death events in otherwise immobilized cells. Biofilm surface topography is measured in the nearly homeostatic limit via white light interferometry. Although biofilms are far from equilibrium systems, measured surface-height fluctuation spectra resemble the spectra of thermal permeable membranes but with an activity-mediated effective temperature, as predicted by Risler, Peilloux, and Prost [Phys. Rev. Lett. 115, 258104 (2015), 10.1103/PhysRevLett.115.258104]. By comparing the activity of killer strains of V. cholerae with that of genetically modified strains that cannot kill each other and validating with individual-based simulations, we demonstrate that extracted effective temperatures increase with the amount of death and reproduction and that death and reproduction can fluidize biofilms. Together, these observations demonstrate the unique physical consequences of activity mediated by death and reproduction events.

Complete Reconstitution of the Vancomycin-Intermediate Staphylococcus aureus Phenotype of Strain Mu50 in Vancomycin-Susceptible S. aureus

PubMed Central

Sekine, Miwa; Hishinuma, Tomomi; Aiba, Yoshifumi; Hiramatsu, Keiichi

2016-01-01

Complete reconstitution of the vancomycin-intermediate Staphylococcus aureus (VISA) phenotype of strain Mu50 was achieved by sequentially introducing mutations into six genes of vancomycin-susceptible S. aureus (VSSA) strain N315ΔIP. The six mutated genes were detected in VISA strain Mu50 but not in N315ΔIP. Introduction of the mutation Ser329Leu into vraS, encoding the sensor histidine kinase of the vraSR two-component regulatory (TCR) system, and another mutation, Glu146Lys, into msrR, belonging to the LytR-CpsA-Psr (LCP) family, increased the level of vancomycin resistance to that detected in heterogeneous vancomycin-intermediate S. aureus (hVISA) strain Mu3. Introduction of two more mutations, Asn197Ser into graR of the graSR TCR system and His481Tyr into rpoB, encoding the β subunit of RNA polymerase, converted the hVISA strain into a VISA strain with the same level of vancomycin resistance as Mu50. Surprisingly, however, the constructed quadruple mutant strain ΔIP4 did not have a thickened cell wall, a cardinal feature of the VISA phenotype. Subsequent study showed that cell wall thickening was an inducible phenotype in the mutant strain, whereas it was a constitutive one in Mu50. Finally, introduction of the Ala297Val mutation into fdh2, which encodes a putative formate dehydrogenase, or a 67-amino-acid sequence deletion into sle1 [sle1(Δ67aa)], encoding the hydrolase of N-acetylmuramyl-l-alanine amidase in the peptidoglycan, converted inducible cell wall thickening into constitutive cell wall thickening. sle1(Δ67aa) was found to cause a drastic decrease in autolysis activity. Thus, all six mutated genes required for acquisition of the VISA phenotype were directly or indirectly involved in the regulation of cell physiology. The VISA phenotype seemed to be achieved through multiple genetic events accompanying drastic changes in cell physiology. PMID:27067329

Macrophage activating activity of pyrrole alkaloids from Morus alba fruits.

PubMed

Kim, Seon Beom; Chang, Bo Yoon; Jo, Yang Hee; Lee, Sang Hoon; Han, Sang-Bae; Hwang, Bang Yeon; Kim, Sung Yeon; Lee, Mi Kyeong

2013-01-09

The fruits of Morus alba have been traditionally used as a tonic to enhance immune responses. The macrophage activating constituents of Morus alba fruits were purified using various column chromatography techniques. The structures of isolated compounds were determined on the basis of spectroscopic data interpretation such as 1D and 2D NMR analysis. The macrophage activating activities of isolated compounds were evaluated by measuring the production of nitric oxide, TNF-α and IL-12 in RAW 264.7 cells. The phagocytic activity was also evaluated. Five pyrrole alkaloids, 5-(hydroxymethyl)-1H-pyrrole-2-carboxaldehyde (1), 2-formyl-1H-pyrrole-1-butanoic acid (2), 2-formyl-5-(hydroxymethyl)-1H-pyrrole-1-butanoic acid (3), 2-formyl-5-(methoxymethyl)-1H-pyrrole-1-butanoic acid (4) and Morrole A (5) were isolated from the fruits of Morus alba. Morrole A (5) is first reported in nature and other pyrrole alkaloids (1-4) are first reported from Morus species. Among the isolated compounds, compounds 3 and 4 significantly activated macrophage activity by the enhancement of nitric oxide, TNF-α and IL-12 production, and the stimulation of phagocytic activity in RAW 264.7 cells. Pyrrole alkaloids, including a new compound, were isolated from Morus alba fruits. These compounds activated macrophage activity in RAW 264.7 cells. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Is constipation associated with decreased physical activity in normally active subjects?

PubMed

Tuteja, Ashok K; Talley, Nicholas J; Joos, Sandra K; Woehl, James V; Hickam, David H

2005-01-01

The effectiveness of physical activity in the management of constipation remains controversial. We examined the associations among physical activity, constipation, and quality of life (QoL) in a population of employed adults to determine whether the risk of constipation is related to physical activity. A total of 1,069 employees (age range 24-77) of the Veterans Affairs (VA) Black Hills Health Care System were mailed validated questionnaires (response rate 72%), inquiring about bowel habits, QoL (SF 36), and physical activity (modified Baecke questionnaire). Constipation was defined using the Rome I criteria. One hundred and forty (19.4%, 95% CI 16.2-22.4) employees reported constipation. The average total physical activity and all subscales of physical activity were not significantly different in subjects with and without constipation (all p > or = 0.2). Subjects with constipation had lower QoL scores than subjects without constipation, and physical activity was positively correlated with physical functioning and health perception. Physical activity appears to be unrelated to the risk of constipation in employed adults, but higher physical activity was associated with improved QoL. Recommendations to increase physical activity may not alter symptoms of constipation but may improve overall well-being.

Comparison of passive and active leisure activities and life satisfaction with aging.

PubMed

Cho, Dongwook; Post, Jay; Kim, Sung Kyeom

2018-03-01

Many older adults face limitations to participating in active leisure activities as a result of their physical constraints from aging. Passive leisure activities become alternative leisure activities for older adults as a result of limited physical capacity. The present study sought to determine whether there exists a difference in the frequency of participation in passive and active leisure activities, and the effect of participation in passive and active leisure activities on the life satisfaction level of old adults. A total of 460 participants aged 60-95 years were randomly selected from 21 sites in the USA. The Life Satisfaction Index - Z and the Meaningful Activity Participation Assessment were analyzed to examine older adults' life satisfaction and frequency of active or passive activities. The results showed that participation in passive leisure activities, such reading, talking on the telephone and watching TV/listening to the radio, is more frequent among older adults (P = 0.000). The regression coefficient found that club/organization or volunteering (P = 0.008), homemaking/maintenance (P = 0.017) and traveling (P = 0.017) for active leisure activities were statistically significant predictors of Life Satisfaction Index - Z for older adults. The current study shows that older adults spent much more times participating in passive leisure activities, such as radio/watching TV, talking on the phone and reading. The result also showed that active leisure activities, such as club/organization or volunteering, home making/maintenance and traveling, were significant predictors of life satisfaction for older adults controlling for covariates. The current study suggests marketing and programming plans to overcome the constraints that influence older adults' life satisfaction. Geriatr Gerontol Int 2018; 18: 380-386. © 2017 Japan Geriatrics Society.

Transcriptional activation of peroxisome proliferator-activated receptor-{gamma} requires activation of both protein kinase A and Akt during adipocyte differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Sang-pil; Ha, Jung Min; Yun, Sung Ji

2010-08-13

Research highlights: {yields} Elevated cAMP activates both PKA and Epac. {yields} PKA activates CREB transcriptional factor and Epac activates PI3K/Akt pathway via Rap1. {yields} Akt modulates PPAR-{gamma} transcriptional activity in concert with CREB. -- Abstract: Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is required for the conversion of pre-adipocytes. However, the mechanism underlying activation of PPAR-{gamma} is unclear. Here we showed that cAMP-induced activation of protein kinase A (PKA) and Akt is essential for the transcriptional activation of PPAR-{gamma}. Hormonal induction of adipogenesis was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), by a protein kinase A (PKA) inhibitor (H89), and by amore » Rap1 inhibitor (GGTI-298). Transcriptional activity of PPAR-{gamma} was markedly enhanced by 3-isobutyl-1-methylxanthine (IBMX), but not insulin and dexamethasone. In addition, IBMX-induced PPAR-{gamma} transcriptional activity was blocked by PI3K/Akt, PKA, or Rap1 inhibitors. 8-(4-Chlorophenylthio)-2'-O-methyl-cAMP (8-pCPT-2'-O-Me-cAMP) which is a specific agonist for exchanger protein directly activated by cAMP (Epac) significantly induced the activation of Akt. Furthermore, knock-down of Akt1 markedly attenuated PPAR-{gamma} transcriptional activity. These results indicate that both PKA and Akt signaling pathways are required for transcriptional activation of PPAR-{gamma}, suggesting post-translational activation of PPAR-{gamma} might be critical step for adipogenic gene expression.« less

Activity Engagement and Activity-Related Experiences: The Role of Personality.

PubMed

Newton, Nicky J; Pladevall-Guyer, Jana; Gonzalez, Richard; Smith, Jacqui

2016-08-12

The associations of personality with activity participation and well-being have been well studied. However, less is known concerning the relationship between personality and specific aspects of activity engagement in older adults. We conducted a fine-grained examination of the effects of extraversion and conscientiousness on reported activity engagement-which we define as participation, time allocated, and affective experience-during 8 everyday activities. Data were obtained using a day reconstruction measure from a subgroup of participants in the 2012 Health and Retirement Study (HRS: N = 5,484; mean age = 67.98 years). We found mixed support for hypotheses suggesting that specific personality traits would be associated with activity participation, time allocated, and activity-affective experience. For example, extraverts were more likely to socialize and experienced higher socializing-related positive affect, but did not spend more time socializing. Results are discussed in light of the value of including personality in, and its contribution to, studies of activity engagement in later life. In addition, the need to acknowledge the complexity of the concept of activity engagement in future research is highlighted. © The Author 2016. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Multi-day activity scheduling reactions to planned activities and future events in a dynamic model of activity-travel behavior

NASA Astrophysics Data System (ADS)

Nijland, Linda; Arentze, Theo; Timmermans, Harry

2014-01-01

Modeling multi-day planning has received scarce attention in activity-based transport demand modeling so far. However, new dynamic activity-based approaches are being developed at the current moment. The frequency and inflexibility of planned activities and events in activity schedules of individuals indicate the importance of incorporating those pre-planned activities in the new generation of dynamic travel demand models. Elaborating and combining previous work on event-driven activity generation, the aim of this paper is to develop and illustrate an extension of a need-based model of activity generation that takes into account possible influences of pre-planned activities and events. This paper describes the theory and shows the results of simulations of the extension. The simulation was conducted for six different activities, and the parameter values used were consistent with an earlier estimation study. The results show that the model works well and that the influences of the parameters are consistent, logical, and have clear interpretations. These findings offer further evidence of face and construct validity to the suggested modeling approach.

Toxicity tests, antioxidant activity, and antimicrobial activity of chitosan

NASA Astrophysics Data System (ADS)

Kurniasih, M.; Purwati; Dewi, R. S.

2018-04-01

Chitosan is a naturally occurring cationic biopolymer, obtained by alkaline deacetylation of chitin. This research aims to investigate the toxicity, antioxidant activity and antibacterial activity of chitosan from shrimp chitin. In this study, chitin extracted from shrimp waste material. Chitin is then deacetylation with 60% NaOH so that chitosan produced. Degrees of deacetylation, molecular weight, toxicity test, antioxidant activity and antimicrobial activity of chitosan then evaluated. Toxicity test using Brine Shrimp Lethality Test. The antioxidant analysis was performed using DPPH method (2, 2-diphenyl-1-picrylhydrazyl) and FTC method (ferric thiocyanate) in which the radical formed will reduce Ferro to Ferri resulting in a complex with thiocyanate. To determine the antibacterial activity of Staphylococcus aureus, antifungal in Candida albicans and Aspergillus niger by measuring antimicrobial effects and minimum inhibitory concentrations (MIC). Based on the result of research, the value of degrees of deacetylation, molecular weight, and LC50 values of chitosan synthesis was 94,32, 1052.93 g/mol and 1364.41 ppm, respectively. In general, the antioxidative activities increased as the concentration of chitosan increased. MIC value of chitosan against S. aureus, C. albicans, and A. niger was 10 ppm, 15.6 ppm, and 5 ppm, respectively.

Differential Activity-Driven Instabilities in Biphasic Active Matter

NASA Astrophysics Data System (ADS)

Weber, Christoph A.; Rycroft, Chris H.; Mahadevan, L.

2018-06-01

Active stresses can cause instabilities in contractile gels and living tissues. Here we provide a generic hydrodynamic theory that treats these systems as a mixture of two phases of varying activity and different mechanical properties. We find that differential activity between the phases causes a uniform mixture to undergo a demixing instability. We follow the nonlinear evolution of the instability and characterize a phase diagram of the resulting patterns. Our study complements other instability mechanisms in mixtures driven by differential adhesion, differential diffusion, differential growth, and differential motion.

Physical activity behavior and related characteristics of highly active eighth-grade girls.

PubMed

Taverno Ross, Sharon E; Dowda, Marsha; Beets, Michael W; Pate, Russell R

2013-06-01

Although girls are generally less physically active than boys, some girls regularly engage in high levels of physical activity (PA); however, very little is known about these girls and how they differ from those who are less physically active. This study examined the PA behavior and related characteristics of highly active adolescent girls and compared them with those who are less active. Data from 1,866 eighth-grade girls from six field centers across the United States participating in the Trial of Activity for Adolescent Girls (TAAG) were included in the present analysis. Mixed-model analysis of variance examined differences in sociodemographic, anthropometric, psychosocial, and physical activity (accelerometry and self-report) variables between high- and low-active girls; effect sizes were calculated for the differences. High-active girls were taller, had lower body mass indices and body fat, and were less sedentary. High-active girls scored higher on self-efficacy, enjoyment of PA, self-management strategies, outcome-expectancy value, and support from family and friends than low-active girls. Low-active girls participated in more leisure time and educational sedentary activities than high-active girls. High-active girls participated in more PA classes/lessons outside of school, team sports, and individual sports. They were also more likely to participate in sports in an organized setting in the community or at school than low-active girls. Health promotion efforts should focus on decreasing the amount of time girls spend in sedentary activities and replacing that time with organized PA opportunities; such efforts should seek to minimize perceived barriers and increase self-efficacy and support for PA. Copyright © 2013 Society for Adolescent Health and Medicine. Published by Elsevier Inc. All rights reserved.

Physical activity energy expenditure in Dutch adolescents: contribution of active transport to school, physical education, and leisure time activities.

PubMed

Slingerland, Menno; Borghouts, Lars B; Hesselink, Matthijs K C

2012-05-01

Detailed knowledge about physical activity energy expenditure (PAEE) can guide the development of school interventions aimed at reducing overweight in adolescents. However, relevant components of PAEE have never been objectively quantified in this population. This study investigated the contribution of active transport to and from school, physical education (PE), and leisure time activities to total PAEE during a regular school week in adolescents. Seventy-three adolescents (mean age: 15.7 years) wore an individually calibrated combined heart rate-acceleration monitor and kept an activity diary during a regular school week. Branched equation modeling was used to calculate PAEE of the specific activity categories, and their relative contribution to total PAEE was determined. Active transport and PE contributed 30.0% and 17.4%, respectively, to school-related PAEE. Active transport to and from school contributed 15% to total PAEE. Youth with a high physical activity level (PAL) spent 4 hours less in sedentary behavior than subjects with a medium or low PAL (F = 77.415 (2.70), p < .001) and had higher PAEE during leisure time sports (F = 9.135 (2.70), p < .001) and work-related activities (F = 10.583 (2.70), p < .001) than youth with medium or low PAL values. Active transport and PE contribute significantly to PAEE during school hours in adolescents. To achieve an increase in total PAEE in the least active group of adolescents, promising strategies might be to reduce inactive behavior, increase participation in leisure time sports, and possibly to replace inactive for active jobs. © 2012, American School Health Association.

Nursing activities score.

PubMed

Miranda, Dinis Reis; Nap, Raoul; de Rijk, Angelique; Schaufeli, Wilmar; Iapichino, Gaetano

2003-02-01

The instruments used for measuring nursing workload in the intensive care unit (e.g., Therapeutic Intervention Scoring System-28) are based on therapeutic interventions related to severity of illness. Many nursing activities are not necessarily related to severity of illness, and cost-effectiveness studies require the accurate evaluation of nursing activities. The aim of the study was to determine the nursing activities that best describe workload in the intensive care unit and to attribute weights to these activities so that the score describes average time consumption instead of severity of illness. To define by consensus a list of nursing activities, to determine the average time consumption of these activities by use of a 1-wk observational cross-sectional study, and to compare these results with those of the Therapeutic Intervention Scoring System-28. A total of 99 intensive care units in 15 countries. Consecutive admissions to the intensive care units. Daily recording of nursing activities at a patient level and random multimoment recording of these activities. A total of five new items and 14 subitems describing nursing activities in the intensive care unit (e.g., monitoring, care of relatives, administrative tasks) were added to the list of therapeutic interventions in Therapeutic Intervention Scoring System-28. Data from 2,041 patients (6,451 nursing days and 127,951 multimoment recordings) were analyzed. The new activities accounted for 60% of the average nursing time; the new scoring system (Nursing Activities Score) explained 81% of the nursing time (vs. 43% in Therapeutic Intervention Scoring System-28). The weights in the Therapeutic Intervention Scoring System-28 are not derived from the use of nursing time. Our study suggests that the Nursing Activities Score measures the consumption of nursing time in the intensive care unit. These results should be validated in independent databases.

[Positive Activities Campaign.

ERIC Educational Resources Information Center

Substance Abuse and Mental Health Services Administration (DHHS/PHS), Rockville, MD. Center for Substance Abuse Prevention.

This packet contains four pamphlets that are part of a campaign to encourage adults to provide and promote positive activities for youth and to serve as role models for young people. "Positive Activities: A Campaign for Youth" includes information on what positive activities are, how to get involved in helping to provide positive activities for…

Active Volcanism on Io: Global Distribution and Variations in Activity

USGS Publications Warehouse

Lopes-Gautier, R.; McEwen, A.S.; Smythe, W.B.; Geissler, P.E.; Kamp, L.; Davies, A.G.; Spencer, J.R.; Keszthelyi, L.; Carlson, R.; Leader, F.E.; Mehlman, R.; Soderblom, L.

1999-01-01

Io's volcanic activity has been monitored by instruments aboard the Galileo spacecraft since June 28, 1996. We present results from observations by the near-infrared mapping spectrometer (NIMS) for the first 10 orbits of Galileo, correlate them with results from the Solid State Imaging System (SSI) and from groundbased observations, and compare them to what was known about Io's volcanic activity from observations made during the two Voyager flybys in 1979. A total of 61 active volcanic centers have been identified from Voyager, groundbased, and Galileo observations. Of these, 41 are hot spots detected by NIMS and/or SSI. Another 25 locations were identified as possible active volcanic centers, mostly on the basis of observed surface changes. Hot spots are correlated with surface colors, particularly dark and red deposits, and generally anti-correlated with white, SO2-rich areas. Surface features corresponding to the hot spots, mostly calderas or flows, were identified from Galileo and Voyager images. Hot spot temperatures obtained from both NIMS and SSI are consistent with silicate volcanism, which appears to be widespread on Io. Two types of hot spot activity are present: persistent-type activity, lasting from months to years, and sporadic events, which may represent either short-lived activity or low-level activity that occasionally flares up. Sporadic events are not often detected, but may make an important contribution to Io's heat flow and resurfacing. The distribution of active volcanic centers on the surface does not show any clear correlation with latitude, longitude, Voyager-derived global topography, or heat flow patterns predicted by the asthenosphere and deep mantle tidal dissipation models. However, persistent hot spots and active plumes are concentrated toward lower latitudes, and this distribution favors the asthenosphere rather than the deep mantle tidal dissipation model. ?? 1999 Academic Press.

Association between sedentary behavior, physical activity, and obesity: inactivity among active kids.

PubMed

Wong, Suzy L; Leatherdale, Scott T

2009-01-01

Sedentary behavior and physical activity are not mutually exclusive behaviors. The relative risk of overweight for adolescents who are highly sedentary and highly physically active is unclear. A better understanding of the relationship between sedentary behaviors, physical activity, and body mass index (BMI) would provide insight for developing interventions to prevent or reduce overweight. Using the physical activity module of the School Health Action, Planning and Evaluation System (SHAPES), we collected data from 25,060 students in grades 9 through 12 from 76 secondary schools in Ontario, Canada. Sex-specific logistic regression analyses were performed to examine how BMI, weight perceptions, social influences, team sports participation, and smoking behavior were associated with being 1) high active-high sedentary, 2) low active-low sedentary, and 3) low active-high sedentary. Low active-high sedentary boys were more likely to be overweight than high active-low sedentary boys (adjusted odds ratio [AOR], 1.60; 95% confidence interval [CI], 1.01-2.58). When compared with high active-low sedentary girls, girls who were low active-high sedentary (OR, 2.24; 95% CI, 1.23-4.09) or high active-high sedentary (OR, 1.91; 95% CI, 1.01-3.61) were more likely to be overweight. Sedentary behavior may moderate the relationship between physical activity and overweight. Developing a better understanding of sedentary behavior in relation to physical activity and overweight is critical for preventing and reducing overweight among youth.

Worksite physical activity policies and environments in relation to employee physical activity.

PubMed

Crespo, Noe C; Sallis, James F; Conway, Terry L; Saelens, Brian E; Frank, Lawrence D

2011-01-01

Examine associations between worksite physical activity promotion strategies and employees' physical activity and sedentary behaviors. Cross-sectional. Seattle-King County, Washington and Baltimore, Maryland-Washington, D.C. regions. Adults working outside the home (n = 1313). Mean age was 45 ± 10 years, 75.8% of participants were non-Hispanic white, 56% were male, and 51% had income ≥$70,000/year. Participants reported demographic characteristics and presence/absence of nine physical activity promotion environment and policy strategies in their work environment (e.g., showers, lockers, physical activity programs). A worksite physical activity promotion index was a tally of strategies. Total sedentary and moderate-to-vigorous physical activity (MVPA) min/d were objectively assessed via 7-day accelerometry. Total job-related physical activity minutes and recreational physical activity minutes were self-reported with the International Physical Activity Questionnaire. Mixed-effects models and generalized estimating equations evaluated the association of the worksite promotion index with physical activity and sedentary behavior, adjusting for demographics. A higher worksite promotion index was significantly associated with higher total sedentary behavior (β = 3.97), MVPA (β = 1.04), recreational physical activity (β = 1.1 and odds ratio = 1.39; away from work and at work, respectively) and negatively with job-related physical activity (β = .90). Multiple worksite physical activity promotion strategies based on environmental supports and policies may increase recreational physical activity and should be evaluated in controlled trials. These findings are particularly important given the increasingly sedentary nature of employment.

Forum: Communication Activism Pedagogy. Turning Communication Activism Pedagogy Teaching into Communication Activism Pedagogy Research

ERIC Educational Resources Information Center

Frey, Lawrence R.; Palmer, David L.

2017-01-01

In this rejoinder to this forum's respondents to the stimulus essay, "Communication Activism Pedagogy and Research: Communication Education Scholarship to Promote Social Justice," Lawrence Frey and David Palmer state that the forum editors asked them and the invited respondents to focus on communication activism pedagogy (CAP) research…

Leisure-time physical activity in relation to occupational physical activity among women.

PubMed

Ekenga, Christine C; Parks, Christine G; Wilson, Lauren E; Sandler, Dale P

2015-05-01

The objective of this study is to examine the association between occupational physical activity and leisure-time physical activity among US women in the Sister Study. We conducted a cross-sectional study of 26,334 women who had been employed in their current job for at least 1 year at baseline (2004-2009). Occupational physical activity was self-reported and leisure-time physical activity was estimated in metabolic equivalent hours per week. Log multinomial regression was used to evaluate associations between occupational (sitting, standing, manually active) and leisure-time (insufficient, moderate, high) activity. Models were adjusted for age, race/ethnicity, education, income, geographic region, and body mass index. Only 54% of women met or exceeded minimum recommended levels of leisure-time physical activity (moderate 32% and high 22%). Women who reported sitting (prevalence ratio (PR)=0.82, 95% confidence interval (CI): 0.74-0.92) or standing (PR=0.84, 95% CI: 0.75-0.94) most of the time at work were less likely to meet the requirements for high leisure-time physical activity than manually active workers. Associations were strongest among women living in the Northeast and the South. In this nationwide study, low occupational activity was associated with lower leisure-time physical activity. Women who are not active in the workplace may benefit from strategies to promote leisure-time physical activity. Published by Elsevier Inc.

Leisure-time physical activity in relation to occupational physical activity among women

PubMed Central

Ekenga, Christine C.; Parks, Christine G.; Wilson, Lauren E.; Sandler, Dale P.

2017-01-01

Objective To examine the association between occupational physical activity and leisure-time physical activity among US women in the Sister Study. Methods We conducted a cross-sectional study of 26,334 women who had been employed in their current job for at least 1 year at baseline (2004–2009). Occupational physical activity was self-reported and leisure-time physical activity was estimated in metabolic equivalent hours per week. Log multinomial regression was used to evaluate associations between occupational (sitting, standing, manually active) and leisure-time (insufficient, moderate, high) activity. Models were adjusted for age, race/ethnicity, education, income, geographic region, and body mass index. Results Only 54% of women met or exceeded minimum recommended levels of leisure-time physical activity (moderate 32% and high 22%). Women who reported sitting (PR = 0.82, 95% CI: 0.74–0.92) or standing (PR = 0.84, 95% CI: 0.75–0.94) most of the time at work were less likely to meet the requirements for high leisure-time physical activity than manually active workers. Associations were strongest among women living in the Northeast and the South. Conclusion In this nationwide study, low occupational activity was associated with lower leisure-time physical activity. Women who are not active in the workplace may benefit from strategies to promote leisure-time physical activity. PMID:25773471

Emodin Regulates Glucose Utilization by Activating AMP-activated Protein Kinase*

PubMed Central

Song, Parkyong; Kim, Jong Hyun; Ghim, Jaewang; Yoon, Jong Hyuk; Lee, Areum; Kwon, Yonghoon; Hyun, Hyunjung; Moon, Hyo-Youl; Choi, Hueng-Sik; Berggren, Per-Olof; Suh, Pann-Ghill; Ryu, Sung Ho

2013-01-01

AMP-activated protein kinase has been described as a key signaling protein that can regulate energy homeostasis. Here, we aimed to characterize novel AMP-activated kinase (AMPK)-activating compounds that have a much lower effective concentration than metformin. As a result, emodin, a natural anthraquinone derivative, was shown to stimulate AMPK activity in skeletal muscle and liver cells. Emodin enhanced GLUT4 translocation and [14C]glucose uptake into the myotube in an AMPK-dependent manner. Also, emodin inhibited glucose production by suppressing the expression of key gluconeogenic genes, such as phosphoenolpyruvate carboxykinase and glucose-6-phosphatase, in hepatocytes. Furthermore, we found that emodin can activate AMPK by inhibiting mitochondrial respiratory complex I activity, leading to increased reactive oxygen species and Ca2+/calmodulin-dependent protein kinase kinase activity. Finally, we confirmed that a single dose administration of emodin significantly decreased the fasting plasma glucose levels and improved glucose tolerance in C57Bl/6J mice. Increased insulin sensitivity was also confirmed after daily injection of emodin for 8 days using an insulin tolerance test and insulin-stimulated PI3K phosphorylation in wild type and high fat diet-induced diabetic mouse models. Our study suggests that emodin regulates glucose homeostasis in vivo by AMPK activation and that this may represent a novel therapeutic principle in the treatment of type 2 diabetic models. PMID:23303186

Lactate dehydrogenase activity drives hair follicle stem cell activation

PubMed Central

Aimee, Flores; John, Schell; Abby, Krall; David, Jelinek; Matilde, Miranda; Melina, Grigorian; Daniel, Braas; White Andrew, C; Jessica, Zhou; Nick, Graham; Thomas, Graeber; Pankaj, Seth; Denis, Evseenko; Hilary, Coller; Jared, Rutter; Heather, Christofk; Lowry William, E

2017-01-01

Summary While normally dormant, Hair Follicle Stem Cells (HFSCs) quickly become activated to divide during a new hair cycle. The quiescence of HFSCs is known to be regulated by a number of intrinsic and extrinsic mechanisms. Here we provide several lines of evidence to demonstrate that HFSCs utilize glycolytic metabolism and produce significantly more lactate than other cells in the epidermis. Furthermore, lactate generation appears to be critical for the activation of HFSCs as deletion of lactate dehydrogenase (Ldha) prevented their activation. Conversely, genetically promoting lactate production in HFSCs through mitochondrial pyruvate carrier (Mpc1) deletion accelerated their activation and the hair cycle. Finally, we identify small molecules that increase lactate production by stimulating Myc levels or inhibiting Mpc1 carrier activity and can topically induce the hair cycle. These data suggest that HFSCs maintain a metabolic state that allow them to remain dormant and yet quickly respond to appropriate proliferative stimuli. PMID:28812580

Physical Activity Basics

MedlinePlus

... Weight Breastfeeding Micronutrient Malnutrition State and Local Programs Physical Activity Basics Recommend on Facebook Tweet Share Compartir How much physical activity do you need? Regular physical activity helps improve ...

Riluzole activates TRPC5 channels independently of PLC activity

PubMed Central

Richter, Julia M; Schaefer, Michael; Hill, Kerstin

2014-01-01

BACKGROUND AND PURPOSE The transient receptor potential channel C5 (TRPC5) is a Ca2+-permeable cation channel, which is predominantly expressed in the brain. TRPC5 is activated in a PLC-dependent manner by, as yet, unidentified endogenous messengers. Recently, modulators of TRPC5, like Ca2+, pH and phospholipids, have been identified. However, the role of TRPC5 in vivo is only poorly understood. Novel specific modulators of TRPC5 might help to elucidate its function. EXPERIMENTAL APPROACH Novel modulators of TRPC5 were identified in a compound screening of approved drugs and natural compounds. The potency and selectivity of TRPC5-activating compounds were determined by fluorometric calcium imaging. The biophysical properties of channel activation by these compounds were analysed using electrophysiological measurements. KEY RESULTS Riluzole was identified as a novel activator of TRPC5 (EC50 9.2 ± 0.5 μM) and its mechanism of action was shown to be independent of G protein signalling and PLC activity. Riluzole-induced TRPC5 currents were potentiated by La3+ and, utilizing TRPC5 mutants that lack La3+ binding sites, it was confirmed that riluzole and La3+ activate TRPC5 by different mechanisms. Recordings of excised inside-out patches revealed a relatively direct effect of riluzole on TRPC5. CONCLUSIONS AND IMPLICATIONS Riluzole can activate TRPC5 heterologously expressed in HEK293 cells as well as those endogenously expressed in the U-87 glioblastoma cell line. Riluzole does not activate any other member of the TRPC family and could, therefore, despite its action on other ion channels, be a useful pharmacological tool for identifying TRPC5-specific currents in immortalized cell lines or in acutely isolated primary cells. PMID:24117252

Requirement of GM2 ganglioside activator for phospholipase D activation

PubMed Central

Nakamura, Shun-ichi; Akisue, Toshihiro; Jinnai, Hitoshi; Hitomi, Tomohiro; Sarkar, Sukumar; Miwa, Noriko; Okada, Taro; Yoshida, Kimihisa; Kuroda, Shun’ichi; Kikkawa, Ushio; Nishizuka, Yasutomi

1998-01-01

Sequence analysis of a heat-stable protein necessary for the activation of ADP ribosylation factor-dependent phospholipase D (PLD) reveals that this protein has a structure highly homologous to the previously known GM2 ganglioside activator whose deficiency results in the AB-variant of GM2 gangliosidosis. The heat-stable activator protein indeed has the capacity to enhance enzymatic conversion of GM2 to GM3 ganglioside that is catalyzed by β-hexosaminidase A. Inversely, GM2 ganglioside activator purified separately from tissues as described earlier [Conzelmann, E. & Sandhoff, K. (1987) Methods Enzymol. 138, 792–815] stimulates ADP ribosylation factor-dependent PLD in a dose-dependent manner. At higher concentrations of ammonium sulfate, the PLD activator protein apparently substitutes for protein kinase C and phosphatidylinositol 4,5-bisphosphate, both of which are known as effective stimulators of the PLD reaction. The mechanism of action of the heat-stable PLD activator protein remains unknown. PMID:9770472

Activity Based Curriculum for Elementary Education. Additional Activities, K-6.

ERIC Educational Resources Information Center

Wichita Public Schools, KS.

This elementary curriculum is a vehicle to provide manipulative activities that reinforce academic skills through meaningful, relevant, activity-based awareness of modern society. The twenty-six activity plans included in the curriculum place a major emphasis upon realistic or concrete experiences that deal with the manipulation and exploration of…

Demo of Active Resonators on the Active Noise Control Fan

NASA Image and Video Library

1998-10-01

NASA has been involved in several projects to reduce aircraft engine noise. In the 1990s Active Noise Control was investigated as a potential method to reduce aircraft engine noise. A description and demonstration of Active Resonators on the Active Noise Control Fan is shown in this video.

Examining Activism in Practice: A Qualitative Study of Archival Activism

ERIC Educational Resources Information Center

Novak, Joy Rainbow

2013-01-01

While archival literature has increasingly discussed activism in the context of archives, there has been little examination of the extent to which archivists in the field have accepted or incorporated archival activism into practice. Scholarship that has explored the practical application of archival activism has predominately focused on case…

Active8! Technology-Based Intervention to Promote Physical Activity in Hospital Employees.

PubMed

Blake, Holly; Suggs, L Suzanne; Coman, Emil; Aguirre, Lucia; Batt, Mark E

2017-03-01

Increase physical activity in health care employees using health messaging, and compare e-mail with mobile phone short-message service (SMS) as delivery channels. Randomized controlled trial Setting. U.K. hospital workplace. Two hundred ninety-six employees (19-67 years, 53% of study Web site visitors). Twelve-week messaging intervention designed to increase physical activity and delivered via SMS (n =147) or e-mail (n =149); content tailored using theory of planned behavior (TPB) and limited to 160 characters. Baseline and 6, 12, and 16 weeks. Online measures included TPB constructs, physical activity behavior on the Global Physical Activity Questionnaire, and health-related quality of life on the Short-Form 12. General linear models for repeated measures. Increase in duration (mean h/d) of moderate work-related activity and moderate recreational activity from baseline to 16 weeks. Short-lived increase in frequency (d/wk) of vigorous recreational activity from baseline to 6 weeks. Increase in duration and frequency of active travel from baseline to 16 weeks. E-mails generated greater changes than SMS in active travel and moderate activity (work and recreational). Minimal physical activity promotion delivered by SMS or e-mail can increase frequency and duration of active travel and duration of moderate intensity physical activity at work and for leisure, which is maintained up to 1 month after messaging ends. Both channels were useful platforms for health communication; e-mails were particularly beneficial with hospital employees.

Activated carbon from peach stones using phosphoric acid activation at medium temperatures.

PubMed

Kim, Dong-Su

2004-01-01

In the present study, the activation features of phosphoric acid have been investigated using waste peach stones as the raw material in the production of granular activated carbon. Thermogravimetry/differential thermal analysis was conducted to characterize the thermal behavior of peach stone and titration method was used to evaluate the adsorption capacity of the produced activated carbon. It was observed that the iodine value of the activated carbon increased with activation temperature. However, temperatures higher than 500 degrees C caused a thermal destruction, which resulted in the decrease of the adsorption capacity. Activation longer than 1.5 h at 500 degrees C resulted in thermal degradation of the porous structure of the activated carbon. The adsorption capacity was enhanced with increasing of amounts of phosphoric acid, however, excessive phosphoric acid caused a decrease in the iodine value. In addition, it was found that the carbon yields generally decreased with activation temperature and activation time. Scanning electron microscopy analysis was conducted to observe the changes in the poros structure of the activated carbon produced in different temperatures. Activation of carbon by phosphoric acid was found to be superior to that by CaCl2 and gas activation. The activated carbon produced from peach stone was applied as an adsorbent in the treatment of synthesized wastewater containing cadmium ion and its adsorption capacity was found to be as good as that of the commercial one.

Simultaneous Fluorescent Gram Staining and Activity Assessment of Activated Sludge Bacteria

PubMed Central

Forster, Scott; Snape, Jason R.; Lappin-Scott, Hilary M.; Porter, Jonathan

2002-01-01

Wastewater treatment is one of the most important commercial biotechnological processes, and yet the component bacterial populations and their associated metabolic activities are poorly understood. The novel fluorescent dye hexidium iodide allows assessment of Gram status by differential absorption through bacterial cell walls. Differentiation between gram-positive and gram-negative wastewater bacteria was achieved after flow cytometric analysis. This study shows that the relative proportions of gram-positive and gram-negative bacterial cells identified by traditional microscopy and hexidium iodide staining were not significantly different. Dual staining of cells for Gram status and activity proved effective in analyzing mixtures of cultured bacteria and wastewater populations. Levels of highly active organisms at two wastewater treatment plants, both gram positive and gram negative, ranged from 1.5% in activated sludge flocs to 16% in the activated sludge fluid. Gram-positive organisms comprised <5% of the total bacterial numbers but accounted for 19 and 55% of the highly active organisms within flocs at the two plants. Assessment of Gram status and activity within activated sludge samples over a 4-day period showed significant differences over time. This method provides a rapid, quantitative measure of Gram status linked with in situ activity within wastewater systems. PMID:12324319

Simultaneous fluorescent gram staining and activity assessment of activated sludge bacteria.

PubMed

Forster, Scott; Snape, Jason R; Lappin-Scott, Hilary M; Porter, Jonathan

2002-10-01

Wastewater treatment is one of the most important commercial biotechnological processes, and yet the component bacterial populations and their associated metabolic activities are poorly understood. The novel fluorescent dye hexidium iodide allows assessment of Gram status by differential absorption through bacterial cell walls. Differentiation between gram-positive and gram-negative wastewater bacteria was achieved after flow cytometric analysis. This study shows that the relative proportions of gram-positive and gram-negative bacterial cells identified by traditional microscopy and hexidium iodide staining were not significantly different. Dual staining of cells for Gram status and activity proved effective in analyzing mixtures of cultured bacteria and wastewater populations. Levels of highly active organisms at two wastewater treatment plants, both gram positive and gram negative, ranged from 1.5% in activated sludge flocs to 16% in the activated sludge fluid. Gram-positive organisms comprised <5% of the total bacterial numbers but accounted for 19 and 55% of the highly active organisms within flocs at the two plants. Assessment of Gram status and activity within activated sludge samples over a 4-day period showed significant differences over time. This method provides a rapid, quantitative measure of Gram status linked with in situ activity within wastewater systems.

Pharmacological Activation of Peroxisome Proliferator-Activated Receptor {Delta} Increases Sphingomyelin Synthase Activity in THP-1 Macrophage-Derived Foam Cell.

PubMed

Mou, Dongsheng; Yang, Hua; Qu, Changhua; Chen, Juan; Zhang, Chaogui

2016-08-01

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors, which mediate glucose and lipid homeostasis by regulating the expression of a large number of transcription factors. Sphingomyelin synthase (SMS) is a key enzyme in the synthesis of sphingomyelin (SM), and its expression and activity have been reported to be associated with atherosclerosis (AS). Although there have been many functional PPAR and SMS studies on atherosclerosis in recent years, few have investigated the correlation between the activation of PPARδ and the activity of SMS. In his study, macrophage-induced foam cells were utilized to model important pathological changes that occur in AS. The influence of PPARδ agonism by GW501516 on SMS and its product molecule SM were measured. Results indicated that the activation of PPARδ was correlated in a positive manner with the activity of SMS2, and the content of SM was dose dependently increased by GW501516. Together, this study represents the first to suggest that PPARδ activation may be a potential risk of AS through enhancing activity of SMS2.

Testicular cell-conditioned medium supports embryonic stem cell differentiation toward germ lineage and to spermatocyte- and oocyte-like cells.

PubMed

Shah, Syed M; Saini, Neha; Singh, Manoj K; Manik, Radheysham; Singla, Suresh K; Palta, Prabhat; Chauhan, Manmohan S

2016-08-01

Testicular cells are believed to secrete various growth factors that activate signaling pathways finally leading to gametogenesis. In vitro gametogenesis is an obscure but paramountly important task primarily because of paucity of the precursor cells and first trimester gonadal tissues. To overcome these limitations for development of in vitro gametes, the present study was designed to induce differentiation of buffalo embryonic stem (ES) cells into germ lineage cells on stimulation by testicular cell-conditioned medium (TCM), on the basis of the assumption that ES cells have the intrinsic property to differentiate into any cell type and TCM would provide the necessary growth factors for differentiation toward germ cell lineage. For this purpose, buffalo ES cells were differentiated as embryoid bodies (EB) in floating cultures and as monolayer adherent cultures in different doses (10%, 20%, and 40%) of TCM for different culture intervals (4, 8, and 14 days), to identify the optimum dose-and-time period. We observed that 40% TCM dose induces highest expression of primordial germ cell-specific (DAZL, VASA, and PLZF), meiotic (SYCP3, MLH1, TNP1/2, and PRM2), spermatocyte-specific (BOULE and TEKT1), and oocyte-specific genes (GDF9 and ZP2/3) for a culture period of 14 days under both floating and adherent differentiation. Immunocytochemical analysis of EBs and adherent cultures revealed presence of primordial germ cell markers (c-KIT, DAZL, and VASA), meiotic markers (SYCP3, MLH1 and PROTAMINE1), spermatocyte markers (ACROSIN and HAPRIN), and oocyte markers (GDF9 and ZP4), indicating progression into post-meiotic gametogenesis. The detection of germ cell-specific proteins in Day 14 EBs like VASA, GDF9, and ZP4 by Western blotting further confirmed germ lineage differentiation. The significantly lower (P < 0.05) concentration of 5-methyl-2-deoxycytidine in optimally differentiated EBs is suggestive of the process of methylation erasure. Oocyte-like structures

Rethinking food anticipatory activity in the activity-based anorexia rat model.

PubMed

Wu, Hemmings; van Kuyck, Kris; Tambuyzer, Tim; Luyten, Laura; Aerts, Jean-Marie; Nuttin, Bart

2014-01-29

When a rat is on a limited fixed-time food schedule with full access to a running wheel (activity-based anorexia model, ABA), its activity level will increase hours prior to the feeding period. This activity, called food-anticipatory activity (FAA), is a hypothesized parallel to the hyperactivity symptom in human anorexia nervosa. To investigate in depth the characteristics of FAA, we retrospectively analyzed the level of FAA and activities during other periods in ABA rats. To our surprise, rats with the most body weight loss have the lowest level of FAA, which contradicts the previously established link between FAA and the severity of ABA symptoms. On the contrary, our study shows that postprandial activities are more directly related to weight loss. We conclude that FAA alone may not be sufficient to reflect model severity, and activities during other periods may be of potential value in studies using ABA model.

The effect of physical activity homework on physical activity among college students.

PubMed

Claxton, David; Wells, Gayle M

2009-03-01

This study examined the effect of using physical activity homework on physical activity levels of college students. Students in randomly assigned sections of a university health course were assigned 30 minutes of physical activity homework 3 days a week or no homework for 12 weeks. Participants completed self-reports of physical activity before the homework intervention and again at the conclusion of the 12 weeks of physical activity homework. Participants in all course sections reported significant increases in the number of days per week of moderate and vigorous physical activity. Participants in homework sections additionally showed significant increases in the days they engaged in muscular strength/endurance training and activities to manage weight. Participants in sections without homework showed a significant increase in the number of days engaged in flexibility training. Comparison of gain scores showed statistically significant increases by the homework group in the days they participated in activities designed to manage weight. Physical activity homework was deemed to be an effective method of increasing college students' levels of physical activity.

Don't worry, be active: positive affect and habitual physical activity.

PubMed

Pasco, Julie A; Jacka, Felice N; Williams, Lana J; Brennan, Sharon L; Leslie, Eva; Berk, Michael

2011-12-01

The aim of ths study was to examine the association between habitual physical activity and positive and negative affect. This cross-sectional study included 276 women aged 20 +, from the Geelong Osteoporosis Study. Habitual physical activity and other lifestyle exposures were assessed by questionnaire, concurrent with anthropometric assessments. Physical activity was categorized as very active, moderately active or sedentary. Positive and negative affect scores were derived from the validated 20 item Positive and Negative Affect Schedule (PANAS) self-report and were categorized into tertiles. There was a pattern of lower positive affect scores for lower levels of physical activity. With very active as the reference category, the odds for having a positive affect score in the highest tertile were sequentially lower for those who were moderately active (OR = 0.53, 95%CI 0.28-1.01) and sedentary (OR = 0.28, 95%CI 0.10-0.75). Associations were sustained after adjusting for body mass index and polypharmacy (OR = 0.50, 95%CI 0.26-0.96 and OR = 0.25, 95%CI 0.09-0.72, respectively). These associations were not explained by age, negative affect score or other exposures. No association was detected between physical activity and negative affect scores. This study reports that higher positive affect scores, encompassing emotions such as interest, excitement, enthusiasm and alertness, are associated with higher levels of habitual physical activity. These observations warrant further investigations into possible mechanistic interplay between neurobiological and psychosocial factors that underpin this association.

Youth Physical Activity Resource Use and Activity Measured by Accelerometry

ERIC Educational Resources Information Center

Maslow, Andra L.; Colabianchi, Natalie

2011-01-01

Objectives: To examine whether use of physical activity resources (e.g., parks) was associated with daily physical activity measured by accelerometry. Methods: One hundred eleven adolescents completed a travel diary with concurrent accelerometry. The main exposure was self-reported use of a physical activity resource (none /1 resources). The main…

Predicting Activity Energy Expenditure Using the Actical[R] Activity Monitor

ERIC Educational Resources Information Center

Heil, Daniel P.

2006-01-01

This study developed algorithms for predicting activity energy expenditure (AEE) in children (n = 24) and adults (n = 24) from the Actical[R] activity monitor. Each participant performed 10 activities (supine resting, three sitting, three house cleaning, and three locomotion) while wearing monitors on the ankle, hip, and wrist; AEE was computed…

Structure-activity analysis of synthetic alpha-thrombin-receptor-activating peptides.

PubMed

Van Obberghen-Schilling, E; Rasmussen, U B; Vouret-Craviari, V; Lentes, K U; Pavirani, A; Pouysségur, J

1993-06-15

alpha-Thrombin stimulates G-protein-coupled effectors leading to secretion and aggregation in human platelets, and to a mitogenic response in CCL39 hamster fibroblasts. alpha-Thrombin receptors can be activated by synthetic peptides corresponding to the receptor sequence starting with serine-42, at the proposed cleavage site. We have previously determined that the agonist domain of receptor-activating peptides resides within the five N-terminal residues [Vouret-Craviari, Van Obberghen-Schilling, Rasmussen, Pavirani, Lecocq and Pouysségur (1992) Mol. Biol. Cell. 3, 95-102], although the 7-residue peptide (SFFLRNP) corresponding to the hamster alpha-thrombin receptor was 10 times more potent than the 5-residue peptide for activation of human platelets. In the present study we have analysed the role of individual amino acids in receptor activation by using a series of modified hexa- or hepta-peptides derived from the human alpha-thrombin-receptor sequence. Cellular events examined here include phospholipase C activation, adenylyl cyclase inhibition and DNA synthesis stimulation in non-transformed CCL39 fibroblasts and a tumorigenic variant of that line (A71 cells). Modification of the peptide sequence had similar functional consequence for each of the assays described, indicating that either a unique receptor or pharmacologically indistinguishable receptor subtypes activate distinct G-protein signalling pathways. Furthermore, we found that: (1) the N-terminal serine can be replaced by small or intermediately sized amino acids (+/- hydroxyl groups) without loss of activity. However, its replacement by an aromatic side-chain or omission of the N-terminal amino group severely reduces activity. (2) An aromatic side-chain on the penultimate N-terminal residue appears to play a critical role since phenylalanine in this position can be substituted by tyrosine without complete loss of activity whereas an alanine in its place is not tolerated. (3) Deletion of the first

Structure-activity analysis of synthetic alpha-thrombin-receptor-activating peptides.

PubMed Central

Van Obberghen-Schilling, E; Rasmussen, U B; Vouret-Craviari, V; Lentes, K U; Pavirani, A; Pouysségur, J

1993-01-01

alpha-Thrombin stimulates G-protein-coupled effectors leading to secretion and aggregation in human platelets, and to a mitogenic response in CCL39 hamster fibroblasts. alpha-Thrombin receptors can be activated by synthetic peptides corresponding to the receptor sequence starting with serine-42, at the proposed cleavage site. We have previously determined that the agonist domain of receptor-activating peptides resides within the five N-terminal residues [Vouret-Craviari, Van Obberghen-Schilling, Rasmussen, Pavirani, Lecocq and Pouysségur (1992) Mol. Biol. Cell. 3, 95-102], although the 7-residue peptide (SFFLRNP) corresponding to the hamster alpha-thrombin receptor was 10 times more potent than the 5-residue peptide for activation of human platelets. In the present study we have analysed the role of individual amino acids in receptor activation by using a series of modified hexa- or hepta-peptides derived from the human alpha-thrombin-receptor sequence. Cellular events examined here include phospholipase C activation, adenylyl cyclase inhibition and DNA synthesis stimulation in non-transformed CCL39 fibroblasts and a tumorigenic variant of that line (A71 cells). Modification of the peptide sequence had similar functional consequence for each of the assays described, indicating that either a unique receptor or pharmacologically indistinguishable receptor subtypes activate distinct G-protein signalling pathways. Furthermore, we found that: (1) the N-terminal serine can be replaced by small or intermediately sized amino acids (+/- hydroxyl groups) without loss of activity. However, its replacement by an aromatic side-chain or omission of the N-terminal amino group severely reduces activity. (2) An aromatic side-chain on the penultimate N-terminal residue appears to play a critical role since phenylalanine in this position can be substituted by tyrosine without complete loss of activity whereas an alanine in its place is not tolerated. (3) Deletion of the first

Transcription co-activator SAYP mediates the action of STAT activator

PubMed Central

Panov, Vladislav V.; Kuzmina, Julia L.; Doronin, Semen A.; Kopantseva, Marina R.; Nabirochkina, Elena N.; Georgieva, Sofia G.; Vorobyeva, Nadezhda E.; Shidlovskii, Yulii V.

2012-01-01

Jak/STAT is an important signaling pathway mediating multiple events in development. We describe participation of metazoan co-activator SAYP/PHF10 in this pathway downstream of STAT. The latter, via its activation domain, interacts with the conserved core of SAYP. STAT is associated with the SAYP-containing co-activator complex BTFly and recruits BTFly onto genes. SAYP is necessary for stimulating STAT-driven transcription of numerous genes. Mutation of SAYP leads to maldevelopments similar to those observed in STAT mutants. Thus, SAYP is a novel co-activator mediating the action of STAT. PMID:22123744

Transcription co-activator SAYP mediates the action of STAT activator.

PubMed

Panov, Vladislav V; Kuzmina, Julia L; Doronin, Semen A; Kopantseva, Marina R; Nabirochkina, Elena N; Georgieva, Sofia G; Vorobyeva, Nadezhda E; Shidlovskii, Yulii V

2012-03-01

Jak/STAT is an important signaling pathway mediating multiple events in development. We describe participation of metazoan co-activator SAYP/PHF10 in this pathway downstream of STAT. The latter, via its activation domain, interacts with the conserved core of SAYP. STAT is associated with the SAYP-containing co-activator complex BTFly and recruits BTFly onto genes. SAYP is necessary for stimulating STAT-driven transcription of numerous genes. Mutation of SAYP leads to maldevelopments similar to those observed in STAT mutants. Thus, SAYP is a novel co-activator mediating the action of STAT.

Physical Activity Assessment

Cancer.gov

Current evidence convincingly indicates that physical activity reduces the risk of colon and breast cancer. Physical activity may also reduce risk of prostate cancer. Scientists are also evaluating potential relationships between physical activity and other cancers.

Integration of active pauses and pattern of muscular activity during computer work.

PubMed

St-Onge, Nancy; Samani, Afshin; Madeleine, Pascal

2017-09-01

Submaximal isometric muscle contractions have been reported to increase variability of muscle activation during computer work; however, other types of active contractions may be more beneficial. Our objective was to determine which type of active pause vs. rest is more efficient in changing muscle activity pattern during a computer task. Asymptomatic regular computer users performed a standardised 20-min computer task four times, integrating a different type of pause: sub-maximal isometric contraction, dynamic contraction, postural exercise and rest. Surface electromyographic (SEMG) activity was recorded bilaterally from five neck/shoulder muscles. Root-mean-square decreased with isometric pauses in the cervical paraspinals, upper trapezius and middle trapezius, whereas it increased with rest. Variability in the pattern of muscular activity was not affected by any type of pause. Overall, no detrimental effects on the level of SEMG during active pauses were found suggesting that they could be implemented without a cost on activation level or variability. Practitioner Summary: We aimed to determine which type of active pause vs. rest is best in changing muscle activity pattern during a computer task. Asymptomatic computer users performed a standardised computer task integrating different types of pauses. Muscle activation decreased with isometric pauses in neck/shoulder muscles, suggesting their implementation during computer work.

syk kinase activation by a src kinase-initiated activation loop phosphorylation chain reaction

PubMed Central

El-Hillal, O.; Kurosaki, T.; Yamamura, H.; Kinet, J.-P.; Scharenberg, A. M.

1997-01-01

Activation of the syk tyrosine kinase occurs almost immediately following engagement of many types of antigen receptors, including Fc receptors, but the mechanism through which syk is activated is currently unclear. Here we demonstrate that Fc receptor-induced syk activation occurs as the result of phosphorylation of the syk activation loop by both src family kinases and other molecules of activated syk, suggesting that syk activation occurs as the result of a src kinase-initiated activation loop phosphorylation chain reaction. This type of activation mechanism predicts that syk activation would exhibit exponential kinetics, providing a potential explanation for its rapid and robust activation by even weak antigen receptor stimuli. We propose that a similar mechanism may be responsible for generating rapid activation of other cytoplasmic tyrosine kinases, such as those of the Bruton tyrosine kinase/tec family, as well. PMID:9050880

Activity of antimicrobial peptide mimetics in the oral cavity: II. Activity against periopathogenic biofilms and anti-inflammatory activity

PubMed Central

Hua, J; Scott, R.W.; Diamond, G

2011-01-01

Whereas periodontal disease is ultimately of bacterial etiology, from multispecies biofilms of gram-negative anaerobic microorganisms, much of the deleterious effects are caused by the resultant epithelial inflammatory response. Hence, development of a treatment that combines anti-biofilm antibiotic activity with anti-inflammatory activity would be of great utility. Antimicrobial peptides (AMPs) such as defensins are naturally occurring peptides that exhibit broad-spectrum activity as well as a variety of immunomodulatory activities. Furthermore, bacteria do not readily develop resistance to these agents. However, clinical studies have suggested that they do not represent optimal candidates for exogenous therapeutic agents. Small-molecule mimetics of these AMPs exhibit similar activities to the parent peptides, in addition to having low toxicity, high stability and low cost. To determine whether AMP mimetics have the potential for treatment of periodontal disease, we examined the activity of one mimetic, mPE, against biofilm cultures of Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis. Metabolic assays as well as culture and biomass measurement assays demonstrated that mPE exhibits potent activity against biofilm cultures of both species. Furthermore, as little as 2 µg ml−1 mPE was sufficient to inhibit interleukin-1β-induced secretion of interleukin-8 in both gingival epithelial cells and THP-1 cells. This anti-inflammatory activity is associated with a reduction in activation of nuclear factor-κB, suggesting that mPE can act both as an anti-biofilm agent in an anaerobic environment and as an anti-inflammatory agent in infected tissues. PMID:21040516

Recess, Extracurricular Activities, and Active Classrooms: Means for Increasing Elementary School Students' Physical Activity

ERIC Educational Resources Information Center

Kahan, David

2008-01-01

Physical education is traditionally thought of as the primary means of providing physical activity in the school environment. However, only 17 to 22 percent of elementary schools offer daily physical education with a cumulative duration of about 85 to 98 minutes per week. Based on pedometer counts of weekday physical activity, lunch recess and…

Glycogen synthase kinase 3β promotes liver innate immune activation by restraining AMP-activated protein kinase activation.

PubMed

Zhou, Haoming; Wang, Han; Ni, Ming; Yue, Shi; Xia, Yongxiang; Busuttil, Ronald W; Kupiec-Weglinski, Jerzy W; Lu, Ling; Wang, Xuehao; Zhai, Yuan

2018-07-01

Glycogen synthase kinase 3β (Gsk3β [Gsk3b]) is a ubiquitously expressed kinase with distinctive functions in different types of cells. Although its roles in regulating innate immune activation and ischaemia and reperfusion injuries (IRIs) have been well documented, the underlying mechanisms remain ambiguous, in part because of the lack of cell-specific tools in vivo. We created a myeloid-specific Gsk3b knockout (KO) strain to study the function of Gsk3β in macrophages in a murine liver partial warm ischaemia model. Compared with controls, myeloid Gsk3b KO mice were protected from IRI, with diminished proinflammatory but enhanced anti-inflammatory immune responses in livers. In bone marrow-derived macrophages, Gsk3β deficiency resulted in an early reduction of Tnf gene transcription but sustained increase of Il10 gene transcription on Toll-like receptor 4 stimulation in vitro. These effects were associated with enhanced AMP-activated protein kinase (AMPK) activation, which led to an accelerated and higher level of induction of the novel innate immune negative regulator small heterodimer partner (SHP [Nr0b2]). The regulatory function of Gsk3β on AMPK activation and SHP induction was confirmed in wild-type bone marrow-derived macrophages with a Gsk3 inhibitor. Furthermore, we found that this immune regulatory mechanism was independent of Gsk3β Ser9 phosphorylation and the phosphoinositide 3-kinase-Akt signalling pathway. In vivo, myeloid Gsk3β deficiency facilitated SHP upregulation by ischaemia-reperfusion in liver macrophages. Treatment of Gsk3b KO mice with either AMPK inhibitor or SHP small interfering RNA before the onset of liver ischaemia restored liver proinflammatory immune activation and IRI in these otherwise protected hosts. Additionally, pharmacological activation of AMPK protected wild-type mice from liver IRI, with reduced proinflammatory immune activation. Inhibition of the AMPK-SHP pathway by liver ischaemia was demonstrated in tumour resection

Antioxidative activities and active compounds of extracts from Catalpa plant leaves.

PubMed

Xu, Hongyu; Hu, Gege; Dong, Juane; Wei, Qin; Shao, Hongbo; Lei, Ming

2014-01-01

In order to screen the Catalpa plant with high antioxidant activity and confirm the corresponding active fractions from Catalpa ovata G. Don, C. fargesii Bur., and C. bungei C. A. Mey., total flavonoid contents and antioxidant activities of the extracts/fractions of Catalpa plant leaves were determined. The determined total flavonoid content and antioxidant activity were used as assessment criteria. Those compounds with antioxidant activity were isolated with silica gel column chromatography and ODS column chromatography. Our results showed that the total flavonoid content in C. bungei C. A. Mey. (30.07 mg/g · DW) was the highest, followed by those in C. fargesii Bur. (25.55 mg/g · DW) and C. ovata G. Don (24.96 mg/g · DW). According to the determination results of total flavonoid content and antioxidant activity in 3 clones of leaves of C. bungei C. A. Mey., the total flavonoid content and antioxidant activity in crude extracts from C. bungei C. A. Mey. 6 (CA6) leaves were the highest. Moreover, the results showed that the total flavonoid content and antioxidant activities of ethyl acetate (EA) fraction in ethanol crude extracts in CA6 leaves were the highest, followed by n-butanol, petroleum ether (PE), and water fractions. Two flavonoid compounds with antioxidant activity were firstly isolated based on EA fraction. The two compounds were luteolin (1) and apigenin (2), respectively.

Activated AMPK inhibits PPAR-{alpha} and PPAR-{gamma} transcriptional activity in hepatoma cells.

PubMed

Sozio, Margaret S; Lu, Changyue; Zeng, Yan; Liangpunsakul, Suthat; Crabb, David W

2011-10-01

AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor-α (PPAR-α) are critical regulators of short-term and long-term fatty acid oxidation, respectively. We examined whether the activities of these molecules were coordinately regulated. H4IIEC3 cells were transfected with PPAR-α and PPAR-γ expression plasmids and a peroxisome-proliferator-response element (PPRE) luciferase reporter plasmid. The cells were treated with PPAR agonists (WY-14,643 and rosiglitazone), AMPK activators 5-aminoimidazole-4-carboxamide riboside (AICAR) and metformin, and the AMPK inhibitor compound C. Both AICAR and metformin decreased basal and WY-14,643-stimulated PPAR-α activity; compound C increased agonist-stimulated reporter activity and partially reversed the effect of the AMPK activators. Similar effects on PPAR-γ were seen, with both AICAR and metformin inhibiting PPRE reporter activity. Compound C increased basal PPAR-γ activity and rosiglitazone-stimulated activity. In contrast, retinoic acid receptor-α (RAR-α), another nuclear receptor that dimerizes with retinoid X receptor (RXR), was largely unaffected by the AMPK activators. Compound C modestly increased AM580 (an RAR agonist)-stimulated activity. The AMPK activators did not affect PPAR-α binding to DNA, and there was no consistent correlation between effects of the AMPK activators and inhibitor on PPAR and the nuclear localization of AMPK-α subunits. Expression of either a constitutively active or dominant negative AMPK-α inhibited basal and WY-14,643-stimulated PPAR-α activity and basal and rosiglitazone-stimulated PPAR-γ activity. We concluded that the AMPK activators AICAR and metformin inhibited transcriptional activities of PPAR-α and PPAR-γ, whereas inhibition of AMPK with compound C activated both PPARs. The effects of AMPK do not appear to be mediated through effects on RXR or on PPAR/RXR binding to DNA. These effects are independent of kinase activity and instead appear to

Get Active Orlando: changing the built environment to increase physical activity.

PubMed

McCreedy, Malisa; Leslie, Jill G

2009-12-01

Active Living by Design's Get Active Orlando partnership (GAO) focused on downtown Orlando's Community Redevelopment Area, including the Parramore Heritage District, home to many low-income and ethnically diverse residents, including many seniors. The area had undergone substantial development, and GAO aimed to incorporate active living considerations into the city's changing landscape. Get Active Orlando conducted a baseline survey of all streets, sidewalks, and bicycle lanes in the project area and identified a sequence of plans and policies in which to incorporate changes identified in the assessment. To create more immediate opportunities for active living, the partnership initiated a senior walking program, a bicycle refurbishment and giveaway program, and community bicycle-riding events, and led a social-marketing campaign that emphasized simple lifestyle changes. Get Active Orlando influenced adoption of public policies supporting active living in Orlando, including the Downtown Transportation Plan, Streetscape Guidelines, Design Standards Review Checklist, and growth management policies. Establishment of the Mayor's Advisory Council on Active Living is testament to the heightened significance of active living in Orlando. Initial assessment data served as a strong platform for policy change. Creating connections across disciplines including land-use planning, transportation, public health, and economic development allowed GAO to secure substantial policy change to influence design of the built environment. Engaging community members, including youth, as leaders was an important factor in program success. The physical environment in Orlando's Community Redevelopment Area is beginning to change as a reflection of a new policy framework designed to support active living.

Curcumin Suppresses T Cell Activation by Blocking Ca2+ Mobilization and Nuclear Factor of Activated T Cells (NFAT) Activation

PubMed Central

Kliem, Christian; Merling, Anette; Giaisi, Marco; Köhler, Rebecca; Krammer, Peter H.; Li-Weber, Min

2012-01-01

Curcumin is the active ingredient of the spice turmeric and has been shown to have a number of pharmacologic and therapeutic activities including antioxidant, anti-microbial, anti-inflammatory, and anti-carcinogenic properties. The anti-inflammatory effects of curcumin have primarily been attributed to its inhibitory effect on NF-κB activity due to redox regulation. In this study, we show that curcumin is an immunosuppressive phytochemical that blocks T cell-activation-induced Ca2+ mobilization with IC50 = ∼12.5 μm and thereby prevents NFAT activation and NFAT-regulated cytokine expression. This finding provides a new mechanism for curcumin-mediated anti-inflammatory and immunosuppressive function. We also show that curcumin can synergize with CsA to enhance immunosuppressive activity because of different inhibitory mechanisms. Furthermore, because Ca2+ is also the secondary messenger crucial for the TCR-induced NF-κB signaling pathway, our finding also provides another mechanism by which curcumin suppresses NF-κB activation. PMID:22303019

Homogeneous near surface activity distribution by double energy activation for TLA

NASA Astrophysics Data System (ADS)

Takács, S.; Ditrói, F.; Tárkányi, F.

2007-10-01

Thin layer activation (TLA) is a versatile tool for activating thin surface layers in order to study real-time the surface loss by wear, corrosion or erosion processes of the activated parts, without disassembling or stopping running mechanical structures or equipment. The research problem is the determination of the irradiation parameters to produce point-like or large area optimal activity-depth distribution in the sample. Different activity-depth profiles can be produced depending on the type of the investigated material and the nuclear reaction used. To produce activity that is independent of the depth up to a certain depth is desirable when the material removed from the surface by wear, corrosion or erosion can be collected completely. By applying dual energy irradiation the thickness of this quasi-constant activity layer can be increased or the deviation of the activity distribution from a constant value can be minimized. In the main, parts made of metals and alloys are suitable for direct activation, but by using secondary particle implantation the wear of other materials can also be studied in a surface range a few micrometers thick. In most practical cases activation of a point-like spot (several mm2) is enough to monitor the wear, corrosion or erosion, but for special problems relatively large surfaces areas of complicated spatial geometry need to be activated uniformly. Two ways are available for fulfilling this task, (1) production of large area beam spot or scanning the beam over the surface in question from the accelerator side, or (2) a programmed 3D movement of the sample from the target side. Taking into account the large variability of tasks occurring in practice, the latter method was chosen as the routine solution in our cyclotron laboratory.

Activation of Neurotensin Receptor Type 1 Attenuates Locomotor Activity

PubMed Central

Vadnie, Chelsea A.; Hinton, David J.; Choi, Sun; Choi, YuBin; Ruby, Christina L.; Oliveros, Alfredo; Prieto, Miguel L.; Park, Jun Hyun; Choi, Doo-Sup

2014-01-01

Intracerebroventricular administration of neurotensin (NT) suppresses locomotor activity. However, the brain regions that mediate the locomotor depressant effect of NT and receptor subtype-specific mechanisms involved are unclear. Using a brain-penetrating, selective NT receptor type 1 (NTS1) agonist PD149163, we investigated the effect of systemic and brain region-specific NTS1 activation on locomotor activity. Systemic administration of PD149163 attenuated the locomotor activity of C57BL/6J mice both in a novel environment and in their homecage. However, mice developed tolerance to the hypolocomotor effect of PD149163 (0.1 mg/kg, i.p.). Since NTS1 is known to modulate dopaminergic signaling, we examined whether PD149163 blocks dopamine receptor-mediated hyperactivity. Pretreatment with PD149163 (0.1 or 0.05 mg/kg, i.p.) inhibited D2R agonist bromocriptine (8 mg/kg, i.p.)-mediated hyperactivity. D1R agonist SKF81297 (8 mg/kg, i.p.)-induced hyperlocomotion was only inhibited by 0.1 mg/kg of PD149163. Since the nucleus accumbens (NAc) and medial prefrontal cortex (mPFC) have been implicated in the behavioral effects of NT, we examined whether microinjection of PD149163 into these regions reduces locomotion. Microinjection of PD149163 (2 pmol) into the NAc, but not the mPFC suppressed locomotor activity. In summary, our results indicate that systemic and intra-NAc activation of NTS1 is sufficient to reduce locomotion and NTS1 activation inhibits D2R-mediated hyperactivity. Our study will be helpful to identify pharmacological factors and a possible therapeutic window for NTS1-targeted therapies for movement disorders. PMID:24929110

Parent and child physical activity and sedentary time: do active parents foster active children?

PubMed

Jago, Russell; Fox, Kenneth R; Page, Angie S; Brockman, Rowan; Thompson, Janice L

2010-04-15

Physical activity has many positive effects on children's health while TV viewing has been associated with adverse health outcomes. Many children do not meet physical activity recommendations and exceed TV viewing guidelines. Parents are likely to be an important influence on their children's behaviour. There is an absence of information about the associations between parents' and children's physical activity and TV viewing. Year 6 children and their parent were recruited from 40 primary schools. Results are presented for the 340 parent-child dyads with accelerometer data that met a > or = 3 day inclusion criteria and the 431 parent-child dyads with complete self-reported TV viewing. Over 80% of the dyads with valid TV viewing data included mothers and their child. Mean minutes of moderate to vigorous physical activity (MVPA), minutes of sedentary time per day and counts per minute were assessed by accelerometer. Self-reported hours of TV viewing were coded into 3 groups (< 2 hours per day, 2-4 hours per day and >4 hours per day. Linear and multi-nominal regression models were run by child gender to examine parent-child associations. In linear regression models there was an association for the overall sedentary time of girls and their parents (t = 2.04. p = .020) but there was no association between girls' and parents' physical activity. There were no associations between parents' and boys' sedentary or physical activity time. For girls, the risk of watching more than 4 hours of TV per day, (reference = 2 hours of TV per day), was 3.67 times higher if the girl's parent watched 2-4 hours of TV per day (p = 0.037). For boys, the risk of watching more than 4 hours of TV per day, was 10.47 times higher if the boy's parent watched more than 4 hours of TV per day (p = 0.038). There are associations in the sedentary time of parents and daughters. Higher parental TV viewing was associated with an increased risk of high levels of TV viewing for both boys and girls. There

Active Play: Exploring the Influences on Children's School Playground Activities

ERIC Educational Resources Information Center

Hyndman, Brendon; Benson, Amanda; Telford, Amanda

2016-01-01

Because children spend so much of their time in schools, their playgrounds offer a good setting for promoting active play in young lives. Teachers, instead of considering active play a taxing demand on their busy day, have begun to develop an informal curriculum for it. The authors review the research on children's active play and explores its…

4-Aminoquinolines Active against Chloroquine-Resistant Plasmodium falciparum: Basis of Antiparasite Activity and Quantitative Structure-Activity Relationship Analyses▿

PubMed Central

Hocart, Simon J.; Liu, Huayin; Deng, Haiyan; De, Dibyendu; Krogstad, Frances M.; Krogstad, Donald J.

2011-01-01

Chloroquine (CQ) is a safe and economical 4-aminoquinoline (AQ) antimalarial. However, its value has been severely compromised by the increasing prevalence of CQ resistance. This study examined 108 AQs, including 68 newly synthesized compounds. Of these 108 AQs, 32 (30%) were active only against CQ-susceptible Plasmodium falciparum strains and 59 (55%) were active against both CQ-susceptible and CQ-resistant P. falciparum strains (50% inhibitory concentrations [IC50s], ≤25 nM). All AQs active against both CQ-susceptible and CQ-resistant P. falciparum strains shared four structural features: (i) an AQ ring without alkyl substitution, (ii) a halogen at position 7 (Cl, Br, or I but not F), (iii) a protonatable nitrogen at position 1, and (iv) a second protonatable nitrogen at the end of the side chain distal from the point of attachment to the AQ ring via the nitrogen at position 4. For activity against CQ-resistant parasites, side chain lengths of ≤3 or ≥10 carbons were necessary but not sufficient; they were identified as essential factors by visual comparison of 2-dimensional (2-D) structures in relation to the antiparasite activities of the AQs and were confirmed by computer-based 3-D comparisons and differential contour plots of activity against P. falciparum. The advantage of the method reported here (refinement of quantitative structure-activity relationship [QSAR] descriptors by random assignment of compounds to multiple training and test sets) is that it retains QSAR descriptors according to their abilities to predict the activities of unknown test compounds rather than according to how well they fit the activities of the compounds in the training sets. PMID:21383099

Structure-activity relationships of rosiglitazone for peroxisome proliferator-activated receptor gamma transrepression.

PubMed

Toyota, Yosuke; Nomura, Sayaka; Makishima, Makoto; Hashimoto, Yuichi; Ishikawa, Minoru

2017-06-15

Anti-inflammatory effects of peroxisome proliferator-activated receptor gamma (PPRAγ) ligands are thought to be largely due to PPARγ-mediated transrepression. Thus, transrepression-selective PPARγ ligands without agonistic activity or with only partial agonistic activity should exhibit anti-inflammatory properties with reduced side effects. Here, we investigated the structure-activity relationships (SARs) of PPARγ agonist rosiglitazone, focusing on transrepression activity. Alkenic analogs showed slightly more potent transrepression with reduced efficacy of transactivating agonistic activity. Removal of the alkyl group on the nitrogen atom improved selectivity for transrepression over transactivation. Among the synthesized compounds, 3l exhibited stronger transrepressional activity (IC 50 : 14μM) and weaker agonistic efficacy (11%) than rosiglitazone or pioglitazone. Copyright © 2017 Elsevier Ltd. All rights reserved.

Intrinsic resting-state activity predicts working memory brain activation and behavioral performance.

PubMed

Zou, Qihong; Ross, Thomas J; Gu, Hong; Geng, Xiujuan; Zuo, Xi-Nian; Hong, L Elliot; Gao, Jia-Hong; Stein, Elliot A; Zang, Yu-Feng; Yang, Yihong

2013-12-01

Although resting-state brain activity has been demonstrated to correspond with task-evoked brain activation, the relationship between intrinsic and evoked brain activity has not been fully characterized. For example, it is unclear whether intrinsic activity can also predict task-evoked deactivation and whether the rest-task relationship is dependent on task load. In this study, we addressed these issues on 40 healthy control subjects using resting-state and task-driven [N-back working memory (WM) task] functional magnetic resonance imaging data collected in the same session. Using amplitude of low-frequency fluctuation (ALFF) as an index of intrinsic resting-state activity, we found that ALFF in the middle frontal gyrus and inferior/superior parietal lobules was positively correlated with WM task-evoked activation, while ALFF in the medial prefrontal cortex, posterior cingulate cortex, superior frontal gyrus, superior temporal gyrus, and fusiform gyrus was negatively correlated with WM task-evoked deactivation. Further, the relationship between the intrinsic resting-state activity and task-evoked activation in lateral/superior frontal gyri, inferior/superior parietal lobules, superior temporal gyrus, and midline regions was stronger at higher WM task loads. In addition, both resting-state activity and the task-evoked activation in the superior parietal lobule/precuneus were significantly correlated with the WM task behavioral performance, explaining similar portions of intersubject performance variance. Together, these findings suggest that intrinsic resting-state activity facilitates or is permissive of specific brain circuit engagement to perform a cognitive task, and that resting activity can predict subsequent task-evoked brain responses and behavioral performance. Copyright © 2012 Wiley Periodicals, Inc.

Antioxidative Activities and Active Compounds of Extracts from Catalpa Plant Leaves

PubMed Central

Xu, Hongyu; Hu, Gege; Dong, Juane; Wei, Qin; Shao, Hongbo; Lei, Ming

2014-01-01

In order to screen the Catalpa plant with high antioxidant activity and confirm the corresponding active fractions from Catalpa ovata G. Don, C. fargesii Bur., and C. bungei C. A. Mey., total flavonoid contents and antioxidant activities of the extracts/fractions of Catalpa plant leaves were determined. The determined total flavonoid content and antioxidant activity were used as assessment criteria. Those compounds with antioxidant activity were isolated with silica gel column chromatography and ODS column chromatography. Our results showed that the total flavonoid content in C. bungei C. A. Mey. (30.07 mg/g·DW) was the highest, followed by those in C. fargesii Bur. (25.55 mg/g·DW) and C. ovata G. Don (24.96 mg/g·DW). According to the determination results of total flavonoid content and antioxidant activity in 3 clones of leaves of C. bungei C. A. Mey., the total flavonoid content and antioxidant activity in crude extracts from C. bungei C. A. Mey. 6 (CA6) leaves were the highest. Moreover, the results showed that the total flavonoid content and antioxidant activities of ethyl acetate (EA) fraction in ethanol crude extracts in CA6 leaves were the highest, followed by n-butanol, petroleum ether (PE), and water fractions. Two flavonoid compounds with antioxidant activity were firstly isolated based on EA fraction. The two compounds were luteolin (1) and apigenin (2), respectively. PMID:25431795

Impact of an active video game on healthy children's physical activity.

PubMed

Baranowski, Tom; Abdelsamad, Dina; Baranowski, Janice; O'Connor, Teresia Margareta; Thompson, Debbe; Barnett, Anthony; Cerin, Ester; Chen, Tzu-An

2012-03-01

This naturalistic study tests whether children receiving a new (to them) active video game spontaneously engage in more physical activity than those receiving an inactive video game, and whether the effect would be greater among children in unsafe neighborhoods, who might not be allowed to play outside. Participants were children 9 to 12 years of age, with a BMI >50th percentile, but <99th percentile; none of these children a medical condition that would preclude physical activity or playing video games. A randomized clinical trial assigned children to receiving 2 active or 2 inactive video games, the peripherals necessary to run the games, and a Wii console. Physical activity was monitored by using accelerometers for 5 weeks over the course of a 13-week experiment. Neighborhood safety was assessed with a 12 item validated questionnaire. There was no evidence that children receiving the active video games were more active in general, or at anytime, than children receiving the inactive video games. The outcomes were not moderated by parent perceived neighborhood safety, child BMI z score, or other demographic characteristics. These results provide no reason to believe that simply acquiring an active video game under naturalistic circumstances provides a public health benefit to children.

Evaluation of a novel canine activity monitor for at-home physical activity analysis.

PubMed

Yashari, Jonathan M; Duncan, Colleen G; Duerr, Felix M

2015-07-04

Accelerometers are motion-sensing devices that have been used to assess physical activity in dogs. However, the lack of a user-friendly, inexpensive accelerometer has hindered the widespread use of this objective outcome measure in veterinary research. Recently, a smartphone-based, affordable activity monitor (Whistle) has become available for measurement of at-home physical activity in dogs. The aim of this research was to evaluate this novel accelerometer. Eleven large breed, privately owned dogs wore a collar fitted with both the Whistle device and a previously validated accelerometer-based activity monitor (Actical) for a 24-h time period. Owners were asked to have their dogs resume normal daily activities. Total activity time obtained from the Whistle device in minutes was compared to the total activity count from the Actical device. Activity intensity from the Whistle device was calculated manually from screenshots of the activity bars displayed in the smartphone-application and compared to the activity count recorded by the Actical in the same 3-min time period. A total of 3740 time points were compared. There was a strong correlation between activity intensity of both devices for individual time points (Pearson's correlation coefficient 0.81, p < 0.0001). An even stronger correlation was observed between the total activity data between the two devices (Pearson's correlation coefficient 0.925, p < 0.0001). Activity data provided by the Whistle activity monitor may be used as an objective outcome measurement in dogs. The total activity time provided by the Whistle application offers an inexpensive method for obtaining at-home, canine, real-time physical activity data. Limitations of the Whistle device include the limited battery life, the need for manual derivation of activity intensity data and data transfer, and the requirement of Wi-Fi and Bluetooth availability for data transmission.

Serotonin Activates Overall Feeding by Activating Two Separate Neural Pathways in Caenorhabditis elegans

PubMed Central

Song, Bo-mi; Avery, Leon

2012-01-01

Food intake in the nematode Caenorhabditis elegans requires two distinct feeding motions, pharyngeal pumping and isthmus peristalsis. Bacteria, the natural food of C. elegans, activate both feeding motions (Croll, 1978; Horvitz et al., 1982; Chiang et al., 2006). The mechanisms by which bacteria activate the feeding motions are largely unknown. To understand the process, we studied how serotonin, an endogenous pharyngeal pumping activator whose action is triggered by bacteria, activates feeding motions. Here, we show that serotonin, like bacteria, activates overall feeding by activating isthmus peristalsis as well as pharyngeal pumping. During active feeding, the frequencies and the timing of onset of the two motions were distinct, but each isthmus peristalsis was coupled to the preceding pump. We found that serotonin activates the two feeding motions mainly by activating two separate neural pathways in response to bacteria. For activating pumping, the SER-7 serotonin receptor in the MC motor neurons in the feeding organ activated cholinergic transmission from MC to the pharyngeal muscles by activating the Gsα signaling pathway. For activating isthmus peristalsis, SER-7 in the M4 (and possibly M2) motor neuron in the feeding organ activated the G12α signaling pathway in a cell-autonomous manner, which presumably activates neurotransmission from M4 to the pharyngeal muscles. Based on our results and previous calcium imaging of pharyngeal muscles (Shimozono et al., 2004), we propose a model that explains how the two feeding motions are separately regulated yet coupled. The feeding organ may have evolved this way to support efficient feeding. PMID:22323705

Activation of neurotensin receptor type 1 attenuates locomotor activity.

PubMed

Vadnie, Chelsea A; Hinton, David J; Choi, Sun; Choi, YuBin; Ruby, Christina L; Oliveros, Alfredo; Prieto, Miguel L; Park, Jun Hyun; Choi, Doo-Sup

2014-10-01

Intracerebroventricular administration of neurotensin (NT) suppresses locomotor activity. However, the brain regions that mediate the locomotor depressant effect of NT and receptor subtype-specific mechanisms involved are unclear. Using a brain-penetrating, selective NT receptor type 1 (NTS1) agonist PD149163, we investigated the effect of systemic and brain region-specific NTS1 activation on locomotor activity. Systemic administration of PD149163 attenuated the locomotor activity of C57BL/6J mice both in a novel environment and in their homecage. However, mice developed tolerance to the hypolocomotor effect of PD149163 (0.1 mg/kg, i.p.). Since NTS1 is known to modulate dopaminergic signaling, we examined whether PD149163 blocks dopamine receptor-mediated hyperactivity. Pretreatment with PD149163 (0.1 or 0.05 mg/kg, i.p.) inhibited D2R agonist bromocriptine (8 mg/kg, i.p.)-mediated hyperactivity. D1R agonist SKF-81297 (8 mg/kg, i.p.)-induced hyperlocomotion was only inhibited by 0.1 mg/kg of PD149163. Since the nucleus accumbens (NAc) and medial prefrontal cortex (mPFC) have been implicated in the behavioral effects of NT, we examined whether microinjection of PD149163 into these regions reduces locomotion. Microinjection of PD149163 (2 pmol) into the NAc, but not the mPFC suppressed locomotor activity. In summary, our results indicate that systemic and intra-NAc activation of NTS1 is sufficient to reduce locomotion and NTS1 activation inhibits D2R-mediated hyperactivity. Our study will be helpful to identify pharmacological factors and a possible therapeutic window for NTS1-targeted therapies for movement disorders. Copyright © 2014 Elsevier Ltd. All rights reserved.

Vision Drives Correlated Activity without Patterned Spontaneous Activity in Developing Xenopus Retina

PubMed Central

Demas, James A.; Payne, Hannah; Cline, Hollis T.

2011-01-01

Developing amphibians need vision to avoid predators and locate food before visual system circuits fully mature. Xenopus tadpoles can respond to visual stimuli as soon as retinal ganglion cells (RGCs) innervate the brain, however, in mammals, chicks and turtles, RGCs reach their central targets many days, or even weeks, before their retinas are capable of vision. In the absence of vision, activity-dependent refinement in these amniote species is mediated by waves of spontaneous activity that periodically spread across the retina, correlating the firing of action potentials in neighboring RGCs. Theory suggests that retinorecipient neurons in the brain use patterned RGC activity to sharpen the retinotopy first established by genetic cues. We find that in both wild type and albino Xenopus tadpoles, RGCs are spontaneously active at all stages of tadpole development studied, but their population activity never coalesces into waves. Even at the earliest stages recorded, visual stimulation dominates over spontaneous activity and can generate patterns of RGC activity similar to the locally correlated spontaneous activity observed in amniotes. In addition, we show that blocking AMPA and NMDA type glutamate receptors significantly decreases spontaneous activity in young Xenopus retina, but that blocking GABAA receptor blockers does not. Our findings indicate that vision drives correlated activity required for topographic map formation. They further suggest that developing retinal circuits in the two major subdivisions of tetrapods, amphibians and amniotes, evolved different strategies to supply appropriately patterned RGC activity to drive visual circuit refinement. PMID:21312343

Active, capable, and potentially active faults - a paleoseismic perspective

USGS Publications Warehouse

Machette, M.N.

2000-01-01

Maps of faults (geologically defined source zones) may portray seismic hazards in a wide range of completeness depending on which types of faults are shown. Three fault terms - active, capable, and potential - are used in a variety of ways for different reasons or applications. Nevertheless, to be useful for seismic-hazards analysis, fault maps should encompass a time interval that includes several earthquake cycles. For example, if the common recurrence in an area is 20,000-50,000 years, then maps should include faults that are 50,000-100,000 years old (two to five typical earthquake cycles), thus allowing for temporal variability in slip rate and recurrence intervals. Conversely, in more active areas such as plate boundaries, maps showing faults that are <10,000 years old should include those with at least 2 to as many as 20 paleoearthquakes. For the International Lithosphere Programs' Task Group II-2 Project on Major Active Faults of the World our maps and database will show five age categories and four slip rate categories that allow one to select differing time spans and activity rates for seismic-hazard analysis depending on tectonic regime. The maps are accompanied by a database that describes evidence for Quaternary faulting, geomorphic expression, and paleoseismic parameters (slip rate, recurrence interval and time of most recent surface faulting). These maps and databases provide an inventory of faults that would be defined as active, capable, and potentially active for seismic-hazard assessments.

Light and dark-activated biocidal activity of conjugated polyelectrolytes.

PubMed

Ji, Eunkyung; Corbitt, Thomas S; Parthasarathy, Anand; Schanze, Kirk S; Whitten, David G

2011-08-01

This Spotlight on Applications provides an overview of a research program that has focused on the development and mechanistic study of cationic conjugated polyelectrolytes (CPEs) that function as light- and dark-active biocidal agents. Investigation has centered on poly-(phenylene ethynylene) (PPE) type conjugated polymers that are functionalized with cationic quaternary ammonium solubilizing groups. These polymers are found to interact strongly with Gram-positive and Gram-negative bacteria, and upon illumination with near-UV and visible light act to rapidly kill the bacteria. Mechanistic studies suggest that the cationic PPE-type polymers efficiently sensitize singlet oxygen ((1)O(2)), and this cytotoxic agent is responsible for initiating the sequence of events that lead to light-activated bacterial killing. Specific CPEs also exhibit dark-active antimicrobial activity, and this is believed to arise due to interactions between the cationic/lipophilic polymers and the negatively charged outer membrane characteristic of Gram-negative bacteria. Specific results are shown where a cationic CPE with a degree of polymerization of 49 exhibits pronounced light-activated killing of E. coli when present in the cell suspension at a concentration of 1 μg mL(-1).

Forecasting Occurrences of Activities.

PubMed

Minor, Bryan; Cook, Diane J

2017-07-01

While activity recognition has been shown to be valuable for pervasive computing applications, less work has focused on techniques for forecasting the future occurrence of activities. We present an activity forecasting method to predict the time that will elapse until a target activity occurs. This method generates an activity forecast using a regression tree classifier and offers an advantage over sequence prediction methods in that it can predict expected time until an activity occurs. We evaluate this algorithm on real-world smart home datasets and provide evidence that our proposed approach is most effective at predicting activity timings.

Theory-based physical activity beliefs by race and activity levels among older adults.

PubMed

Kosma, Maria; Cardinal, Bradley J

2016-01-01

Given the benefits of physical activity and the high proportion of inactivity among older adults, the purpose was to elicit theory-based behavioral, normative, and control physical activity beliefs among 140 educationally and economically diverse older adults and compare their beliefs by race (Blacks vs. Whites) and physical activity levels (inactive/underactive vs. highly active individuals). This was an elicitation study that took place in eight, mostly rural community settings in a Southeastern US state, such as Council of Aging Offices, retirement centers, and churches. Participants' behavioral, normative, and control beliefs were elicited via in person interviews. A valid and reliable questionnaire was also used to assess their physical activity levels. According to the content analysis, inactive/underactive participants reported fewer physical activity advantages than highly active participants. Common physical activity advantages between the two groups were overall health, emotional functioning, and physical functioning. Similar physical activity advantages were reported among Blacks and Whites with overall health being the most important advantage. The most common physical activity disadvantages and barriers for all four groups were falls, injuries, pain, and health issues. Inactive/underactive individuals and Blacks tended to report more disadvantages and barriers than their peers. Common physical activity supporters were family members, friends and peers, and health-care professionals. In their physical activity motivational programs, health promoters should reinforce physical activity benefits, social support, access to activity programs, and safety when intervening among older adults.

College Student Environmental Activism: How Experiences and Identities Influence Environmental Activism Approaches

ERIC Educational Resources Information Center

King, Laura A. H.

2016-01-01

College student environmental activism is one way students civically engage in addressing social issues. This study explores the environmental activism of twelve college students and how their experiences outside of college and in college influenced their activism. In addition, how students' identities influenced their approach to activism was…

The Role of Physical Activity Assessments for School-Based Physical Activity Promotion

ERIC Educational Resources Information Center

Welk, Gregory J.

2008-01-01

The emphasis in public health on lifestyle physical activity in recent years has focused attention on the promotion of lifetime physical activity as the primary objective of physical education. If used properly, physical activity and physical fitness assessments can enhance individual promotion of physical activity and also provide valuable…

Ladders to Literacy: A Preschool Activity Book [and] A Kindergarten Activity Book.

ERIC Educational Resources Information Center

Notari-Syverson, Angela; O'Connor, Rollanda E.; Vadasy, Patricia F.

This preschool activity book and kindergarten activity book set provide a curriculum of 114 activities to develop emergent literacy skills in preschool and kindergarten children. The curriculum is based on research findings and is intended to be ecologically valid, embedding activities into developmentally appropriate, daily classroom routines and…

Active Cytokinins

PubMed Central

Mornet, René; Theiler, Jane B.; Leonard, Nelson J.; Schmitz, Ruth Y.; Moore, F. Hardy; Skoog, Folke

1979-01-01

Four series of azidopurines have been synthesized and tested for cytokinin activity in the tobacco callus bioassay: 2- and 8-azido-N6-benzyladenines, -N6-(Δ2-isopentenyl)adenines, and -zeatins, and N6-(2- and 4-azidobenzyl)adenines. The compounds having 2-azido substitution on the adenine ring are as active as the corresponding parent compounds, while those with 8-azido substitution are about 10 or more times as active. The 8-azidozeatin, which is the most active cytokinin observed, exhibited higher than minimal detectable activity at 1.2 × 10−5 micromolar, the lowest concentration tested. The shape of the growth curve indicates that even a concentration as low as 5 × 10−6 micromolar would probably be effective. By comparison, the lowest active concentration ever reported for zeatin has been 5 × 10−5 micromolar, representing a sensitivity rarely attained. All of the azido compounds have been submitted to photolysis in aqueous ethanol, and the photoproducts have been detected and identified by low and high resolution mass spectrometry. They are rationalized as products of abstraction and insertion reactions of the intermediate nitrenes. The potential of the major released products as cytokinins was also assessed by bioassay. 2-Azido-N6-(Δ2-isopentenyl)adenine competed with [14C]kinetin for the cytokinin-binding protein isolated from wheat germ. When the azido compound was photolysed in the presence of this protein, its attachment effectively blocked the binding of [14C]kinetin. PMID:16661017

Computer-aided drug design for AMP-activated protein kinase activators.

PubMed

Wang, Zhanli; Huo, Jianxin; Sun, Lidan; Wang, Yongfu; Jin, Hongwei; Yu, Hui; Zhang, Liangren; Zhou, Lishe

2011-09-01

AMP-activated protein kinase (AMPK) is an important therapeutic target for the potential treatment of metabolic disorders, cardiovascular disease and cancer. Recently, various classes of compounds that activate AMPK by direct or indirect interactions have been reported. The importance of computer-aided drug design approaches in the search for potent activators of AMPK is now established, including structure-based design, ligand-based design, fragment-based design, as well as structural analysis. This review article highlights the computer-aided drug design approaches utilized to discover of activators targeting AMPK. The principles, advantages or limitation of the different methods are also being discussed together with examples of applications taken from the literatures.

Inferring deep-brain activity from cortical activity using functional near-infrared spectroscopy

PubMed Central

Liu, Ning; Cui, Xu; Bryant, Daniel M.; Glover, Gary H.; Reiss, Allan L.

2015-01-01

Functional near-infrared spectroscopy (fNIRS) is an increasingly popular technology for studying brain function because it is non-invasive, non-irradiating and relatively inexpensive. Further, fNIRS potentially allows measurement of hemodynamic activity with high temporal resolution (milliseconds) and in naturalistic settings. However, in comparison with other imaging modalities, namely fMRI, fNIRS has a significant drawback: limited sensitivity to hemodynamic changes in deep-brain regions. To overcome this limitation, we developed a computational method to infer deep-brain activity using fNIRS measurements of cortical activity. Using simultaneous fNIRS and fMRI, we measured brain activity in 17 participants as they completed three cognitive tasks. A support vector regression (SVR) learning algorithm was used to predict activity in twelve deep-brain regions using information from surface fNIRS measurements. We compared these predictions against actual fMRI-measured activity using Pearson’s correlation to quantify prediction performance. To provide a benchmark for comparison, we also used fMRI measurements of cortical activity to infer deep-brain activity. When using fMRI-measured activity from the entire cortex, we were able to predict deep-brain activity in the fusiform cortex with an average correlation coefficient of 0.80 and in all deep-brain regions with an average correlation coefficient of 0.67. The top 15% of predictions using fNIRS signal achieved an accuracy of 0.7. To our knowledge, this study is the first to investigate the feasibility of using cortical activity to infer deep-brain activity. This new method has the potential to extend fNIRS applications in cognitive and clinical neuroscience research. PMID:25798327

Tea enhances insulin activity.

PubMed

Anderson, Richard A; Polansky, Marilyn M

2002-11-20

The most widely known health benefits of tea relate to the polyphenols as the principal active ingredients in protection against oxidative damage and in antibacterial, antiviral, anticarcinogenic, and antimutagenic activities, but polyphenols in tea may also increase insulin activity. The objective of this study was to determine the insulin-enhancing properties of tea and its components. Tea, as normally consumed, was shown to increase insulin activity >15-fold in vitro in an epididymal fat cell assay. Black, green, and oolong teas but not herbal teas, which are not teas in the traditional sense because they do not contain leaves of Camellia senensis, were all shown to increase insulin activity. High-performance liquid chromatography fractionation of tea extracts utilizing a Waters SymmetryPrep C18 column showed that the majority of the insulin-potentiating activity for green and oolong teas was due to epigallocatechin gallate. For black tea, the activity was present in several regions of the chromatogram corresponding to, in addition to epigallocatechin gallate, tannins, theaflavins, and other undefined compounds. Several known compounds found in tea were shown to enhance insulin with the greatest activity due to epigallocatechin gallate followed by epicatechin gallate, tannins, and theaflavins. Caffeine, catechin, and epicatechin displayed insignificant insulin-enhancing activities. Addition of lemon to the tea did not affect the insulin-potentiating activity. Addition of 5 g of 2% milk per cup decreased the insulin-potentiating activity one-third, and addition of 50 g of milk per cup decreased the insulin-potentiating activity approximately 90%. Nondairy creamers and soy milk also decreased the insulin-enhancing activity. These data demonstrate that tea contains in vitro insulin-enhancing activity and the predominant active ingredient is epigallocatechin gallate.

Bonus Activity Book.

ERIC Educational Resources Information Center

Learning, 1992

1992-01-01

Provides on-task activities to fill in unexpected extra moments in elementary classes. The activities require little preparation and take 5-15 minutes to complete. There are activities for math, language arts, social science, science, critical thinking, and computer. An outer space board game is also included. (SM)

Active chiral fluids.

PubMed

Fürthauer, S; Strempel, M; Grill, S W; Jülicher, F

2012-09-01

Active processes in biological systems often exhibit chiral asymmetries. Examples are the chirality of cytoskeletal filaments which interact with motor proteins, the chirality of the beat of cilia and flagella as well as the helical trajectories of many biological microswimmers. Here, we derive constitutive material equations for active fluids which account for the effects of active chiral processes. We identify active contributions to the antisymmetric part of the stress as well as active angular momentum fluxes. We discuss four types of elementary chiral motors and their effects on a surrounding fluid. We show that large-scale chiral flows can result from the collective behavior of such motors even in cases where isolated motors do not create a hydrodynamic far field.

Porous properties of activated carbons from waste newspaper prepared by chemical and physical activation.

PubMed

Okada, Kiyoshi; Yamamoto, Nobuo; Kameshima, Yoshikazu; Yasumori, Atsuo

2003-06-01

Activated carbons were prepared from old newspaper and paper prepared from simulated paper sludge by chemical activation using various alkali carbonates and hydroxides as activating reagents and also by physical activation using steam. In the chemical activation, the influence of oxidation, carbonization, and activation on the porous properties of the resulting activated carbons was investigated. The specific surface areas (S(BET)) of the activated carbons prepared by single-step activation (direct activation without oxidation and carbonization) were higher than those resulting from two-step activation (oxidation-activation and carbonization-activation) and three-step activation (oxidation-carbonization-activation) methods. The S(BET) values were strongly dependent on the activating reagents and the activating conditions, being >1000 m(2)/g using K(2)CO(3), Rb(2)CO(3), Cs(2)CO(3), and KOH as activating reagents but <1000 m(2)/g using Li(2)CO(3), Na(2)CO(3), and NaOH. These differences in S(BET) values are suggested to be related to the ionic radii of the alkalis used as activating reagents. The microstructures of the higher S(BET) samples show a complete loss of fiber shape but those of the lower S(BET) samples maintain the shape. In the physical activation, the porous properties of the activated carbons prepared by the single-step method were examined as a function of the production conditions such as activation temperature, activation time, steam concentration, and flow rate of the carrier gas. The maximum S(BET) and total pore volume (V(P)) were 1086 m(2)/g and 1.01 ml/g, obtained by activation at 850 degrees C for 2 h, flowing 20 mol% of steam in nitrogen gas at 0.5 l/min. A correlation was found between S(BET) and the yield of the product, the maximum S(BET) value corresponding to a product yield of about 10%. This result is suggested to result from competition between pore formation and surface erosion. Compared with chemically activated carbons using K(2)CO

Trunk muscle activation during dynamic weight-training exercises and isometric instability activities.

PubMed

Hamlyn, Nicolle; Behm, David G; Young, Warren B

2007-11-01

The purpose of this study was to examine the extent of activation in various trunk muscles during dynamic weight-training and isometric instability exercises. Sixteen subjects performed squats and deadlifts with 80% 1 repetition maximum (1RM), as well as with body weight as resistance and 2 unstable calisthenic-type exercises (superman and sidebridge). Electromyographic (EMG) activity was measured from the lower abdominals (LA), external obliques (EO), upper lumbar erector spinae (ULES), and lumbar-sacral erector spinae (LSES) muscle groups. Results indicated that the LSES EMG activity during the 80% 1RM squat significantly exceeded 80% 1RM deadlift LSES EMG activity by 34.5%. The LSES EMG activity of the 80% 1RM squat also exceeded the body weight squat, deadlift, superman, and sidebridge by 56, 56.6, 65.5, and 53.1%, respectively. The 80% 1RM deadlift ULES EMG activity significantly exceeded the 80% 1RM squat exercise by 12.9%. In addition, the 80% 1RM deadlift ULES EMG activity also exceeded the body weight squat, deadlift, superman, and sidebridge exercises by 66.7, 65.5, 69.3, and 68.6%, respectively. There were no significant changes in EO or LA activity. Therefore, the augmented activity of the LSES and ULES during 80% 1RM squat and deadlift resistance exercises exceeded the activation levels achieved with the same exercises performed with body weight and selected instability exercises. Individuals performing upright, resisted, dynamic exercises can achieve high trunk muscle activation and thus may not need to add instability device exercises to augment core stability training.

Physical activity during hospitalization: Activities and preferences of adults versus older adults.

PubMed

Meesters, Jorit; Conijn, D; Vermeulen, H M; Vliet Vlieland, Tpm

2018-04-16

Inactivity during hospitalization leads to a functional decline and an increased risk of complications. To date, studies focused on older adults. This study aims to compare the physical activities performed by older adult and adult hospitalized patients. Patients hospitalized for >3 days at a university hospital completed a questionnaire regarding their physical activities (% of days on which an activity was performed divided by the length of stay) and physical activity needs during hospitalization. Crude and adjusted comparisons of older adult (>60 years) and adult (≤60 years) patients were performed using parametric testing and regression analyses. Of 524 patients, 336 (64%) completed the questionnaire, including 166 (49%) older adult patients. On average, the patients were physically active on 35% or less of the days during their hospitalization. Linear regression analysis showed no significant associations between being an older adult and performing physical activities after adjusting for gender, length of stay, surgical intervention, and meeting physical activity recommendations prior to hospitalization. Most patients were well informed regarding physical activity during hospitalization; however, the older adult patients reported a need for information regarding physical activities after hospitalization more frequently (odds ratios, 2.47) after adjusting for educational level, gender, and physical therapy during hospitalization. Both older adult and adult patients are physically inactive during hospitalization, and older adult patients express a greater need for additional information regarding physical activity after hospitalization than adult patients. Therefore, personalized strategies that inform and motivate patients to resume physical activities during hospitalization are needed regardless of age.

Association between recent sports activity, sports activity in young adulthood, and stroke.

PubMed

Grau, Armin J; Barth, Cordula; Geletneky, Beate; Ling, Paul; Palm, Frederik; Lichy, Christoph; Becher, Heiko; Buggle, Florian

2009-02-01

Leisure-time physical activity protects from stroke. It is insufficiently established whether early lifetime physical activity is independently protective and whether some etiologic stroke subgroups particularly benefit from physical activity. We tested the hypothesis that both recent and early-adulthood sports activities are associated with reduced odds of stroke and analyzed their effects in stroke subtypes. We performed a case-control study of 370 patients with acute stroke or transient ischemic attack (TIA) and 370 age- and sex-matched control subjects randomly selected from the population and assessed recent and young adulthood sports activities and their weekly duration in standardized interviews. Recent regular sports activities were less often reported by patients (94/370, 25.4%) than by control subjects (162/370, 43.8%; P<0.0001). After adjustment for vascular risk factors, education, and other factors, recent participation in sports was significantly associated with reduced odds of stroke/TIA (odds ratio=0.64; 95% CI, 0.43 to 0.96). Both groups did not differ with regard to sports activities in young adulthood. More control subjects (69/365, 18.9%) than patients (25/361, 6.9%) participated in sports recently after not having been active in young adulthood, and such a pattern was associated with reduced odds of stroke/TIA in multivariable analysis (odds ratio=0.37; 95% CI, 0.21 to 0.85). Our study supports previous results that have shown stroke protection by physical activity. Results suggest that continuous lifetime activity or starting activities during later adulthood is required to reduce stroke risk.

Control of Promatrilysin (MMP7) Activation and Substrate-specific Activity by Sulfated Glycosaminoglycans*

PubMed Central

Ra, Hyun-Jeong; Harju-Baker, Susanna; Zhang, Fuming; Linhardt, Robert J.; Wilson, Carole L.; Parks, William C.

2009-01-01

Matrix metalloproteinases are maintained in an inactive state by a bond between the thiol of a conserved cysteine in the prodomain and a zinc atom in the catalytic domain. Once this bond is disrupted, MMPs become active proteinases and can act on a variety of extracellular protein substrates. In vivo, matrilysin (MMP7) activates pro-α-defensins (procryptdins), but in vitro, processing of these peptides is slow, with about 50% conversion in 8–12 h. Similarly, autolytic activation of promatrilysin in vitro can take up to 12–24 h for 50% conversion. These inefficient reactions suggest that natural cofactors enhance the activation and activity of matrilysin. We determined that highly sulfated glycosaminoglycans (GAG), such as heparin, chondroitin-4,6-sulfate (CS-E), and dermatan sulfate, markedly enhanced (>50-fold) the intermolecular autolytic activation of promatrilysin and the activity of fully active matrilysin to cleave specific physiologic substrates. In contrast, heparan sulfate and less sulfated forms of chondroitin sulfate did not augment matrilysin activation or activity. Chondroitin-2,6-sulfate (CS-D) also did not enhance matrilysin activity, suggesting that the presentation of sulfates is more important than the overall degree of sulfation. Surface plasmon resonance demonstrated that promatrilysin bound heparin (KD, 400 nm) and CS-E (KD, 630 nm). Active matrilysin bound heparin (KD, 150 nm) but less so to CS-E (KD, 60 μm). Neither form bound heparan sulfate. These observations demonstrate that sulfated GAGs regulate matrilysin activation and its activity against specific substrates. PMID:19654318

Strand displacement activated peroxidase activity of hemin for fluorescent DNA sensing.

PubMed

Wang, Quanbo; Xu, Nan; Gui, Zhen; Lei, Jianping; Ju, Huangxian; Yan, Feng

2015-10-07

To efficiently regulate the catalytic activity of the peroxidase mimic hemin, this work designs a double-stranded DNA probe containing an intermolecular dimer of hemin, whose peroxidase activity can be activated by a DNA strand displacement reaction. The double-stranded probe is prepared by annealing two strands of hemin labelled DNA oligonucleotides. Using the fluorescent oxidation product of tyramine by H2O2 as a tracing molecule, the low peroxidase activity of the hemin dimer ensures a low fluorescence background. The strand displacement reaction of the target DNA dissociates the hemin dimer and thus significantly increases the catalytic activity of hemin to produce a large amount of dityramine for fluorescence signal readout. Based on the strand displacement regulated peroxidase activity, a simple and sensitive homogeneous fluorescent DNA sensing method is proposed. The detection can conveniently be carried out in a 96-well plate within 20 min with a detection limit of 0.18 nM. This method shows high specificity, which can effectively distinguish single-base mismatched DNA from perfectly matched target DNA. The DNA strand displacement regulated catalytic activity of hemin has promising application in the determination of various DNA analytes.

Influence of plasma-activated compounds on melanogenesis and tyrosinase activity

PubMed Central

Ali, Anser; Ashraf, Zaman; Kumar, Naresh; Rafiq, Muhammad; Jabeen, Farukh; Park, Ji Hoon; Choi, Ki Hong; Lee, SeungHyun; Seo, Sung-Yum; Choi, Eun Ha; Attri, Pankaj

2016-01-01

Many organic chemists around the world synthesize medicinal compounds or extract multiple compounds from plants in order to increase the activity and quality of medicines. In this work, we synthesized new eugenol derivatives (ED) and then treated them with an N2 feeding gas atmospheric pressure plasma jet (APPJ) to increase their utility. We studied the tyrosinase-inhibition activity (activity test) and structural changes (circular dichroism) of tyrosinase with ED and plasma activated eugenol derivatives (PAED) in a cell-free environment. Later, we used docking studies to determine the possible interaction sites of ED and PAED compounds with tyrosinase enzyme. Moreover, we studied the possible effect of ED and PAED on melanin synthesis and its mechanism in melanoma (B16F10) cells. Additionally, we investigated the structural changes that occurred in activated ED after plasma treatment using nuclear magnetic resonance (NMR). Hence, this study provides a new perspective on PAED for the field of plasma medicine. PMID:26931617

Antioxidant activity and antibacterial activity of peach gum derived oligosaccharides.

PubMed

Yao, Xing-Cun; Cao, Yan; Wu, Sheng-Jun

2013-11-01

In this study, peach gum derived oligosaccharides (PGDO) were prepared from peach gum polysaccharides by hydrolysis using hydrogen peroxide (H2O2) under following conditions: time, 8h; temperature, 55 °C; H2O2 concentration, 4% (v/v); and NaOH concentration, 2.0 M. The antioxidant activity and antibacterial activity of PGDO were estimated. There were no significant chemical changes in the backbones of the peach gum polysaccharides treated with H2O2. The PGDO showed high hydroxyl radical scavenging activity (86.12%) and 2, 2-diphenyl-β-picrylhydrazyl radical scavenging activity (91.70%) at the concentration of 100 μg/mL as well as high reducing capacity at the concentration of 50 μg/mL. In addition the PGDO had high antibacterial activity against Bacillus subtilis, Staphylococcus aureus, and Escherichia coli at the concentration of 100 μg/mL. Copyright © 2013 Elsevier B.V. All rights reserved.

Active Flow Control Activities at NASA Langley

NASA Technical Reports Server (NTRS)

Anders, Scott G.; Sellers, William L., III; Washburn, Anthony E.

2004-01-01

NASA Langley continues to aggressively investigate the potential advantages of active flow control over more traditional aerodynamic techniques. This paper provides an update to a previous paper and describes both the progress in the various research areas and the significant changes in the NASA research programs. The goals of the topics presented are focused on advancing the state of knowledge and understanding of controllable fundamental mechanisms in fluids as well as to address engineering challenges. An organizational view of current research activities at NASA Langley in active flow control as supported by several projects is presented. On-center research as well as NASA Langley funded contracts and grants are discussed at a relatively high level. The products of this research are to be demonstrated either in bench-top experiments, wind-tunnel investigations, or in flight as part of the fundamental NASA R&D program and then transferred to more applied research programs within NASA, DOD, and U.S. industry.

Optimizing post activation potentiation for explosive activities in competitive sports

PubMed Central

Gołaś, Artur; Maszczyk, Adam; Mikołajec, Kazimierz; Stastny, Petr

2016-01-01

Abstract Post activation potentiation (PAP) has shown improved performance during movements requiring large muscular power output following contractions under near maximal load conditions. PAP can be described as an acute enhancement of performance or an enhancement of factors determining an explosive sports activity following a preload stimulus. In practice, PAP has been achieved by complex training, which involves a combination of a heavy loaded exercise followed by a biomechanically similar explosive activity, best if specific for a particular sport discipline. The main objective of this study was to investigate the effects of PAP on performance in explosive motor activities specific for basketball, luge and athletics throws. The novel approach to the experiments included individualized recovery time (IRT) between the conditioning exercise and the explosive activity. Additionally, the research groups were homogenous and included only competitive athletes of similar age and training experience. Thirty one well trained athletes from 3 different sport disciplines participated in the study. All athletes performed a heavy loaded conditioning activity (80-130%1RM) followed by a biomechanically similar explosive exercise, during which power (W) or the rate of power development (W/s/kg) was evaluated. The results of our experiment confirmed the effectiveness of PAP with well-trained athlets during explosive motor activities such as jumping, throwing and pushing. Additionally, our research showed that eccentric supramaximal intensities (130% 1RM) can be effective in eliciting PAP in strength trained athletes. Our experiments also showed that the IRT should be individualized because athletes differ in the strength level, training experience and muscle fiber structure. In the three experiments conducted with basketball players, track and field athletes and luge athletes, the optimal IRT equaled 6 min. This justifies the need to individualize the volume and intensity of the

The Use of Multiple Slate Devices to Support Active Reading Activities

ERIC Educational Resources Information Center

Chen, Nicholas Yen-Cherng

2012-01-01

Reading activities in the classroom and workplace occur predominantly on paper. Since existing electronic devices do not support these reading activities as well as paper, users have difficulty taking full advantage of the affordances of electronic documents. This dissertation makes three main contributions toward supporting active reading…

An Active Learning Activity to Reinforce the Design Components of the Corticosteroids

PubMed Central

Mandela, Prashant

2018-01-01

Despite the popularity of active learning applications over the past few decades, few activities have been reported for the field of medicinal chemistry. The purpose of this study is to report a new active learning activity, describe participant contributions, and examine participant performance on the assessment questions mapped to the objective covered by the activity. In this particular activity, students are asked to design two novel corticosteroids as a group (6–8 students per group) based on the design characteristics of marketed corticosteroids covered in lecture coupled with their pharmaceutics knowledge from the previous semester and then defend their design to the class through an interactive presentation model. Although class performance on the objective mapped to this material on the assessment did not reach statistical significance, use of this activity has allowed fruitful discussion of misunderstood concepts and facilitated multiple changes to the lecture presentation. As pharmacy schools continue to emphasize alternative learning pedagogies, publication of previously implemented activities demonstrating their use will help others apply similar methodologies. PMID:29401733

An Active Learning Activity to Reinforce the Design Components of the Corticosteroids.

PubMed

Slauson, Stephen R; Mandela, Prashant

2018-02-05

Despite the popularity of active learning applications over the past few decades, few activities have been reported for the field of medicinal chemistry. The purpose of this study is to report a new active learning activity, describe participant contributions, and examine participant performance on the assessment questions mapped to the objective covered by the activity. In this particular activity, students are asked to design two novel corticosteroids as a group (6-8 students per group) based on the design characteristics of marketed corticosteroids covered in lecture coupled with their pharmaceutics knowledge from the previous semester and then defend their design to the class through an interactive presentation model. Although class performance on the objective mapped to this material on the assessment did not reach statistical significance, use of this activity has allowed fruitful discussion of misunderstood concepts and facilitated multiple changes to the lecture presentation. As pharmacy schools continue to emphasize alternative learning pedagogies, publication of previously implemented activities demonstrating their use will help others apply similar methodologies.

Active touch sensing

PubMed Central

Prescott, Tony J.; Diamond, Mathew E.; Wing, Alan M.

2011-01-01

Active sensing systems are purposive and information-seeking sensory systems. Active sensing usually entails sensor movement, but more fundamentally, it involves control of the sensor apparatus, in whatever manner best suits the task, so as to maximize information gain. In animals, active sensing is perhaps most evident in the modality of touch. In this theme issue, we look at active touch across a broad range of species from insects, terrestrial and marine mammals, through to humans. In addition to analysing natural touch, we also consider how engineering is beginning to exploit physical analogues of these biological systems so as to endow robots with rich tactile sensing capabilities. The different contributions show not only the varieties of active touch—antennae, whiskers and fingertips—but also their commonalities. They explore how active touch sensing has evolved in different animal lineages, how it serves to provide rapid and reliable cues for controlling ongoing behaviour, and even how it can disintegrate when our brains begin to fail. They demonstrate that research on active touch offers a means both to understand this essential and primary sensory modality, and to investigate how animals, including man, combine movement with sensing so as to make sense of, and act effectively in, the world. PMID:21969680

FL Activities & Festivals.

ERIC Educational Resources Information Center

American Council on the Teaching of Foreign Languages, Hastings-on-Hudson, NY.

A collection of student, class, and school foreign language activities suggests a variety of projects and describes three specific school efforts. The suggested activities include: (1) individual student efforts such as writing to pen-pals; (2) group activities such as a foreign language auction or sing-along; (3) group projects for the school…

Comparison of laser-activated tissue solders and thrombin-activated cryoprecipitate for wound closure

NASA Astrophysics Data System (ADS)

Kayton, Mark L.; Libutti, Steven K.; Bessler, Marc; Allendorf, John D. F.; Eiref, Simon D.; Marx, Gerard; Mou, Xiaode; Morales, Alfredo M.; Treat, Michael R.; Nowygrod, Roman

1994-09-01

To determine the relative strengths of various biologic adhesives at several timepoints, we compared thrombin-activated SD (solvent-detergent treated) cryoprecipitate with laser- activated SD cryoprecipitate and a laser-activated, albumin-based glue. Male Sprague-Dawley rats (n equals 79) received four, 3-cm, dorsal skin incisions which were closed with either laser- activated cryoprecipitate, laser-activated albumin solder, thrombin-activated cryoprecipitate, or standard skin staples. The cryoprecipitate was derived from pooled human plasma and was treated with a solvent-detergent process, rendering it free of envelope-coated viruses (i.e., HBV, HIV). An 808-nm diode laser was used to activate each solder with an average duration of exposure of 75 seconds per incision. Animals were sacrificed for evaluation of wound tensile strength and histology at 0 hours, 2 hours, 4 hours, and 4 days. At all timepoints tested, laser-activated solders were significantly stronger than thrombin-activated cryoprecipitate (p < 0.03) and control wounds (p < 0.003). There was no significant difference in tensile strength between the two types of laser-activated solder at any timepoint.

Perceiving active listening activates the reward system and improves the impression of relevant experiences.

PubMed

Kawamichi, Hiroaki; Yoshihara, Kazufumi; Sasaki, Akihiro T; Sugawara, Sho K; Tanabe, Hiroki C; Shinohara, Ryoji; Sugisawa, Yuka; Tokutake, Kentaro; Mochizuki, Yukiko; Anme, Tokie; Sadato, Norihiro

2015-01-01

Although active listening is an influential behavior, which can affect the social responses of others, the neural correlates underlying its perception have remained unclear. Sensing active listening in social interactions is accompanied by an improvement in the recollected impressions of relevant experiences and is thought to arouse positive feelings. We therefore hypothesized that the recognition of active listening activates the reward system, and that the emotional appraisal of experiences that had been subject to active listening would be improved. To test these hypotheses, we conducted functional magnetic resonance imaging (fMRI) on participants viewing assessments of their own personal experiences made by evaluators with or without active listening attitude. Subjects rated evaluators who showed active listening more positively. Furthermore, they rated episodes more positively when they were evaluated by individuals showing active listening. Neural activation in the ventral striatum was enhanced by perceiving active listening, suggesting that this was processed as rewarding. It also activated the right anterior insula, representing positive emotional reappraisal processes. Furthermore, the mentalizing network was activated when participants were being evaluated, irrespective of active listening behavior. Therefore, perceiving active listening appeared to result in positive emotional appraisal and to invoke mental state attribution to the active listener.

Perceiving active listening activates the reward system and improves the impression of relevant experiences

PubMed Central

Kawamichi, Hiroaki; Yoshihara, Kazufumi; Sasaki, Akihiro T.; Sugawara, Sho K.; Tanabe, Hiroki C.; Shinohara, Ryoji; Sugisawa, Yuka; Tokutake, Kentaro; Mochizuki, Yukiko; Anme, Tokie; Sadato, Norihiro

2015-01-01

Although active listening is an influential behavior, which can affect the social responses of others, the neural correlates underlying its perception have remained unclear. Sensing active listening in social interactions is accompanied by an improvement in the recollected impressions of relevant experiences and is thought to arouse positive feelings. We therefore hypothesized that the recognition of active listening activates the reward system, and that the emotional appraisal of experiences that had been subject to active listening would be improved. To test these hypotheses, we conducted functional magnetic resonance imaging (fMRI) on participants viewing assessments of their own personal experiences made by evaluators with or without active listening attitude. Subjects rated evaluators who showed active listening more positively. Furthermore, they rated episodes more positively when they were evaluated by individuals showing active listening. Neural activation in the ventral striatum was enhanced by perceiving active listening, suggesting that this was processed as rewarding. It also activated the right anterior insula, representing positive emotional reappraisal processes. Furthermore, the mentalizing network was activated when participants were being evaluated, irrespective of active listening behavior. Therefore, perceiving active listening appeared to result in positive emotional appraisal and to invoke mental state attribution to the active listener. PMID:25188354

101 Environmental Education Activities. Booklet 5--Science & Social Studies (Interdisciplinary) Activities.

ERIC Educational Resources Information Center

Whitney, Helen, Comp.

Forestry is the main focus of this fifth booklet in the series "101 Environmental Education Activities" by the Upper Mississippi River ECO-Center. Designed for students in the intermediate grades and junior high school, the booklet contains 9 science and social studies activities and 5 interdisciplinary activities. Most activity…

Cultural Activation of Consumers.

PubMed

Siegel, Carole E; Reid-Rose, Lenora; Joseph, Adriana M; Hernandez, Jennifer C; Haugland, Gary

2016-02-01

This column discusses "cultural activation," defined as a consumer's recognition of the importance of providing cultural information to providers about cultural affiliations, challenges, views about, and attitudes toward behavioral health and general medical health care, as well as the consumer's confidence in his or her ability to provide this information. An aid to activation, "Cultural Activation Prompts," and a scale that measures a consumer's level of activation, the Cultural Activation Measurement Scale, are described. Suggestions are made about ways to introduce cultural activation as a component of usual care.

TRAF6 and Src kinase activity regulates Cot activation by IL-1.

PubMed

Rodríguez, Cristina; Pozo, Maite; Nieto, Elvira; Fernández, Margarita; Alemany, Susana

2006-09-01

Cot is one of the MAP kinase kinase kinases that regulates the ERK1/ERK2 pathway under physiological conditions. Cot is activated by LPS, by inducing its dissociation from the inactive p105 NFkappaB-Cot complex in macrophages. Here, we show that IL-1 promotes a 10-fold increase in endogenous Cot activity and that Cot is the only MAP kinase kinase kinase that activates ERK1/ERK2 in response to this cytokine. Moreover, in cells where the expression of Cot is blocked, IL-1 fails to induce an increase in IL-8 and MIP-1betamRNA levels. The activation of Cot-MKK1-ERK1/ERK2 signalling pathway by IL-1 is dependent on the activity of the transducer protein TRAF6. Most important, IL-1-induced ERK1/ERK2 activation is inhibited by PP1, a known inhibitor of Src tyrosine kinases, but this tyrosine kinase activity is not required for IL-1 to activate other MAP kinases such as p38 and JNK. This Src kinases inhibitor does not block the dissociation and subsequently degradation of Cot in response to IL-1, indicating that other events besides Cot dissociation are required to activate Cot. All these data highlight the specific requirements for activation of the Cot-MKK1-ERK1/ERK2 pathway and provide evidence that Cot controls the functions of IL-1 that are mediated by ERK1/ERK2.

Teacher Activity Survey.

ERIC Educational Resources Information Center

Delaware Univ., Newark. School of Education.

The purpose of this survey is to determine what teachers think are (a) the relevant teaching activities, skills, or procedures and (b) the appropriate time and place to develop competence in an activity. Multiple choice responses are given for each of the following teacher activities: (a) utilize more than one approach to controlling and…

Activity and energy expenditure in older people playing active video games.

PubMed

Taylor, Lynne M; Maddison, Ralph; Pfaeffli, Leila A; Rawstorn, Jonathan C; Gant, Nicholas; Kerse, Ngaire M

2012-12-01

Tayl To quantify energy expenditure in older adults playing interactive video games while standing and seated, and secondarily to determine whether participants' balance status influenced the energy cost associated with active video game play. Cross-sectional study. University research center. Community-dwelling adults (N=19) aged 70.7±6.4 years. Participants played 9 active video games, each for 5 minutes, in random order. Two games (boxing and bowling) were played in both seated and standing positions. Energy expenditure was assessed using indirect calorimetry while at rest and during game play. Energy expenditure was expressed in kilojoules per minute and metabolic equivalents (METs). Balance was assessed using the mini-BESTest, the Activities-specific Balance Confidence Scale, and the Timed Up and Go (TUG). Mean ± SD energy expenditure was significantly greater for all game conditions compared with rest (all P≤.01) and ranged from 1.46±.41 METs to 2.97±1.16 METs. There was no significant difference in energy expenditure, activity counts, or perceived exertion between equivalent games played while standing and seated. No significant correlations were observed between energy expenditure or activity counts and balance status. Active video games provide light-intensity exercise in community-dwelling older people, whether played while seated or standing. People who are unable to stand may derive equivalent benefits from active video games played while seated. Further research is required to determine whether sustained use of active video games alters physical activity levels in community settings for this population. Copyright © 2012 American Congress of Rehabilitation Medicine. Published by Elsevier Inc. All rights reserved.

Obesity and lipid stress inhibit carnitine acetyltransferase activity[S

PubMed Central

Seiler, Sarah E.; Martin, Ola J.; Noland, Robert C.; Slentz, Dorothy H.; DeBalsi, Karen L.; Ilkayeva, Olga R.; An, Jie; Newgard, Christopher B.; Koves, Timothy R.; Muoio, Deborah M.

2014-01-01

Carnitine acetyltransferase (CrAT) is a mitochondrial matrix enzyme that catalyzes the interconversion of acetyl-CoA and acetylcarnitine. Emerging evidence suggests that this enzyme functions as a positive regulator of total body glucose tolerance and muscle activity of pyruvate dehydrogenase (PDH), a mitochondrial enzyme complex that promotes glucose oxidation and is feedback inhibited by acetyl-CoA. Here, we used tandem mass spectrometry-based metabolic profiling to identify a negative relationship between CrAT activity and muscle content of lipid intermediates. CrAT specific activity was diminished in muscles from obese and diabetic rodents despite increased protein abundance. This reduction in enzyme activity was accompanied by muscle accumulation of long-chain acylcarnitines (LCACs) and acyl-CoAs and a decline in the acetylcarnitine/acetyl-CoA ratio. In vitro assays demonstrated that palmitoyl-CoA acts as a direct mixed-model inhibitor of CrAT. Similarly, in primary human myocytes grown in culture, nutritional and genetic manipulations that promoted mitochondrial influx of fatty acids resulted in accumulation of LCACs but a pronounced decrease of CrAT-derived short-chain acylcarnitines. These results suggest that lipid-induced antagonism of CrAT might contribute to decreased PDH activity and glucose disposal in the context of obesity and diabetes. PMID:24395925

Choleretic Activity of Turmeric and its Active Ingredients.

PubMed

Wang, Yonglu; Wang, Liyao; Zhu, Xinyi; Wang, Dong; Li, Xueming

2016-07-01

Turmeric, a rhizome of Curcumin longa L. is widely used as both a spice and an herbal medicine. The traditional use of turmeric in gastroenterology is mainly based on its choleretic activity. The aim of this study is to determine the effects of turmeric on bile flow (BF) and total bile acids (TBAs) excretion in a bile fistula rat model after acute duodenal administration. A significant dose-dependent enhancement in both BF and TBAs was detected after treatment with the turmeric decoctions which suggested the choleretic activity was bile acid-dependent secretion. In order to direct the active group of compounds, aqueous (AE), ethyl acetate (EtOAc), and petroleum ether (PE) extracts were investigated. The EtOAc and PE extracts showing high effects were purified to locate the active ingredients. Three curcuminoids (curcumin, demethoxycurcumin, and bisdemethoxycurcumin) and 2 sesquiterpenes (bisacurone B and ar-turmerone) were isolated. It was found Bisacurone B was the most potent choleretic ingredient followed by ar-turmerone, bisdemethoxycurcumin demethoxycurcumin, and then curcumin. The amounts of the active ingredients were quantitatively analyzed by high-performance liquid chromatography. The EtOAc and PE extracts had high sesquiterpenes and curcuminoids content, while the AE extract had poor content of sesquiterpenes and curcuminoids which affected neither BF nor TBAs. Based on the results of multiple linear regression analysis, the content of BIS and TUR were dominant factors (P < 0.01) of controlling BL and TBAs in EtOAC and PE extracts. © 2016 Institute of Food Technologists®

Active and non-active video gaming among Dutch adolescents: who plays and how much?

PubMed

Simons, Monique; de Vet, Emely; Brug, Johannes; Seidell, Jaap; Chinapaw, Mai J M

2014-11-01

The aim of study was to determine prevalence and identify demographic correlates of active and non-active gaming among adolescents. Cross-sectional. A survey, assessing game behavior and correlates, was conducted among adolescents (12-16 years, n = 373), recruited via schools. Multivariable logistic regression analyses were conducted to examine demographic correlates of active gaming (≥ 1 h per week) and non-active gaming (>7h per week). Of all participants (n=373), 3% reported to play exclusively active games, 40% active games and non-active games, 40% exclusively non-active games, and 17% not playing video games at all. Active gaming adolescents played active games on average on 1.5 (sd = 1.2) days per school week for 36 (sd = 32.9)min and 1 (sd = 0.54) day per weekend for 42 (sd = 36.5)min. Non-active gaming adolescents played on average on 3.3 (sd = 1.6) days per school week for 65 (sd = 46.0)min and 1.4 (sd = 0.65) days per weekend for 80 (sd = 50.8)min. Adolescents attending lower levels of education were more likely to play active games ≥ 1 h per week than adolescents attending higher educational levels. Boys and older adolescents were more likely to play non-active games >7h per week, than girls or younger adolescents. Many adolescents play active games, especially those following a lower educational level, but time spent in this activity is relatively low compared to non-active gaming. To be feasible as a public health strategy, active gaming interventions should achieve more time is spent on active gaming at the expense of non-active gaming. Copyright © 2013 Sports Medicine Australia. Published by Elsevier Ltd. All rights reserved.

Physical Activity Pattern of Malaysian Preschoolers: Environment, Barriers, and Motivators for Active Play.

PubMed

Lee, Shoo Thien; Wong, Jyh Eiin; Ong, Wei Wen; Ismail, Mohd Noor; Deurenberg, Paul; Poh, Bee Koon

2016-07-01

Children's physical activity has been correlated with child characteristics and social or physical environment. This study aimed to compare preschoolers' physical activity among various sociodemographic characteristics and to determine barriers, motivators, and environmental factors for active play. A total of 835 preschoolers were included in this analysis. Time spent on active play, quiet play, and screen time was reported by parents. Boys spent significantly more time on active play and screen time than girls. Time spent on quiet play was highest in East Coast Peninsular Malaysia and lowest in Sarawak. Some 40% of children achieved active play recommendation while 27% exceeded daily screen time recommendation. Most parents reported that their child played actively in the house area; and that the main barrier and motivator to active play were safety and child's enjoyment, respectively. These findings demonstrate that sociodemographic characteristics and environment should be considered in designing physical activity intervention programs. © 2016 APJPH.

Outdoor Activities.

ERIC Educational Resources Information Center

Minneapolis Independent School District 275, Minn.

Twenty-four activities suitable for outdoor use by elementary school children are outlined. Activities designed to make children aware of their environment include soil painting, burr collecting, insect and pond water collecting, studies of insect galls and field mice, succession studies, and a model of natural selection using dyed toothpicks. A…

Astronomy Activities.

ERIC Educational Resources Information Center

Greenstone, Sid

This document consists of activities and references for teaching astronomy. The activities (which include objectives, list of materials needed, and procedures) focus on: observing the Big Dipper and locating the North Star; examining the Big Dipper's stars; making and using an astrolabe; examining retograde motion of Mars; measuring the Sun's…

Motives for Physical Activity among Active and Inactive Persons in Their Mid-Thirties

PubMed Central

Aaltonen, Sari; Rottensteiner, Mirva; Kaprio, Jaakko; Kujala, Urho M

2013-01-01

The purpose of this study was to examine the motives for leisure time physical activity among active and inactive men and women in their mid-thirties. We used both cross-sectional and longitudinal designs. Altogether, 2308 participants (mean age 33.9 years, 53.4 % women) were identified from the population-based FinnTwin16 Cohort. Physically active and inactive individuals were identified on the basis of their leisure time MET hours/day. We evaluated participants’ physical activity motivation with a modified version of the Recreational Exercise Motivation Measure. Comparisons between active and inactive individuals were analysed using the Wald test for equality of means, and effect sizes were calculated as Cohen’s d. Motives related to mastery, physical fitness, social aspect of physical activity, psychological state, enjoyment, willingness to be fitter/look better than others and appearance were significantly more important for the active than inactive participants. Conforming to others’ expectations was the only item on which the inactive persons scored higher than active persons. The longitudinal results for physical activity were parallel to the cross-sectional results. This study supports to the view that motivation factors differ between active and inactive persons, and that intrinsic motives are associated with consistent leisure time physical activity. PMID:23331765

Contact activation: a revision.

PubMed

Schmaier, A H

1997-07-01

In conclusion, a revised view of the contact system has been presented. This system has little to do with the initiation of hemostasis. Like lupus anticoagulants, deficiencies of contact proteins give prolonged APTTs but may be risk factors for thrombosis. BK from kininogens is a potent modulator of vascular biology inducing vasodilation, tissue plasminogen activator release, and prostacyclin liberation. Kininogens, themselves, are selective inhibitors of alpha-thrombin-induced platelet activation preventing alpha-thrombin from cleaving the cloned thrombin receptor after arginine41. Kininogens' alpha-thrombin inhibitory activity exists in intact kininogens, BK, and all of BK's breakdown products. HK also is the pivotal protein for contact protein assembly on endothelium. It is the receptor for prekallikrein which when bound to HK becomes activated to kallikrein by an endothelial cell enzyme system independent of activated forms of plasma factor XII. Prekallikrein activation on endothelial cells results in kinetically favorable single chain urokinase and plasminogen activation. Thus the "physiologic, negatively charged surface" for contact system activation is really the assembly of these proteins on cell membranes and activation by membrane-associated enzymes.

Active site densities, oxygen activation and adsorbed reactive oxygen in alcohol activation on npAu catalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Lu-Cun; Friend, C. M.; Fushimi, Rebecca

The activation of molecular O 2as well as the reactivity of adsorbed oxygen species is of central importance in aerobic selective oxidation chemistry on Au-based catalysts. Herein, we address the issue of O 2activation on unsupported nanoporous gold (npAu) catalysts by applying a transient pressure technique, a temporal analysis of products (TAP) reactor, to measure the saturation coverage of atomic oxygen, its collisional dissociation probability, the activation barrier for O 2dissociation, and the facility with which adsorbed O species activate methanol, the initial step in the catalytic cycle of esterification. The results from these experiments indicate that molecular O 2dissociationmore » is associated with surface silver, that the density of reactive sites is quite low, that adsorbed oxygen atoms do not spill over from the sites of activation onto the surrounding surface, and that methanol reacts quite facilely with the adsorbed oxygen atoms. In addition, the O species from O 2dissociation exhibits reactivity for the selective oxidation of methanol but not for CO. The TAP experiments also revealed that the surface of the npAu catalyst is saturated with adsorbed O under steady state reaction conditions, at least for the pulse reaction.« less

Active site densities, oxygen activation and adsorbed reactive oxygen in alcohol activation on npAu catalysts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Lu-Cun; Friend, C. M.; Fushimi, Rebecca

2016-01-01

The activation of molecular O 2as well as the reactivity of adsorbed oxygen species is of central importance in aerobic selective oxidation chemistry on Au-based catalysts. Herein, we address the issue of O 2activation on unsupported nanoporous gold (npAu) catalysts by applying a transient pressure technique, a temporal analysis of products (TAP) reactor, to measure the saturation coverage of atomic oxygen, its collisional dissociation probability, the activation barrier for O 2dissociation, and the facility with which adsorbed O species activate methanol, the initial step in the catalytic cycle of esterification. The results from these experiments indicate that molecular O 2dissociationmore » is associated with surface silver, that the density of reactive sites is quite low, that adsorbed oxygen atoms do not spill over from the sites of activation onto the surrounding surface, and that methanol reacts quite facilely with the adsorbed oxygen atoms. In addition, the O species from O 2dissociation exhibits reactivity for the selective oxidation of methanol but not for CO. The TAP experiments also revealed that the surface of the npAu catalyst is saturated with adsorbed O under steady state reaction conditions, at least for the pulse reaction.« less

Activation of human peroxisome-activated receptor-gamma ...

EPA Pesticide Factsheets

Obesity in children has become an epidemic and recent research suggests a possible contribution from exposure to environmental chemicals. Several chemicals, such as phthalates, brominated flame retardants, and perfluorinated chemicals, are common in house dust on floors where children play and are suspected obesogens. Obesogens can act via a mechanism that involves activation of peroxisome proliferator-activated receptor-gamma (PPARy). A previous study found that dust collected from children’s homes binds to PPARy. Here, we investigated the ability of house dust to activate PPARy in a transiently transfected cell assay. Dust samples were collected in 2012 from carpeted and hardwood floors in children’s homes using thimbles fitted into a vacuum cleaner hose (“TEO” samples), or from homes in an adult cohort NIEHS study. Dust was extracted with 50:50 hexane:acetone, sonicated, centrifuged, and the organic layer collected. This was repeated 2X. The extracts were filtered to remove particulates, dried with purified nitrogen, and reconstituted in DMS0 at 200 ug/ul. COS-1 cells were transfected for 24 hrs with a human PPARy vector containing a luciferase reporter, and exposed for 24 hrs to negative controls water or DMSO (0.1%), positive controls Troglitazone (3 uM in water) or Rosiglitazone (100 nM in DMSO), or dust extracts serially diluted in DMEM at 50, 100, and 200 ug/ml in 0.1% DMSO. Cells were lysed and luciferase activity was measured. Data were log-tra

A Period-Activity Relation for Active RS CVN Stars

NASA Astrophysics Data System (ADS)

Simon, Theodore

Soft X ray observations of RS CVn binaries point to a correlation between L x /Lbol (the X ray to bolometric luminosity ratio that measures the coronal heating rate) and Omega (the stellar angular velocity). This correlation is almost certainly caused by a stellar dynamo, operating in rapidly-rotating late-type stars with deep convection zones. We are proposing to extend the X ray "rotation-activity relation" to the uv transition region and chromospheric emission lines observable with IUE. If the non-radiative heating rates of stellar transition regions and chromospheres are determined largely by magnetic processes associated with a stellar dynamo, then a similar correlation may be found. We have selected a group of recently discovered active long-period systems, which we believe will be very bright at uv wavelengths. One important goal of this program is to determine whether past studies of the "rotation-activity connection" have been compromised by the omission of active long-period RS CVn systems.

Active colloidal molecules

NASA Astrophysics Data System (ADS)

Löwen, Hartmut

2018-03-01

Like ordinary molecules are composed of atoms, colloidal molecules consist of several species of colloidal particles tightly bound together. If one of these components is self-propelled or swimming, novel “active colloidal molecules” emerge. Active colloidal molecules exist on various levels such as “homonuclear”, “heteronuclear” and “polymeric” and possess a dynamical function moving as propellers, spinners or rotors. Self-assembly of such active complexes has been studied a lot recently and this perspective article summarizes recent progress and gives an outlook to future developments in the rapidly expanding field of active colloidal molecules.

Cognitive Activity Mediates the Association between Social Activity and Cognitive Performance: A Longitudinal Study

PubMed Central

Brown, Cassandra L.; Robitaille, Annie; Zelinski, Elizabeth M.; Dixon, Roger A.; Hofer, Scott M.; Piccinin, Andrea M.

2016-01-01

Social activity is one aspect of an active lifestyle and some evidence indicates it is related to preserved cognitive function in older adulthood. However, the potential mechanisms underlying this association remain unclear. We investigate four potential mediational pathways through which social activity may relate to cognitive performance. A multilevel structural equation modeling approach to mediation was used to investigate whether cognitive activity, physical activity, depressive symptoms, and vascular health conditions mediate the association between social activity and cognitive function in older adults. Using data from the Victoria Longitudinal Study (VLS), we tested four cognitive outcomes: fluency, episodic memory, reasoning, and vocabulary. Three important findings emerged. First, the association between social activity and all four domains of cognitive function was significantly mediated by cognitive activity at the within-person level. Second, we observed a significant indirect effect of social activity on all domains of cognitive function through cognitive activity at the between-person level. Third, we found a within-person indirect relationship of social activity with episodic memory performance through physical activity. For these older adults, engagement in social activities was related to participation in everyday cognitive activities and in turn to better cognitive performance. This pattern is consistent with the interpretation that a lifestyle of social engagement may benefit cognitive performance by providing opportunities or motivation to participate in supportive cognitively stimulating activities. PMID:27929339

Weekly active-learning activities in a drug information and literature evaluation course.

PubMed

Timpe, Erin M; Motl, Susannah E; Eichner, Samantha F

2006-06-15

To incorporate learning activities into the weekly 2-hour Drug Information and Literature Evaluation class sessions to improve student ability and confidence in performing course objectives, as well as to assess student perception of the value of these activities. In-class activities that emphasized content and skills taught within class periods were created and implemented. Three different surveys assessing student ability and confidence in completing drug information and literature retrieval and evaluation tasks were administered prior to and following the appropriate class sessions. At the completion of the course, an additional evaluation was administered to assess the students' impressions of the value of the learning activities. Students reported increased ability and confidence in all course objectives. The teaching activities were also stated to be useful in students' learning of the material. Incorporation of weekly learning activities resulted in an improvement in student ability and confidence to perform course objectives. Students considered these activities to be beneficial and to contribute to the completion of course objectives.

Berberine Promotes Glucose Consumption Independently of AMP-Activated Protein Kinase Activation

PubMed Central

Xiao, Yuanyuan; Hou, Wolin; Yu, Xueying; Shen, Li; Liu, Fang; Wei, Li; Jia, Weiping

2014-01-01

Berberine is a plant alkaloid with anti-diabetic action. Activation of AMP-activated protein kinase (AMPK) pathway has been proposed as mechanism for berberine’s action. This study aimed to examine whether AMPK activation was necessary for berberine’s glucose-lowering effect. We found that in HepG2 hepatocytes and C2C12 myotubes, berberine significantly increased glucose consumption and lactate release in a dose-dependent manner. AMPK and acetyl coenzyme A synthetase (ACC) phosphorylation were stimulated by 20 µmol/L berberine. Nevertheless, berberine was still effective on stimulating glucose utilization and lactate production, when the AMPK activation was blocked by (1) inhibition of AMPK activity by Compound C, (2) suppression of AMPKα expression by siRNA, and (3) blockade of AMPK pathway by adenoviruses containing dominant-negative forms of AMPKα1/α2. To test the effect of berberine on oxygen consumption, extracellular flux analysis was performed in Seahorse XF24 analyzer. The activity of respiratory chain complex I was almost fully blocked in C2C12 myotubes by berberine. Metformin, as a positive control, showed similar effects as berberine. These results suggest that berberine and metformin promote glucose metabolism by stimulating glycolysis, which probably results from inhibition of mitochondrial respiratory chain complex I, independent of AMPK activation. PMID:25072399

Zinc triggers microglial activation.

PubMed

Kauppinen, Tiina M; Higashi, Youichirou; Suh, Sang Won; Escartin, Carole; Nagasawa, Kazuki; Swanson, Raymond A

2008-05-28

Microglia are resident immune cells of the CNS. When stimulated by infection, tissue injury, or other signals, microglia assume an activated, "ameboid" morphology and release matrix metalloproteinases, reactive oxygen species, and other proinflammatory factors. This innate immune response augments host defenses, but it can also contribute to neuronal death. Zinc is released by neurons under several conditions in which microglial activation occurs, and zinc chelators can reduce neuronal death in animal models of cerebral ischemia and neurodegenerative disorders. Here, we show that zinc directly triggers microglial activation. Microglia transfected with a nuclear factor-kappaB (NF-kappaB) reporter gene showed a severalfold increase in NF-kappaB activity in response to 30 microm zinc. Cultured mouse microglia exposed to 15-30 microm zinc increased nitric oxide production, increased F4/80 expression, altered cytokine expression, and assumed the activated morphology. Zinc-induced microglial activation was blocked by inhibiting NADPH oxidase, poly(ADP-ribose) polymerase-1 (PARP-1), or NF-kappaB activation. Zinc injected directly into mouse brain induced microglial activation in wild-type mice, but not in mice genetically lacking PARP-1 or NADPH oxidase activity. Endogenous zinc release, induced by cerebral ischemia-reperfusion, likewise induced a robust microglial reaction, and this reaction was suppressed by the zinc chelator CaEDTA. Together, these results suggest that extracellular zinc triggers microglial activation through the sequential activation of NADPH oxidase, PARP-1, and NF-kappaB. These findings identify a novel trigger for microglial activation and a previously unrecognized mechanism by which zinc may contribute to neurological disorders.

The Structure-Activity Relationship between Marine Algae Polysaccharides and Anti-Complement Activity

PubMed Central

Jin, Weihua; Zhang, Wenjing; Liang, Hongze; Zhang, Quanbin

2015-01-01

In this study, 33 different polysaccharides were prepared to investigate the structure-activity relationships between the polysaccharides, mainly from marine algae, and anti-complement activity in the classical pathway. Factors considered included extraction methods, fractionations, molecular weight, molar ratio of galactose to fucose, sulfate, uronic acid (UA) content, linkage, branching, and the type of monosaccharide. It was shown that the larger the molecular weights, the better the activities. The molar ratio of galactose (Gal) to fucose (Fuc) was a positive factor at a concentration lower than 10 µg/mL, while it had no effect at a concentration more than 10 µg/mL. In addition, sulfate was necessary; however, the sulfate content, the sulfate pattern, linkage and branching had no effect at a concentration of more than 10 µg/mL. Moreover, the type of monosaccharide had no effect. Laminaran and UA fractions had no activity; however, they could reduce the activity by decreasing the effective concentration of the active composition when they were mixed with the active compositions. The effect of the extraction methods could not be determined. Finally, it was observed that sulfated galactofucan showed good anti-complement activity after separation. PMID:26712768

Antifeedant activity of quassinoids.

PubMed

Leskinen, V; Polonsky, J; Bhatnagar, S

1984-10-01

The antifeedant activity of 13 quassinoids of different structural types has been studied against the Mexican bean beetle (Epilachna varivestis Mulsant) 4th instar larvae and the southern armyworm (Spodoptera eridania Crawer) 5th instar larvae. All quassinoids tested displayed significant activity against the Mexican bean beetle and, thus, do not reveal a simple structure-activity relationship. Five quassinoids were active against the southern armyworm. Interestingly, four of these-bruceantin (I), glaucarubinone (VI), isobruceine A (VIII), and simalikalactone D (XI)-possess the required structural features for antineoplastic activity. The noncytotoxic quassin (X) is an exception; it is active against both pests.

Occurrence and activity of Archaea in aerated activated sludge wastewater treatment plants.

PubMed

Gray, Neil D; Miskin, Ian P; Kornilova, Oksana; Curtis, Thomas P; Head, Ian M

2002-03-01

The occurrence, distribution and activity of archaeal populations within two aerated, activated sludge wastewater treatment systems, one treating domestic waste and the second treating mixed domestic and industrial wastewater, were investigated by denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction (PCR)-amplified ribosomal RNA gene fragments and process measurements. In the plant receiving mixed industrial and domestic waste the archaeal populations found in the mixed liquor were very similar to those in the influent sewage, though a small number of DGGE bands specific to the mixed liquor were identified. In contrast, the activated sludge treating principally domestic waste harboured distinct archaeal populations associated with the mixed liquor that were not prevalent in the influent sewage. We deduce that the Archaea in the plant treating mixed wastewater were derived principally from the influent, whereas those in the plant treating solely domestic waste were actively growing in the treatment plant. Archaeal 16S rRNA gene sequences related to the Methanosarcinales, Methanomicrobiales and the Methanobacteriales were detected. Methanogenesis was measured in activated sludge samples incubated under oxic and anoxic conditions, demonstrating that the methanogens present in both activated sludge plants were active only in anoxic incubations. The relatively low rates of methanogenesis measured indicated that, although active, the methanogens play a minor role in carbon turnover in activated sludge.

Meditation leads to reduced default mode network activity beyond an active task

PubMed Central

Garrison, Kathleen A.; Zeffiro, Thomas A.; Scheinost, Dustin; Constable, R. Todd; Brewer, Judson A.

2015-01-01

Meditation has been associated with relatively reduced activity in the default mode network, a brain network implicated in self-related thinking and mind wandering. However, previous imaging studies have typically compared meditation to rest despite other studies reporting differences in brain activation patterns between meditators and controls at rest. Moreover, rest is associated with a range of brain activation patterns across individuals that has only recently begun to be better characterized. Therefore, this study compared meditation to another active cognitive task, both to replicate findings that meditation is associated with relatively reduced default mode network activity, and to extend these findings by testing whether default mode activity was reduced during meditation beyond the typical reductions observed during effortful tasks. In addition, prior studies have used small groups, whereas the current study tested these hypotheses in a larger group. Results indicate that meditation is associated with reduced activations in the default mode network relative to an active task in meditators compared to controls. Regions of the default mode showing a group by task interaction include the posterior cingulate/precuneus and anterior cingulate cortex. These findings replicate and extend prior work indicating that suppression of default mode processing may represent a central neural process in long-term meditation, and suggest that meditation leads to relatively reduced default mode processing beyond that observed during another active cognitive task. PMID:25904238

Validity of physical activity monitors for assessing lower intensity activity in adults.

PubMed

Calabró, M Andrés; Lee, Jung-Min; Saint-Maurice, Pedro F; Yoo, Hyelim; Welk, Gregory J

2014-09-28

Accelerometers can provide accurate estimates of moderate-to-vigorous physical activity (MVPA). However, one of the limitations of these instruments is the inability to capture light activity within an acceptable range of error. The purpose of the present study was to determine the validity of different activity monitors for estimating energy expenditure (EE) of light intensity, semi-structured activities. Forty healthy participants wore a SenseWear Pro3 Armband (SWA, v.6.1), the SenseWear Mini, the Actiheart, ActiGraph, and ActivPAL monitors, while being monitored with a portable indirect calorimetry (IC). Participants engaged in a variety of low intensity activities but no formalized scripts or protocols were used during these periods. The Mini and SWA overestimated total EE on average by 1.0% and 4.0%, respectively, while the AH, the GT3X, and the AP underestimated total EE on average by 7.8%, 25.5%, and 22.2%, respectively. The pattern-recognition monitors yielded non-significant differences in EE estimates during the semi-structured period (p = 0.66, p = 0.27, and p = 0.21 for the Mini, SWA, and AH, respectively). The SenseWear Mini provided more accurate estimates of EE during light to moderate intensity semi-structured activities compared to other activity monitors. This monitor should be considered when there is interest in tracking low intensity activities in groups of individuals.

Polypeptides having beta-glucosidase activity, beta-xylosidase activity, or beta-glucosidase and beta-xylosidase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morant, Marc

The present invention relates to isolated polypeptides having beta-glucosidase activity, beta-xylosidase activity, or beta-glucosidase and beta-xylosidase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Misclassification of Physical Activity Level Due to Exclusion of Workplace Activity

ERIC Educational Resources Information Center

Boslaugh, Sarah E.; Kreuter, Matthew W.; Weaver, Nancy L.; Naleid, Kimberly S.; Brownson, Ross C.

2005-01-01

This study examined the effect of including workplace physical activity in calculating the proportion of adults meeting Centers for Disease Control (CDC) guidelines for physical activity. Data on leisure-time and workplace activity were collected from 1,090 Black and White adults in St. Louis, MO. A series of assumptions were used to equate…

Active Learning Increases Children's Physical Activity across Demographic Subgroups.

PubMed

Bartholomew, John B; Jowers, Esbelle M; Roberts, Gregory; Fall, Anna-Mária; Errisuriz, Vanessa L; Vaughn, Sharon

2018-01-01

Given the need to find more opportunities for physical activity within the elementary school day, this study was designed to asses the impact of I-CAN!, active lessons on: 1) student physical activity (PA) outcomes via accelerometry; and 2) socioeconomic status (SES), race, sex, body mass index (BMI), or fitness as moderators of this impact. Participants were 2,493 fourth grade students (45.9% male, 45.8% white, 21.7% low SES) from 28 central Texas elementary schools randomly assigned to intervention (n=19) or control (n=9). Multilevel regression models evaluated the effect of I-CAN! on PA and effect sizes were calculated. The moderating effects of SES, race, sex, BMI, and fitness were examined in separate models. Students in treatment schools took significantly more steps than those in control schools (β = 125.267, SE = 41.327, p = .002, d = .44). I-CAN! had a significant effect on MVPA with treatment schools realizing 80% (β = 0.796, SE =0.251, p = .001; d = .38) more MVPA than the control schools. There were no significant school-level differences on sedentary behavior (β = -0.177, SE = 0.824, p = .83). SES, race, sex, BMI, and fitness level did not moderate the impact of active learning on step count and MVPA. Active learning increases PA within elementary students, and does so consistently across demographic sub-groups. This is important as these sub-groups represent harder to reach populations for PA interventions. While these lessons may not be enough to help children reach daily recommendations of PA, they can supplement other opportunities for PA. This speaks to the potential of schools to adopt policy change to require active learning.

National physical activity surveillance: Users of wearable activity monitors as a potential data source.

PubMed

Omura, John D; Carlson, Susan A; Paul, Prabasaj; Watson, Kathleen B; Fulton, Janet E

2017-03-01

The objective of this study was to assess usage patterns of wearable activity monitors among US adults and how user characteristics might influence physical activity estimates from this type of sample. We analyzed data on 3367 respondents to the 2015 HealthStyles survey, an annual consumer mail panel survey conducted on a nationwide sample. Approximately 1 in 8 respondents (12.5%) reported currently using a wearable activity monitor. Current use varied by sex, age, and education level. Use increased with physical activity level from 4.3% for inactive adults to 17.4% for active adults. Overall, 49.9% of all adults met the aerobic physical activity guideline, while this prevalence was 69.5% among current activity monitor users. Our findings suggest that current users of wearable activity monitors are not representative of the overall US population. Estimates of physical activity levels using data from wearable activity monitors users may be an overestimate and therefore data from users alone may have a limited role in physical activity surveillance.

Cognitive activity mediates the association between social activity and cognitive performance: A longitudinal study.

PubMed

Brown, Cassandra L; Robitaille, Annie; Zelinski, Elizabeth M; Dixon, Roger A; Hofer, Scott M; Piccinin, Andrea M

2016-12-01

Social activity is 1 aspect of an active lifestyle and some evidence indicates it is related to preserved cognitive function in older adulthood. However, the potential mechanisms underlying this association remain unclear. We investigate 4 potential mediational pathways through which social activity may relate to cognitive performance. A multilevel structural equation modeling approach to mediation was used to investigate whether cognitive activity, physical activity, depressive symptoms, and vascular health conditions mediate the association between social activity and cognitive function in older adults. Using data from the Victoria Longitudinal Study, we tested 4 cognitive outcomes: fluency, episodic memory, reasoning, and vocabulary. Three important findings emerged. First, the association between social activity and all 4 domains of cognitive function was significantly mediated by cognitive activity at the within-person level. Second, we observed a significant indirect effect of social activity on all domains of cognitive function through cognitive activity at the between-person level. Third, we found a within-person indirect relationship of social activity with episodic memory performance through physical activity. For these older adults, engagement in social activities was related to participation in everyday cognitive activities and in turn to better cognitive performance. This pattern is consistent with the interpretation that a lifestyle of social engagement may benefit cognitive performance by providing opportunities or motivation to participate in supportive cognitively stimulating activities. (PsycINFO Database Record (c) 2016 APA, all rights reserved).

Activation of Extracellular Signal-Regulated Kinase but Not of p38 Mitogen-Activated Protein Kinase Pathways in Lymphocytes Requires Allosteric Activation of SOS

PubMed Central

Jun, Jesse E.; Yang, Ming; Chen, Hang; Chakraborty, Arup K.

2013-01-01

Thymocytes convert graded T cell receptor (TCR) signals into positive selection or deletion, and activation of extracellular signal-related kinase (ERK), p38, and Jun N-terminal protein kinase (JNK) mitogen-activated protein kinases (MAPKs) has been postulated to play a discriminatory role. Two families of Ras guanine nucleotide exchange factors (RasGEFs), SOS and RasGRP, activate Ras and the downstream RAF-MEK-ERK pathway. The pathways leading to lymphocyte p38 and JNK activation are less well defined. We previously described how RasGRP alone induces analog Ras-ERK activation while SOS and RasGRP cooperate to establish bimodal ERK activation. Here we employed computational modeling and biochemical experiments with model cell lines and thymocytes to show that TCR-induced ERK activation grows exponentially in thymocytes and that a W729E allosteric pocket mutant, SOS1, can only reconstitute analog ERK signaling. In agreement with RasGRP allosterically priming SOS, exponential ERK activation is severely decreased by pharmacological or genetic perturbation of the phospholipase Cγ (PLCγ)-diacylglycerol-RasGRP1 pathway. In contrast, p38 activation is not sharply thresholded and requires high-level TCR signal input. Rac and p38 activation depends on SOS1 expression but not allosteric activation. Based on computational predictions and experiments exploring whether SOS functions as a RacGEF or adaptor in Rac-p38 activation, we established that the presence of SOS1, but not its enzymatic activity, is critical for p38 activation. PMID:23589333

Optimizing Picture Activities for the Language Classroom: Picture Line-Up Activities.

ERIC Educational Resources Information Center

Schmidt, Ken

1995-01-01

The use of picture line-up activities, classroom exercises in which students must place a series of pictures in correct order, is discussed. The advantages of use of a picture series instead of simple pairs are examined, a basic activity is described, and ideas for implementation, adaptation, and follow-up are outlined. The basic activity uses…

Zinc triggers microglial activation

PubMed Central

Kauppinen, Tiina M.; Higashi, Youichirou; Suh, Sang Won; Escartin, Carole; Nagasawa, Kazuki; Swanson, Raymond A.

2009-01-01

Microglia are resident immune cells of the central nervous system. When stimulated by infection, tissue injury, or other signals, microglia assume an activated, “amoeboid” morphology and release matrix metalloproteinases, reactive oxygen species, and other pro-inflammatory factors. This innate immune response augments host defenses, but it can also contribute to neuronal death. Zinc is released by neurons under several conditions in which microglial activation occurs, and zinc chelators can reduce neuronal death in animal models of cerebral ischemia and neurodegenerative disorders. Here we show that zinc directly triggers microglial activation. Microglia transfected with an NF-kB reporter gene showed a several-fold increase in NF-kB activity in response to 30 μM zinc. Cultured mouse microglia exposed to 15 – 30 μM zinc increased nitric oxide production, increased F4/80 expression, altered cytokine expression, and assumed the activated morphology. Zinc-induced microglial activation was blocked by inhibiting NADPH oxidase, poly(ADP-ribose) polymerase-1 (PARP-1), or NF-κB activation. Zinc injected directly into mouse brain induced microglial activation in wild-type mice, but not in mice genetically lacking PARP-1 or NADPH oxidase activity. Endogenous zinc release, induced by cerebral ischemia-reperfusion, likewise induced a robust microglial reaction, and this reaction was suppressed by the zinc chelator CaEDTA. Together, these results suggest that extracellular zinc triggers microglial activation through the sequential activation of NADPH oxidase, PARP-1, and NF-κB. These findings identify a novel trigger for microglial activation and a previously unrecognized mechanism by which zinc may contribute to neurological disorders. PMID:18509044

Stellar activity with LAMOST - II. Chromospheric activity in open clusters

NASA Astrophysics Data System (ADS)

Fang, Xiang-Song; Zhao, Gang; Zhao, Jing-Kun; Bharat Kumar, Yerra

2018-05-01

We use the LAMOST spectra of member stars in Pleiades, M34, Praesepe, and Hyades to study how chromospheric activity varies as a function of mass and rotation at different age. We measured excess equivalent widths of H α, H β, and Ca II K based on estimated chromospheric contributions from old and inactive field dwarfs, and excess luminosities are obtained by normalizing bolometric luminosity, for more than 700 late-type stars in these open clusters. Results indicate two activity sequences in cool spot coverage and H α excess emission among GK dwarfs in Pleiades and M dwarfs in Praesepe and Hyades, paralleling with well-known rotation sequences. A weak dependence of chromospheric emission on rotation exists among ultrafast rotators in saturated regime with Rossby number Ro ≲ 0.1. In the unsaturated regime, chromospheric and coronal emission show similar dependence on Ro, but with a shift towards larger Ro, indicating chromospheric emission gets easily saturated than coronal emission, and/or convective turnover time-scales based on X-ray data do not work well with chromospheric emission. More interestingly, our analysis shows fully convective slow rotators obey the rotation-chromospheric activity relation similar to hotter stars, confirming the previous finding. We found correlations among H α, H β, and Ca II K emissions, in which H α losses are more important than Ca II K for cooler and more active stars. In addition, a weak correlation is seen between chromospheric emission and photospheric activity that shows dependence on stellar spectral type and activity level, which provides some clues on how spot configuration varies as a function of mass and activity level.

Are joint injury, sport activity, physical activity, obesity, or occupational activities predictors for osteoarthritis? A systematic review.

PubMed

Richmond, Sarah A; Fukuchi, Reginaldo K; Ezzat, Allison; Schneider, Kathryn; Schneider, Geoff; Emery, Carolyn A

2013-08-01

Systematic review with meta-analysis. To identify risk factors for osteoarthritis (OA) of the knee, hip, and ankle, including joint injury, sport, physical activity, overweight/obesity, and occupational activity, in all age groups. OA is a significant health problem worldwide, affecting up to 10% of men and 18% of women over 60 years of age. There has not been a comprehensive review examining modifiable physical risk factors associated with the onset of OA. This evidence is important to inform the physiotherapy management of individuals following onset of OA. Twelve electronic databases were systematically reviewed. The studies selected met the following criteria: (1) original data; (2) joint injury, sport activity, physical activity, overweight/obesity, and/or occupational activity investigated as risk factors; (3) outcomes included OA (hip, knee, and/or ankle); and (4) analytic component of study design. The data extracted included study design, years of follow-up, study population, OA definition, risk factors, and results (effect estimates reported or calculated where available). The quality of evidence was assessed based on a modified version of the Downs and Black checklist. Joint injury, obesity, and occupational activity were associated with an increased risk of OA of the knee and hip. Sport and physical activity produced inconsistent findings. Joint injury was identified as a significant risk factor for knee OA (combined odds ratio = 3.8; 95% confidence interval: 2.0, 7.2) and hip OA (combined odds ratio = 5.0; 95% confidence interval: 1.4, 18.2), as was previous meniscectomy with or without anterior cruciate ligament injury for knee OA (combined odds ratio = 7.4; 95% confidence interval: 4.0, 13.7). There is a paucity of research examining risk factors associated with ankle OA; this review identified only 2 studies with this outcome. Joint injury, obesity, and occupational activity are associated with an increased risk of knee and hip OA. Some findings

Gi proteins regulate adenylyl cyclase activity independent of receptor activation.

PubMed

Melsom, Caroline Bull; Ørstavik, Øivind; Osnes, Jan-Bjørn; Skomedal, Tor; Levy, Finn Olav; Krobert, Kurt Allen

2014-01-01

Despite the view that only β2- as opposed to β1-adrenoceptors (βARs) couple to G(i), some data indicate that the β1AR-evoked inotropic response is also influenced by the inhibition of Gi. Therefore, we wanted to determine if Gi exerts tonic receptor-independent inhibition upon basal adenylyl cyclase (AC) activity in cardiomyocytes. We used the Gs-selective (R,R)- and the Gs- and G(i)-activating (R,S)-fenoterol to selectively activate β2ARs (β1AR blockade present) in combination with Gi inactivation with pertussis toxin (PTX). We also determined the effect of PTX upon basal and forskolin-mediated responses. Contractility was measured ex vivo in left ventricular strips and cAMP accumulation was measured in isolated ventricular cardiomyocytes from adult Wistar rats. PTX amplified both the (R,R)- and (R,S)-fenoterol-evoked maximal inotropic response and concentration-dependent increases in cAMP accumulation. The EC50 values of fenoterol matched published binding affinities. The PTX enhancement of the Gs-selective (R,R)-fenoterol-mediated responses suggests that Gi regulates AC activity independent of receptor coupling to Gi protein. Consistent with this hypothesis, forskolin-evoked cAMP accumulation was increased and inotropic responses to forskolin were potentiated by PTX treatment. In non-PTX-treated tissue, phosphodiesterase (PDE) 3 and 4 inhibition or removal of either constitutive muscarinic receptor activation of Gi with atropine or removal of constitutive adenosine receptor activation with CGS 15943 had no effect upon contractility. However, in PTX-treated tissue, PDE3 and 4 inhibition alone increased basal levels of cAMP and accordingly evoked a large inotropic response. Together, these data indicate that Gi exerts intrinsic receptor-independent inhibitory activity upon AC. We propose that PTX treatment shifts the balance of intrinsic G(i) and Gs activity upon AC towards Gs, enhancing the effect of all cAMP-mediated inotropic agents.

Gi Proteins Regulate Adenylyl Cyclase Activity Independent of Receptor Activation

PubMed Central

Melsom, Caroline Bull; Ørstavik, Øivind; Osnes, Jan-Bjørn; Skomedal, Tor; Levy, Finn Olav; Krobert, Kurt Allen

2014-01-01

Background and purpose Despite the view that only β2- as opposed to β1-adrenoceptors (βARs) couple to Gi, some data indicate that the β1AR-evoked inotropic response is also influenced by the inhibition of Gi. Therefore, we wanted to determine if Gi exerts tonic receptor-independent inhibition upon basal adenylyl cyclase (AC) activity in cardiomyocytes. Experimental approach We used the Gs-selective (R,R)- and the Gs- and Gi-activating (R,S)-fenoterol to selectively activate β2ARs (β1AR blockade present) in combination with Gi inactivation with pertussis toxin (PTX). We also determined the effect of PTX upon basal and forskolin-mediated responses. Contractility was measured ex vivo in left ventricular strips and cAMP accumulation was measured in isolated ventricular cardiomyocytes from adult Wistar rats. Key results PTX amplified both the (R,R)- and (R,S)-fenoterol-evoked maximal inotropic response and concentration-dependent increases in cAMP accumulation. The EC50 values of fenoterol matched published binding affinities. The PTX enhancement of the Gs-selective (R,R)-fenoterol-mediated responses suggests that Gi regulates AC activity independent of receptor coupling to Gi protein. Consistent with this hypothesis, forskolin-evoked cAMP accumulation was increased and inotropic responses to forskolin were potentiated by PTX treatment. In non-PTX-treated tissue, phosphodiesterase (PDE) 3 and 4 inhibition or removal of either constitutive muscarinic receptor activation of Gi with atropine or removal of constitutive adenosine receptor activation with CGS 15943 had no effect upon contractility. However, in PTX-treated tissue, PDE3 and 4 inhibition alone increased basal levels of cAMP and accordingly evoked a large inotropic response. Conclusions and implications Together, these data indicate that Gi exerts intrinsic receptor-independent inhibitory activity upon AC. We propose that PTX treatment shifts the balance of intrinsic Gi and Gs activity upon AC towards Gs

Entropy production of active particles and for particles in active baths

NASA Astrophysics Data System (ADS)

Pietzonka, Patrick; Seifert, Udo

2018-01-01

Entropy production of an active particle in an external potential is identified through a thermodynamically consistent minimal lattice model that includes the chemical reaction providing the propulsion and ordinary translational noise. In the continuum limit, a unique expression follows, comprising a direct contribution from the active process and an indirect contribution from ordinary diffusive motion. From the corresponding Langevin equation, this physical entropy production cannot be inferred through the conventional, yet here ambiguous, comparison of forward and time-reversed trajectories. Generalizations to several interacting active particles and passive particles in a bath of active ones are presented explicitly, further ones are briefly indicated.

Enzymatic analysis of α-ketoglutaramate—A biomarker for hyperammonemia

PubMed Central

Halámková, Lenka; Mailloux, Shay; Halámek, Jan; Cooper, Arthur J.L.; Katz, Evgeny

2012-01-01

Two enzymatic assays were developed for the analysis of α-ketoglutaramate (KGM)—an important biomarker of hepatic encephalopathy and other hyperammonemic diseases. In both procedures, KGM is first converted to α-ketoglutarate (KTG) via a reaction catalyzed by ω-amidase (AMD). In the first procedure, KTG generated in the AMD reaction initiates a biocatalytic cascade in which the concerted action of alanine transaminase and lactate dehydrogenase results in the oxidation of NADH. In the second procedure, KTG generated from KGM is reductively aminated, with the concomitant oxidation of NADH, in a reaction catalyzed by L-glutamic dehydrogenase. In both assays, the decrease in optical absorbance (λ=340 nm) corresponding to NADH oxidation is used to quantify concentrations of KGM. The two analytical procedures were applied to 50% (v/v) human serum diluted with aqueous solutions containing the assay components and spiked with concentrations of KGM estimated to be present in normal human plasma and in plasma from hyperammonemic patients. Since KTG is the product of AMD-catalyzed hydrolysis of KGM, in a separate study, this compound was used as a surrogate for KGM. Statistical analyses of samples mimicking the concentration of KGM assumed to be present in normal and pathological concentration ranges were performed. Both enzymatic assays for KGM were confirmed to discriminate between the predicted normal and pathophysiological concentrations of the analyte. The present study is the first step toward the development of a clinically useful probe for KGM analysis in biological fluids. PMID:23141304

Bacterial SPOR domains are recruited to septal peptidoglycan by binding to glycan strands that lack stem peptides

PubMed Central

Yahashiri, Atsushi; Jorgenson, Matthew A.; Weiss, David S.

2015-01-01

Bacterial SPOR domains bind peptidoglycan (PG) and are thought to target proteins to the cell division site by binding to “denuded” glycan strands that lack stem peptides, but uncertainties remain, in part because septal-specific binding has yet to be studied in a purified system. Here we show that fusions of GFP to SPOR domains from the Escherichia coli cell-division proteins DamX, DedD, FtsN, and RlpA all localize to septal regions of purified PG sacculi obtained from E. coli and Bacillus subtilis. Treatment of sacculi with an amidase that removes stem peptides enhanced SPOR domain binding, whereas treatment with a lytic transglycosylase that removes denuded glycans reduced SPOR domain binding. These findings demonstrate unequivocally that SPOR domains localize by binding to septal PG, that the physiologically relevant binding site is indeed a denuded glycan, and that denuded glycans are enriched in septal PG rather than distributed uniformly around the sacculus. Accumulation of denuded glycans in the septal PG of both E. coli and B. subtilis, organisms separated by 1 billion years of evolution, suggests that sequential removal of stem peptides followed by degradation of the glycan backbone is an ancient feature of PG turnover during bacterial cell division. Linking SPOR domain localization to the abundance of a structure (denuded glycans) present only transiently during biogenesis of septal PG provides a mechanism for coordinating the function of SPOR domain proteins with the progress of cell division. PMID:26305949

Influence of sulfur oxidation state and steric bulk upon trifluoromethyl ketone (TFK) binding kinetics to carboxylesterases and fatty acid amide hydrolase (FAAH)

PubMed Central

Wheelock, Craig E.; Nishi, Kosuke; Ying, Andy; Jones, Paul D.; Colvin, Michael E.; Olmstead, Marilyn M.; Hammock, Bruce D.

2009-01-01

Carboxylesterases metabolize numerous exogenous and endogenous ester-containing compounds including the chemotherapeutic agent CPT-11, anti-influenza viral agent oseltamivir and many agrochemicals. Trifluoromethyl ketone (TFK)-containing compounds with a sulfur atom beta to the ketone moiety are some of the most potent carboxylesterase and amidase inhibitors identified to date. This study examined the effects of alkyl chain length (i.e., steric effects) and sulfur oxidation state upon TFK inhibitor potency (IC50) and binding kinetics (ki). The selective carboxylesterase inhibitor benzil was used as a non-TFK containing control. These effects were examined using two commercial esterases (porcine and rabbit liver esterase) and two human recombinant esterases (hCE-1 and hCE-2) as well as human recombinant fatty acid amide hydrolase (FAAH). In addition, the inhibition mechanism was examined using a combination of 1H NMR, X-ray crystallography and ab initio calculations. Overall, the data show that while sulfur oxidation state profoundly affects both inhibitor potency and binding kinetics, the steric effects dominate and override the contributions of sulfur oxidation. In addition, the data suggest that inclusion of a sulfur atom beta to the ketone contributes an increase (~5-fold) in inhibitor potency due to effects upon ketone hydration and/or intramolecular hydrogen bond formation. These results provide further information on the nature of the TFK binding interaction and will be useful in increasing our understanding of this basic biochemical process. PMID:18023188

Catalyst activator

DOEpatents

McAdon, Mark H.; Nickias, Peter N.; Marks, Tobin J.; Schwartz, David J.

2001-01-01

A catalyst activator particularly adapted for use in the activation of metal complexes of metals of Group 3-10 for polymerization of ethylenically unsaturated polymerizable monomers, especially olefins, comprising two Group 13 metal or metalloid atoms and a ligand structure including at least one bridging group connecting ligands on the two Group 13 metal or metalloid atoms.

α-Enolase Causes Proinflammatory Activation of Pulmonary Microvascular Endothelial Cells and Primes Neutrophils Through Plasmin Activation of Protease-Activated Receptor 2.

PubMed

Bock, Ashley; Tucker, Nicole; Kelher, Marguerite R; Khan, Samina Y; Gonzalez, Eduardo; Wohlauer, Max; Hansen, Kirk; Dzieciatkowska, Monika; Sauaia, Angels; Banerjee, Anirban; Moore, Ernest E; Silliman, Christopher C

2015-08-01

Proinflammatory activation of vascular endothelium leading to increased surface expression of adhesion molecules and neutrophil (PMN) sequestration and subsequent activation is paramount in the development of acute lung injury and organ injury in injured patients. We hypothesize that α-enolase, which accumulates in injured patients, primes PMNs and causes proinflammatory activation of endothelial cells leading to PMN-mediated cytotoxicity. Proteomic analyses of field plasma samples from injured versus healthy patients were used for protein identification. Human pulmonary microvascular endothelial cells (HMVECs) were incubated with α-enolase or thrombin, and intercellular adhesion molecule-1 surface expression was measured by flow cytometry. A two-event in vitro model of PMN cytotoxicity HMVECs activated with α-enolase, thrombin, or buffer was used as targets for lysophosphatidylcholine-primed or buffer-treated PMNs. The PMN priming activity of α-enolase was completed, and lysates from both PMNs and HMVECs were immunoblotted for protease-activated receptor 1 (PAR-1) and PAR-2 and coprecipitation of α-enolase with PAR-2 and plasminogen/plasmin. α-Enolase increased 10.8-fold in injured patients (P < 0.05). Thrombin and α-enolase significantly increased intercellular adhesion molecule-1 surface expression on HMVECs, which was inhibited by antiproteases, induced PMN adherence, and served as the first event in the two-event model of PMN cytotoxicity. α-Enolase coprecipitated with PAR-2 and plasminogen/plasmin on HMVECs and PMNs and induced PMN priming, which was inhibited by tranexamic acid, and enzymatic activity was not required. α-Enolase increases after injury and may activate pulmonary endothelial cells and prime PMNs through plasmin activity and PAR-2 activation. Such proinflammatory endothelial activation may predispose to PMN-mediated organ injury.

Physical Outdoor Activity versus Indoor Activity: Their Influence on Environmental Behaviors.

PubMed

Fang, Wei-Ta; Ng, Eric; Chang, Mei-Chuan

2017-07-17

There are strong evidences linking physical outdoor activity and health benefits; however, little is known about the impact on environmental behaviors. Thus, this study aims to close this gap by investigating the influence of physical outdoor activity on environmental behaviors. A total of 416 surveys were distributed to students in eight public primary schools located near the Hsinchu Science and Industrial Park in Taiwan. Findings from the analysis revealed that subjective norms had a more influential effect on environmental behaviors for participants who engaged in physical activity at outdoor parks. In contrast, descriptive norms had a direct predictive impact on environmental behaviors for participants whose main physical activity venue was at the indoor after-school centers. Research results also highlighted attitude as the strongest predictive variable influence on environmental behaviors for children who engaged in physical indoor and outdoor activities.

Physical Outdoor Activity versus Indoor Activity: Their Influence on Environmental Behaviors

PubMed Central

Fang, Wei-Ta; Ng, Eric; Chang, Mei-Chuan

2017-01-01

There are strong evidences linking physical outdoor activity and health benefits; however, little is known about the impact on environmental behaviors. Thus, this study aims to close this gap by investigating the influence of physical outdoor activity on environmental behaviors. A total of 416 surveys were distributed to students in eight public primary schools located near the Hsinchu Science and Industrial Park in Taiwan. Findings from the analysis revealed that subjective norms had a more influential effect on environmental behaviors for participants who engaged in physical activity at outdoor parks. In contrast, descriptive norms had a direct predictive impact on environmental behaviors for participants whose main physical activity venue was at the indoor after-school centers. Research results also highlighted attitude as the strongest predictive variable influence on environmental behaviors for children who engaged in physical indoor and outdoor activities. PMID:28714934

The influence of active seating on car passengers' perceived comfort and activity levels.

PubMed

Hiemstra-van Mastrigt, S; Kamp, I; van Veen, S A T; Vink, P; Bosch, T

2015-03-01

New technologies have led to an increasingly sedentary lifestyle. Sedentary behaviour is characterised by physical inactivity and is associated with several health risks. This excessive sitting does not only take place in the office or at home, but also during daily commute. Therefore, BMW AG developed an active seating system for the back seat of a car, consisting of sensors in the back rest that register upper body movements of the passenger, with which the passenger controls a game. This study evaluated three different aspects of active seating compared to other tasks (reading, working on laptop, and gaming on tablet). First, discomfort and comfort perception were measured in a 30-minute driving test. Discomfort was very low for all activities and participants felt significantly more challenged, more fit and more refreshed during active seating. Second, heart rate was measured, indicating a light intensity, but nevertheless non-sedentary, activity. Third, average and variability in activity of six postural muscles was measured by electromyography (EMG), showing a higher muscle activity and higher muscle variability for active seating compared to other activities. Active seating might stimulate movements, thereby increasing comfort and well-being. Copyright © 2014 Elsevier Ltd and The Ergonomics Society. All rights reserved.

Thermally Activated Driver

NASA Technical Reports Server (NTRS)

Kinard, William H.; Murray, Robert C.; Walsh, Robert F.

1987-01-01

Space-qualified, precise, large-force, thermally activated driver (TAD) developed for use in space on astro-physics experiment to measure abundance of rare actinide-group elements in cosmic rays. Actinide cosmic rays detected using thermally activated driver as heart of event-thermometer (ET) system. Thermal expansion and contraction of silicone oil activates driver. Potential applications in fluid-control systems where precise valve controls are needed.

Activated carbon from biomass

NASA Astrophysics Data System (ADS)

Manocha, S.; Manocha, L. M.; Joshi, Parth; Patel, Bhavesh; Dangi, Gaurav; Verma, Narendra

2013-06-01

Activated carbon are unique and versatile adsorbents having extended surface area, micro porous structure, universal adsorption effect, high adsorption capacity and high degree of surface reactivity. Activated carbons are synthesized from variety of materials. Most commonly used on a commercial scale are cellulosic based precursors such as peat, coal, lignite wood and coconut shell. Variation occurs in precursors in terms of structure and carbon content. Coir having very low bulk density and porous structure is found to be one of the valuable raw materials for the production of highly porous activated carbon and other important factor is its high carbon content. Exploration of good low cost and non conventional adsorbent may contribute to the sustainability of the environment and offer promising benefits for the commercial purpose in future. Carbonization of biomass was carried out in a horizontal muffle furnace. Both carbonization and activation were performed in inert nitrogen atmosphere in one step to enhance the surface area and to develop interconnecting porosity. The types of biomass as well as the activation conditions determine the properties and the yield of activated carbon. Activated carbon produced from biomass is cost effective as it is easily available as a waste biomass. Activated carbon produced by combination of chemical and physical activation has higher surface area of 2442 m2/gm compared to that produced by physical activation (1365 m2/gm).

Use of active video games to increase physical activity in children: a (virtual) reality?

PubMed

Foley, Louise; Maddison, Ralph

2010-02-01

There has been increased research interest in the use of active video games (in which players physically interact with images onscreen) as a means to promote physical activity in children. The aim of this review was to assess active video games as a means of increasing energy expenditure and physical activity behavior in children. Studies were obtained from computerized searches of multiple electronic bibliographic databases. The last search was conducted in December 2008. Eleven studies focused on the quantification of the energy cost associated with playing active video games, and eight studies focused on the utility of active video games as an intervention to increase physical activity in children. Compared with traditional nonactive video games, active video games elicited greater energy expenditure, which was similar in intensity to mild to moderate intensity physical activity. The intervention studies indicate that active video games may have the potential to increase free-living physical activity and improve body composition in children; however, methodological limitations prevent definitive conclusions. Future research should focus on larger, methodologically sound intervention trials to provide definitive answers as to whether this technology is effective in promoting long-term physical activity in children.

Patterns of Activity in A Global Model of A Solar Active Region

NASA Technical Reports Server (NTRS)

Bradshaw, S. J.; Viall, N. M.

2016-01-01

In this work we investigate the global activity patterns predicted from a model active region heated by distributions of nanoflares that have a range of frequencies. What differs is the average frequency of the distributions. The activity patterns are manifested in time lag maps of narrow-band instrument channel pairs. We combine hydrodynamic and forward modeling codes with a magnetic field extrapolation to create a model active region and apply the time lag method to synthetic observations. Our aim is not to reproduce a particular set of observations in detail, but to recover some typical properties and patterns observed in active regions. Our key findings are the following. (1) Cooling dominates the time lag signature and the time lags between the channel pairs are generally consistent with observed values. (2) Shorter coronal loops in the core cool more quickly than longer loops at the periphery. (3) All channel pairs show zero time lag when the line of sight passes through coronal loop footpoints. (4) There is strong evidence that plasma must be re-energized on a timescale comparable to the cooling timescale to reproduce the observed coronal activity, but it is likely that a relatively broad spectrum of heating frequencies are operating across active regions. (5) Due to their highly dynamic nature, we find nanoflare trains produce zero time lags along entire flux tubes in our model active region that are seen between the same channel pairs in observed active regions.

Meditation leads to reduced default mode network activity beyond an active task.

PubMed

Garrison, Kathleen A; Zeffiro, Thomas A; Scheinost, Dustin; Constable, R Todd; Brewer, Judson A

2015-09-01

Meditation has been associated with relatively reduced activity in the default mode network, a brain network implicated in self-related thinking and mind wandering. However, previous imaging studies have typically compared meditation to rest, despite other studies having reported differences in brain activation patterns between meditators and controls at rest. Moreover, rest is associated with a range of brain activation patterns across individuals that has only recently begun to be better characterized. Therefore, in this study we compared meditation to another active cognitive task, both to replicate the findings that meditation is associated with relatively reduced default mode network activity and to extend these findings by testing whether default mode activity was reduced during meditation, beyond the typical reductions observed during effortful tasks. In addition, prior studies had used small groups, whereas in the present study we tested these hypotheses in a larger group. The results indicated that meditation is associated with reduced activations in the default mode network, relative to an active task, for meditators as compared to controls. Regions of the default mode network showing a Group × Task interaction included the posterior cingulate/precuneus and anterior cingulate cortex. These findings replicate and extend prior work indicating that the suppression of default mode processing may represent a central neural process in long-term meditation, and they suggest that meditation leads to relatively reduced default mode processing beyond that observed during another active cognitive task.

Expression of Active Human Tissue-Type Plasminogen Activator in Escherichia coli

PubMed Central

Qiu, Ji; Swartz, James R.; Georgiou, George

1998-01-01

The formation of native disulfide bonds in complex eukaryotic proteins expressed in Escherichia coli is extremely inefficient. Tissue plasminogen activator (tPA) is a very important thrombolytic agent with 17 disulfides, and despite numerous attempts, its expression in an active form in bacteria has not been reported. To achieve the production of active tPA in E. coli, we have investigated the effect of cooverexpressing native (DsbA and DsbC) or heterologous (rat and yeast protein disulfide isomerases) cysteine oxidoreductases in the bacterial periplasm. Coexpression of DsbC, an enzyme which catalyzes disulfide bond isomerization in the periplasm, was found to dramatically increase the formation of active tPA both in shake flasks and in fermentors. The active protein was purified with an overall yield of 25% by using three affinity steps with, in sequence, lysine-Sepharose, immobilized Erythrina caffra inhibitor, and Zn-Sepharose resins. After purification, approximately 180 μg of tPA with a specific activity nearly identical to that of the authentic protein can be obtained per liter of culture in a high-cell-density fermentation. Thus, heterologous proteins as complex as tPA may be produced in an active form in bacteria in amounts suitable for structure-function studies. In addition, these results suggest the feasibility of commercial production of extremely complex proteins in E. coli without the need for in vitro refolding. PMID:9835579

"I am active": effects of a program to promote active aging.

PubMed

Mendoza-Ruvalcaba, Neyda Ma; Arias-Merino, Elva Dolores

2015-01-01

Active aging involves a general lifestyle strategy that allows preservation of both physical and mental health during the aging process. "I am Active" is a program designed to promote active aging by increased physical activity, healthy nutritional habits, and cognitive functioning. The purpose of this study was to assess the effectiveness of this program. Sixty-four healthy adults aged 60 years or older were recruited from senior centers and randomly allocated to an experimental group (n=31) or a control group (n=33). Baseline, post-test, and 6-month follow-up assessments were performed after the theoretical-practical intervention. Effect sizes were calculated. At the conclusion of the program, the experimental group showed significant improvement compared with the control group in the following domains: physical activity (falls risk, balance, flexibility, self-efficacy), nutrition (self-efficacy and nutritional status), cognitive performance (processing speed and self-efficacy), and quality of life (general, health and functionality, social and economic status). Although some declines were reported, improvements at follow-up remained in self-efficacy for physical activity, self-efficacy for nutrition, and processing speed, and participants had better nutritional status and quality of life overall. Our findings show that this program promotes improvements in domains of active aging, mainly in self-efficacy beliefs as well as in quality of life in healthy elders.

Impact of an Active Video Game on Healthy Children’s Physical Activity

PubMed Central

Abdelsamad, Dina; Baranowski, Janice; O’Connor, Teresia Margareta; Thompson, Debbe; Barnett, Anthony; Cerin, Ester; Chen, Tzu-An

2012-01-01

OBJECTIVE: This naturalistic study tests whether children receiving a new (to them) active video game spontaneously engage in more physical activity than those receiving an inactive video game, and whether the effect would be greater among children in unsafe neighborhoods, who might not be allowed to play outside. METHODS: Participants were children 9 to 12 years of age, with a BMI >50th percentile, but <99th percentile; none of these children a medical condition that would preclude physical activity or playing video games. A randomized clinical trial assigned children to receiving 2 active or 2 inactive video games, the peripherals necessary to run the games, and a Wii console. Physical activity was monitored by using accelerometers for 5 weeks over the course of a 13-week experiment. Neighborhood safety was assessed with a 12 item validated questionnaire. RESULTS: There was no evidence that children receiving the active video games were more active in general, or at anytime, than children receiving the inactive video games. The outcomes were not moderated by parent perceived neighborhood safety, child BMI z score, or other demographic characteristics. CONCLUSIONS: These results provide no reason to believe that simply acquiring an active video game under naturalistic circumstances provides a public health benefit to children. PMID:22371457

Increasing physical activity and decreasing sedentary activity in adolescent girls – The Incorporating More Physical Activity and Calcium in Teens (IMPACT) study

PubMed Central

Jones, Desiree; Hoelscher, Deanna M; Kelder, Steven H; Hergenroeder, Albert; Sharma, Shreela V

2008-01-01

Background Lack of regular physical activity and consequent sub-optimal bone mass acquisition in youth has been implicated as a primary cause of adult-onset osteoporosis. IMPACT was a behavioral theory-based 1 1/2 year randomized controlled field study aimed at increasing bone accretion in middle school girls. The objective of this study was to determine the intervention effects of the IMPACT program upon key physical and sedentary activity endpoints among schools that participated in the IMPACT study. Endpoints examined included weight bearing physical activity (WBPA); moderate to vigorous physical activity (MVPA); vigorous physical activity (VPA); MET (metabolic equivalent) – weighted WBPA and MVPA; sedentary activity; before/after-school physical activity; and weekend physical activity. Methods Primary data analysis using a pretest-posttest control group design was conducted utilizing mixed model analysis of covariance. Data gathered from the IMPACT cohort from 2000–2002 were analyzed to determine baseline versus follow-up differences in activity endpoints. Confounders investigated included ethnicity, body mass index, menarcheal status, participation in 7th grade PE/athletics, friend/familial support and neighborhood safety. Results Follow-up means were higher for participating intervention schools relative to control schools for all physical activity variables but were statistically significant only for the following variables: daily minutes of vigorous physical activity (mean difference between Intervention (I) and Control (C) = 6.00↑ minutes, 95% CI = 5.82–6.18, p = 0.05), daily after school activity minutes (mean difference between I and C = 8.95↑ minutes, 95% CI = 8.69–9.21, p = 0.04), and daily weekend activity minutes (mean difference between I and C = 19.00↑ minutes, 95% CI = 18.40–19.60, p = 0.05). The intervention significantly reduced duration of student daily TV/Video watching (mean difference between I and C = 12.11↓ minutes, 95

Discovering Anti-platelet Drug Combinations with an Integrated Model of Activator-Inhibitor Relationships, Activator-Activator Synergies and Inhibitor-Inhibitor Synergies

PubMed Central

Lombardi, Federica; Golla, Kalyan; Fitzpatrick, Darren J.; Casey, Fergal P.; Moran, Niamh; Shields, Denis C.

2015-01-01

Identifying effective therapeutic drug combinations that modulate complex signaling pathways in platelets is central to the advancement of effective anti-thrombotic therapies. However, there is no systems model of the platelet that predicts responses to different inhibitor combinations. We developed an approach which goes beyond current inhibitor-inhibitor combination screening to efficiently consider other signaling aspects that may give insights into the behaviour of the platelet as a system. We investigated combinations of platelet inhibitors and activators. We evaluated three distinct strands of information, namely: activator-inhibitor combination screens (testing a panel of inhibitors against a panel of activators); inhibitor-inhibitor synergy screens; and activator-activator synergy screens. We demonstrated how these analyses may be efficiently performed, both experimentally and computationally, to identify particular combinations of most interest. Robust tests of activator-activator synergy and of inhibitor-inhibitor synergy required combinations to show significant excesses over the double doses of each component. Modeling identified multiple effects of an inhibitor of the P2Y12 ADP receptor, and complementarity between inhibitor-inhibitor synergy effects and activator-inhibitor combination effects. This approach accelerates the mapping of combination effects of compounds to develop combinations that may be therapeutically beneficial. We integrated the three information sources into a unified model that predicted the benefits of a triple drug combination targeting ADP, thromboxane and thrombin signaling. PMID:25875950

ACTIVE ERK1 IS DIMERISED IN VIVO: BISPHOSPHODIMERS GENERATE PEAK KINASE ACTIVITY AND MONOPHOSPHODIMERS MAINTAIN BASAL ERK1 ACTIVITY

PubMed Central

Philipova, Rada; Whitaker, Michael

2012-01-01

SUMMARY ERK1 and ERK2 are widely involved in cell signalling. Using a recombinant approach, it has been shown that exogenous ERK2 is capable of dimerisation and that preventing dimerisation reduces its nuclear accumulation on stimulation. Dimerisation occurs on phosphorylation; the dimer partner of phosphorylated ERK2 may be either phosphorylated or unphosphorylated. It has been assumed that monophosphodimers are hemiactive. Here we show that ERK1 is capable of dimerisation both in vivo and in vitro. Dimerisation of human recombinant ERK1 in vitro requires both ERK1 phosphorylation and cellular cofactor(s); it leads to the formation of a high molecular weight complex that can be dissociated by treatment with β-mercaptoethanol. We demonstrate for the first time in both sea urchin embryos and human cells that native ERK forms dimers and that high ERK kinase activity is largely associated with bisphosphodimers, not with monophosphodimers or phosphorylated monomers: the activity of the bisphosphodimer is about 20-fold higher than that of the phosphorylated monomer in vitro and the bisphosphodimer shows 5 to 7-fold higher in vivo activity than the basal activity attributable to the monophosphodimer. Thus phosphorylation of both partners in the dimer is a hallmark of ERK activation. Judgments made about ERK kinase activity associated with phosphorylated monomers are at best a proxy for ERK activity. PMID:16317051

Psychological and Pedagogic Conditions of Activating Creative Activity in Students for Successful Learning

ERIC Educational Resources Information Center

Abykanova, Bakytgul; Bilyalova, Zhupar; Makhatova, Valentina; Idrissov, Salamat; Nugumanov, Samal

2016-01-01

Creative activity of a pedagogic process subject depends on the pedagogue's position, on his faith in the abilities to learn successfully, on encouragement of achievements, stimulating the initiative and activity. Successful learning by activating creative activity is possible with the presence of respectful attitude towards the pedagogic process…

Synthesis and characterization of activated carbon from white lotus via single step chemical activation

NASA Astrophysics Data System (ADS)

Andas, Jeyashelly; Midon, Muhammad Dzulfiqar

2017-08-01

Highly porous activated carbon was successfully fabricated from the stalk of Nymphaea odorata via single step chemical activation. ZnCl2 was used as the chemical activating agent in the activation process. The raw material was preliminary characterized using Fourier Transform Infrared (FTIR), ultimate analysis (CHNS/O Analyzer) and Scanning Electron Microscope (SEM). The percentage yield, iodine number (IN) and the textural properties of the activated carbon were optimized under the influence of several synthesizing parameters such as impregnation ratio, activation temperature and activation time using ZnCl2. High IN (750.11 mg/g - 967.16 mg/g) was obtained from Sodium thiosulphate volumetric method and represents the porosity of the synthesized materials. Reduction in several functional groups was observed in the FTIR spectrum of the synthesized activated carbon. SEM analysis of the activated carbon verified the formation of highly porous surface compared to the raw Nymphaea odorata. This study provides a facile synthesis of activated carbon from waste natural resources at benign condition.

Activity Profile and Energy Expenditure Among Active Older Adults, British Columbia, 2011-2012.

PubMed

Madden, Kenneth M; Ashe, Maureen C; Chase, Jocelyn M

2015-07-16

Time spent by young adults in moderate to vigorous activity predicts daily caloric expenditure. In contrast, caloric expenditure among older adults is best predicted by time spent in light activity. We examined highly active older adults to examine the biggest contributors to energy expenditure in this population. Fifty-four community-dwelling men and women aged 65 years or older (mean, 71.4 y) were enrolled in this cross-sectional observational study. All were members of the Whistler Senior Ski Team, and all met current American guidelines for physical activity. Activity levels (sedentary, light, and moderate to vigorous) were recorded by accelerometers worn continuously for 7 days. Caloric expenditure was measured using accelerometry, galvanic skin response, skin temperature, and heat flux. Significant variables were entered into a stepwise multivariate linear model consisting of activity level, age, and sex. The average (standard deviation [SD]) daily nonlying sedentary time was 564 (92) minutes (9.4 [1.5] h) per day. The main predictors of higher caloric expenditure were time spent in moderate to vigorous activity (standardized β = 0.42 [SE, 0.08]; P < .001) and male sex (standardized β = 1.34 [SE, 0.16]; P < .001). A model consisting of only moderate to vigorous physical activity and sex explained 68% of the variation in caloric expenditure. An increase in moderate to vigorous physical activity by 1 minute per day was associated with an additional 16 kcal expended in physical activity. The relationship between activity intensity and caloric expenditure in athletic seniors is similar to that observed in young adults. Active older adults still spend a substantial proportion of the day engaged in sedentary behaviors.

Standardizing Activation Analysis: New Software for Photon Activation Analysis

NASA Astrophysics Data System (ADS)

Sun, Z. J.; Wells, D.; Segebade, C.; Green, J.

2011-06-01

Photon Activation Analysis (PAA) of environmental, archaeological and industrial samples requires extensive data analysis that is susceptible to error. For the purpose of saving time, manpower and minimizing error, a computer program was designed, built and implemented using SQL, Access 2007 and asp.net technology to automate this process. Based on the peak information of the spectrum and assisted by its PAA library, the program automatically identifies elements in the samples and calculates their concentrations and respective uncertainties. The software also could be operated in browser/server mode, which gives the possibility to use it anywhere the internet is accessible. By switching the nuclide library and the related formula behind, the new software can be easily expanded to neutron activation analysis (NAA), charged particle activation analysis (CPAA) or proton-induced X-ray emission (PIXE). Implementation of this would standardize the analysis of nuclear activation data. Results from this software were compared to standard PAA analysis with excellent agreement. With minimum input from the user, the software has proven to be fast, user-friendly and reliable.

Screening of herbal extracts for activation of the human peroxisome proliferator-activated receptor.

PubMed

Rau, O; Wurglics, M; Dingermann, Th; Abdel-Tawab, M; Schubert-Zsilavecz, M

2006-11-01

The peroxisome proliferator-activated receptors play a pivotal role in metazoan lipid and glucose homeostasis. Synthetic activators of PPARalpha (fibrates) and PPARgamma (glitazones) are therefore widely used for treatment of dislipidemia and diabetes, respectively. There is growing evidence for herbal compounds to influence nuclear receptor signalling e.g. the PPARs. We recently reported carnosic acid and carnosol, both being diterpenes found in the labiate herbs sage and rosemary, to be activators of PPARgamma. The subsequent screening of a variety of ethanolic extracts, obtained from traditionally used herbs, for PPAR activation, led to an exceptionally high hit rate. Among 52 extracts nearly the half significantly activated PPARgamma and 14 activated PPARalpha in addition, whereas three of them were pan-PPAR activators, which also activated PPARdelta. The most active extracts, for which a concentration dependent effect could be shown, were the extracts of Alisma plantago aquatica (ze xie/european waterplantain), Catharanthus roseus (madagascar periwinkle), Acorus calamus (sweet calamus), Euphorbia balsamifera (balsam spurge), Jatropha curcas (barbados nut), Origanum majorana (marjoram), Zea mays (corn silk), Capsicum frutescens (chilli) and Urtica dioica (stinging nettle). The results of the present study provide a possible rationale for the traditional use of many herbs as antidiabetics.

Do physical activity facilities near schools affect physical activity in high school girls?

PubMed

Trilk, Jennifer L; Ward, Dianne S; Dowda, Marsha; Pfeiffer, Karin A; Porter, Dwayne E; Hibbert, James; Pate, Russell R

2011-03-01

To investigate associations between the number of physical activity facilities within walking distance of school and physical activity behavior in 12th grade girls during after-school hours. Girls (N=1394) from 22 schools completed a self-report to determine physical activity after 3:00 p.m. The number of physical activity facilities within a 0.75-mile buffer of the school was counted with a Geographic Information System. Associations between the number of facilities and girls' physical activity were examined using linear mixed-model analysis of variance. Overall, girls who attended schools with ≥5 facilities within the buffer reported more physical activity per day than girls in schools with <5 facilities. In addition, girls who attended rural schools with ≥5 facilities reported ∼12% more physical activity per day than girls who attended rural schools with <5 facilities. No difference existed for girls in urban/suburban schools with ≥5 vs. <5 facilities. When school siting decisions are made, the number of physical activity facilities surrounding the school should be considered to encourage physical activity in 12th grade girls. Copyright © 2011 Elsevier Ltd. All rights reserved.

Active Regions Blossoming

NASA Image and Video Library

2015-10-28

As a pair of active regions began to rotate into view, their towering magnetic field lines above them bloomed into a dazzling display of twisting arches (Oct. 27-28, 2015). Some of the lines reached over and connected with the neighboring active region. Active regions are usually the source of solar storms. The images were taken in a wavelength of extreme ultraviolet light. http://photojournal.jpl.nasa.gov/catalog/PIA20048

Activity as Reality in Defining People and Activity as a Cognitive Construct. Activity and the Activity Approach to Understanding People: The Historical Meaning of the Crisis of Cultural-Activity Psychology

ERIC Educational Resources Information Center

Asmolov, A. G.

2015-01-01

In this article we discuss the past, present, and future of the cultural activity approach as a methodology for integrating the humanities and natural sciences as well as psychotechnical and theoretical knowledge. It is suggested that the meaning of historical crisis of cultural activity-psychology consists in reflecting on the future prospects of…

Highly Active Ice Nuclei from Tree Leaf Litters Retain Activity for Decades

NASA Astrophysics Data System (ADS)

Schnell, R. C.; Hill, T. C. J.

2015-12-01

Biogenic ice nuclei (IN) studied since the 1960s were first observed in tree leaf litters, in a few bacteria species and later in fungi and lichens. Recently, viable IN bacteria in precipitation have been used as a metric of their possible role in precipitation formation. Although bacterial IN activity is deactivated by a variety of common environmental stresses, we present data showing that IN found in a potpourri of decayed plant leaves, bacteria, molds and fungi etc. in plant litters are exceptionally stable and active over decades while in storage. As such, their atmospheric IN potential is worthy of further study due to their environmental persistence. In August 1970 litter collected in a grove of deciduous trees near Red Deer, AB, Canada was tested for IN (drop freezing technique). The sample initiated ice at -4C with 109 IN per gram of litter active at -10C. A few kilograms were stored in a plastic bag and tested every few years for IN content; the IN activity remained essentially unchanged over 40 years. In 2011, litter collected in the same grove had the same IN activity as the sample tested over the intervening 40 years. Boiling a gram sample of this litter in 100 grams of water deactivated 99 % of the IN activity down to -13C. None of 88 different bacteria cultures several of which appeared to Pseudomonads (common IN producing bacteria) from the fresh litter produced any active IN. A sample of the litter was placed on the top of a 15 cm column of laboratory grade kaolin and water dripped onto the litter. Ten days later the water reached the bottom of the column. The kaolin was dried and tested for IN. The prior essentially IN free kaolin now exhibited IN activity at -4C with 105 IN active at -10C. The litter exposed kaolin retained the IN activity for 25 years. Baking the kaolin removed the active IN. This suggests that IN activity attributed to kaolin particles sometimes seen at the nucleus of snow crystals could be of biological origin.

A bioluminescent caspase-1 activity assay rapidly monitors inflammasome activation in cells.

PubMed

O'Brien, Martha; Moehring, Danielle; Muñoz-Planillo, Raúl; Núñez, Gabriel; Callaway, Justin; Ting, Jenny; Scurria, Mike; Ugo, Tim; Bernad, Laurent; Cali, James; Lazar, Dan

2017-08-01

Inflammasomes are protein complexes induced by diverse inflammatory stimuli that activate caspase-1, resulting in the processing and release of cytokines, IL-1β and IL-18, and pyroptosis, an immunogenic form of cell death. To provide a homogeneous method for detecting caspase-1 activity, we developed a bioluminescent, plate-based assay that combines a substrate, Z-WEHD-aminoluciferin, with a thermostable luciferase in an optimized lytic reagent added directly to cultured cells. Assay specificity for caspase-1 is conferred by inclusion of a proteasome inhibitor in the lytic reagent and by use of a caspase-1 inhibitor to confirm activity. This approach enables a specific and rapid determination of caspase-1 activation. Caspase-1 activity is stable in the reagent thereby providing assay convenience and flexibility. Using this assay system, caspase-1 activation has been determined in THP-1 cells following treatment with α-hemolysin, LPS, nigericin, gramicidin, MSU, R848, Pam3CSK4, and flagellin. Caspase-1 activation has also been demonstrated in treated J774A.1 mouse macrophages, bone marrow-derived macrophages (BMDMs) from mice, as well as in human primary monocytes. Caspase-1 activity was not detected in treated BMDMs derived from Casp1 -/- mice, further confirming the specificity of the assay. Caspase-1 activity can be measured directly in cultured cells using the lytic reagent, or caspase-1 activity released into medium can be monitored by assay of transferred supernatant. The caspase-1 assay can be multiplexed with other assays to monitor additional parameters from the same cells, such as IL-1β release or cell death. The caspase-1 assay in combination with a sensitive real-time monitor of cell death allows one to accurately establish pyroptosis. This assay system provides a rapid, convenient, and flexible method to specifically and quantitatively monitor caspase-1 activation in cells in a plate-based format. This will allow a more efficient and effective

Environmental Education Activities Manual, Book 3: Middle Elementary Activities.

ERIC Educational Resources Information Center

Stapp, William B., Ed.; Cox, Dorothy A., Ed.

This activities book for the middle elementary grades is the third book of a series of six books designed to provide developmental K-12 experiences designed to support the basic environmental philosophy of spaceship earth presented in Book 1. The aims of the four activity sections of this book are to aid in developing students to make them more…

Environmental Education Activities Manual, Book 2: Lower Elementary Activities.

ERIC Educational Resources Information Center

Stapp, William B., Ed.; Cox, Dorothy A., Ed.

This activities book for lower elementary grades is the second book of a series of six books designed to provide developmental K-12 experiences designed to support the basic environmental philosophy of spaceship earth. The aims of the four activity sections of this book are to aid in developing students to make them more sensitive to their…

Environmental Education Activities Manual, Book 4: Upper Elementary Activities.

ERIC Educational Resources Information Center

Stapp, William B., Ed.; Cox, Dorothy A., Ed.

This activities book for the upper elementary grades is the fourth book of a series of six books designed to provide developmental K-12 experiences designed to support the basic environmental philosophy of spaceship earth presented in Book 1. The aims of the four activity sections of this book are to aid in developing students to make them more…

A Curriculum Activities Guide to Water Pollution and Environmental Studies, Volume I - Activities.

ERIC Educational Resources Information Center

Hershey, John T., Ed.; And Others

This publication, Volume I of a two volume set, consists of many tested water pollution study activities. The activities are grouped into four headings: (1) Hydrologic Cycle, (2) Human Activities, (3) Ecological Perspectives, and (4) Social and Political Factors. Three levels of activities are provided: (1) those which increase awareness, (2)…

UV-activated persulfate oxidation and regeneration of NOM-Saturated granular activated carbon.

PubMed

An, Dong; Westerhoff, Paul; Zheng, Mengxin; Wu, Mengyuan; Yang, Yu; Chiu, Chao-An

2015-04-15

A new method of ultraviolet light (UV) activated persulfate (PS) oxidation was investigated to regenerate granular activated carbon (GAC) in drinking water applications. The improvements in iodine and methylene blue numbers measured in the GAC after ultraviolet- (UV) activated persulfate suggested that the GAC preloaded with natural organic matter (NOM) was chemically regenerated. An experimental matrix for UV-activated persulfate regeneration included a range of persulfate doses and different UV wavelengths. Over 87% of the initial iodine number for GAC was restored under the optimum conditions, perfulfate dosage 60 g/L and UV exposure 1.75 × 10(4) mJ/cm(2). The persulfate dosages had little effect on the recovery of the methylene blue number, which was approximately 65%. Persulfate activation at 185 nm was superior to activation at 254 nm. UV activation of persulfate in the presence of GAC produced acid, lowering the solution pH. Higher persulfate concentrations and UV exposure resulted in greater GAC regeneration. Typical organic and inorganic byproducts (e.g., benzene compounds and sulfate ions) were measured as a component of treated water quality safety. This study provides a proof-of-concept that can be used to optimize pilot-scale and full-scale UV-activated persulfate for regeneration of NOM-saturated GAC. Copyright © 2015 Elsevier Ltd. All rights reserved.

Active Families in the Great Outdoors: A Program to Promote Family Outdoor Physical Activity

ERIC Educational Resources Information Center

Flynn, Jennifer I.; Bassett, David R.; Fouts, Hillary N.; Thompson, Dixie L.; Coe, Dawn P.

2017-01-01

This study evaluated a 4-week program to increase the time families spent engaging in outdoor activity. Parents were provided strategies to increase family outdoor activity and locations to be active. Sixteen families completed the program. Duration and number of family outdoor activity bouts per week, type of activities, locations, and family…

The active video games' narrative impact on children's physical activities

USDA-ARS?s Scientific Manuscript database

Active video games (AVGs) capable of inducing physical activity offer an innovative approach to combating childhood obesity. Unfortunately, children's AVG game play decreases quickly, underscoring the need to identify novel methods for player engagement. Narratives have been demonstrated to influenc...

Camden active spaces: Does the construction of active school playgrounds influence children's physical activity levels? A longitudinal quasi-experiment protocol

PubMed Central

Smith, Lee; Kipps, Courtney; Aggio, Daniel; Fox, Paul; Robinson, Nigel; Trend, Verena; Munnery, Suzie; Kelly, Barry; Hamer, Mark

2014-01-01

Introduction Physical activity is essential for every facet of children's health. However, physical activity levels in British children are low. The school environment is a promising setting to increase children's physical activity but limited empirical evidence exists on how a change in the outdoor physical school environment influences physical activity behaviour. The London Borough of Camden is redesigning seven existing school playgrounds to engage children to become more physically active. The primary aim of this project is to evaluate the impact of the redesigned playgrounds on children's physical activity, well-being and physical function/fitness. Method and analysis This project will use a longitudinal quasi-experimental design. Seven experimental schools and one control school will take part. One baseline data collection session and two follow-ups will be carried out. Between baseline and follow-up, the experimental school playgrounds will be redesigned. At baseline, a series of fitness tests, anthropometric and questionnaire measurements, and 7-day objective physical activity monitoring (Actigraph accelerometer) will be carried out on children (aged 5–16 years). This will be repeated at follow-up. Changes in overall physical activity levels and levels during different times of the day (eg, school breaks) will be examined. Multilevel regression modelling will be used to analyse the data. Ethics and dissemination The results of this study will be disseminated through peer-review publications and scientific presentations. Ethical approval was obtained through the University College London Research Ethics Committee (Reference number: 4400/002). PMID:25232566

Camden active spaces: does the construction of active school playgrounds influence children's physical activity levels? A longitudinal quasi-experiment protocol.

PubMed

Smith, Lee; Kipps, Courtney; Aggio, Daniel; Fox, Paul; Robinson, Nigel; Trend, Verena; Munnery, Suzie; Kelly, Barry; Hamer, Mark

2014-01-01

Physical activity is essential for every facet of children's health. However, physical activity levels in British children are low. The school environment is a promising setting to increase children's physical activity but limited empirical evidence exists on how a change in the outdoor physical school environment influences physical activity behaviour. The London Borough of Camden is redesigning seven existing school playgrounds to engage children to become more physically active. The primary aim of this project is to evaluate the impact of the redesigned playgrounds on children's physical activity, well-being and physical function/fitness. This project will use a longitudinal quasi-experimental design. Seven experimental schools and one control school will take part. One baseline data collection session and two follow-ups will be carried out. Between baseline and follow-up, the experimental school playgrounds will be redesigned. At baseline, a series of fitness tests, anthropometric and questionnaire measurements, and 7-day objective physical activity monitoring (Actigraph accelerometer) will be carried out on children (aged 5–16 years). This will be repeated at follow-up. Changes in overall physical activity levels and levels during different times of the day (eg, school breaks) will be examined. Multilevel regression modelling will be used to analyse the data. The results of this study will be disseminated through peer-review publications and scientific presentations. Ethical approval was obtained through the University College London Research Ethics Committee (Reference number: 4400/002).

Activation Dependence of Stretch Activation in Mouse Skinned Myocardium: Implications for Ventricular Function