Catalytic and hydrodynamic properties of styrene monooxygenases from Rhocodoccus opacus 1CP are modulated by cofactor binding.
Riedel, A. ; Heine, T. ; Westphal, A.H. ; Conrad, C. ; Rathsack, P. ; Berkel, W.J.H. van; Tischler, D. - \ 2015
AMB Express 5 (2015). - ISSN 2191-0855 - 11 p.
recombinant escherichia-coli - pseudomonas-fluorescens st - functional-analysis - crystal-structure - catabolism genes - strain vlb120 - putida ca-3 - degradation - mechanism - oxide
Styrene monooxygenases (SMOs) are flavoenzymes catalyzing the epoxidation of styrene into styrene oxide. SMOs are composed of a monooxygenase (StyA) and a reductase (StyB). The latter delivers reduced FAD to StyA on the expense of NADH. We identified Rhodococcus opacus 1CP as the first microorganism to possess three different StyA isoforms occurring in two systems StyA1/StyA2B and StyA/StyB, respectively. The hydrodynamic properties of StyA isozymes were found to be modulated by the binding of the (reduced) FAD cofactor. StyA1 and SyA2B mainly occur as dimers in their active forms while StyA is a monomer. StyA1 showed the highest epoxidation activity and excellent enantioselectivity in aromatic sulfoxidation. The hydrodynamic and biocatalytic properties of SMOs from strain 1CP are of relevance for investigation of possible industrial applications.
A generic microfluidic biosensor of G protein-coupled receptor activation - impedance measurements of reversible morphological changes of reverse transfected HEK293 cells on microelectrodes
Srivastava, S.K. ; Ramaneti, R. ; Roelse, M. ; Duy Tong, H. ; Vrouwe, E.X. ; Brinkman, A.G.M. ; Smet, L.C.P.M. de; Rijn, C.J.M. van; Jongsma, M.A. - \ 2015
RSC Advances : An international journal to further the chemical sciences 5 (2015). - ISSN 2046-2069 - p. 52563 - 52570.
drug discovery - assays - technology - mechanism - responses - targets - design
Impedance spectroscopy of cell lines on interdigitated electrodes (IDEs) is an established method of monitoring receptor-specific cell shape changes in response to certain analytes. Normally, assays are done in multiwells making it a bulky, static and single use procedure. Here, we present a biosensor allowing sequential application of biological test samples with an automated microfluidic system. It is capable of monitoring relative changes in impedance using castellated IDEs of 250–500 mm diameter, covered with stable or reverse transfected HEK293 cells. Reversible activation of the Neurokinin 1 (NK1) receptor in stable cell lines was observed in response to a series of 5 minute exposures from 1 pM–10 nM of the specific ligand Substance P (SP) using impedance measurements at 10 mV and 15 kHz. An optimal flow speed of 10 ml min 1 was chosen for the 10 ml flow cell. The EC50 of 10 pM was about 10 times lower than the EC50 based on measuring changes in the calcium ion concentration. The method was also shown to work with reverse transfected cells. Plasmid DNA encoding the NK1 gene was spotted onto the electrodes and pre-incubated with a transfection agent. The overlaid HEK293 cells were subsequently transfected by the underlying DNA. After challenge with SP, the cells induced an activation response similar to the stable cell line. The microfluidic micro-electrode reverse transfection system opens up possibilities to perform parallel measurements on IDE arrays with distinct receptors per IDE in a single flow channel .
Controlling the structure and length of self-synthesizing supramolecular polymers through nucleated growth and disassembly
Pal, A. ; Malakoutikhah, M. ; Leonetti, G. ; Tezcan, M. ; Colomb-Delsuc, M. ; Nguyen, V.D. ; Gucht, J. van der; Otto, S. - \ 2015
Angewandte Chemie-International Edition 54 (2015)27. - ISSN 1433-7851 - p. 7852 - 7856.
dynamic combinatorial libraries - systems chemistry - co-micelles - polymerization - replication - driven - complexity - mechanism - selection
Directing self-assembly processes out-of-equilibrium to yield kinetically trapped materials with well-defined dimensions remains a considerable challenge. Kinetically controlled assembly of self-synthesizing peptide-functionalized macrocycles through a nucleation–growth mechanism is reported. Spontaneous fiber formation in this system is effectively shut down as most of the material is diverted into metastable non-assembling trimeric and tetrameric macrocycles. However, upon adding seeds to this mixture, well-defined fibers with controllable lengths and narrow polydispersities are obtained. This seeded growth strategy also allows access to supramolecular triblock copolymers. The resulting noncovalent assemblies can be further stabilized through covalent capture. Taken together, these results show that self-synthesizing materials, through their interplay between dynamic covalent bonds and noncovalent interactions, are uniquely suited for out-of-equilibrium self-assembly.
2-Amino-4,4a-dihydro-4a,7-dimethyl-3H-phenoxazin-3-one as an unexpected product from reduction of 5-methyl-2-nitrophenol
Jansze, S.M. ; Saggiomo, V. ; Marcelis, A.T.M. ; Lutz, M. ; Velders, A.H. - \ 2015
Tetrahedron Letters 56 (2015)9. - ISSN 0040-4039 - p. 1060 - 1062.
aromatic nitro-compounds - phenoxazinone synthase - azo-compounds - aminophenol - mechanism - cells
When attempting to synthesize symmetric 2,2'-dihydroxy-4,4'-dimethyl-azobenzene from 5-methyl-2-nitrophenol by reductive methods based on two literature procedures, an unexpected product was isolated in 30% yield. Full analysis by mass spectrometry, NMR spectroscopy, and single-crystal X-ray structure analysis, proved this product to be tricyclic 2-amino-4,4a-dihydro-4a,7-dimethyl-3H-phenoxazin-3-one. This Letter conveys a warning regarding reductive synthetic routes toward azobenzenes. We also present a novel reductive synthetic route for phenoxazines, an important class of tricyclic compounds.
Effects of nocturnal illumination on life-history decisions and fitness in two wild songbird species
Jong, M.J. de; Ouyang, J. ; Silva, A. Da; Grunsven, R.H.A. van; Kempenaers, B. ; Visser, M.E. ; Spoelstra, K. - \ 2015
Philosophical Transactions of the Royal Society B. Biological sciences 370 (2015). - ISSN 0962-8436 - 8 p.
chemical magnetoreception - photoperiodic control - birds - light - mechanism - success - vision - dawn - date
The effects of artificial night lighting on animal behaviour and fitness are largely unknown. Most studies report short-term consequences in locations that are also exposed to other anthropogenic disturbance. We know little about how the effects of nocturnal illumination vary with different light colour compositions. This is increasingly relevant as the use of LED lights becomes more common, and LED light colour composition can be easily adjusted. We experimentally illuminated previously dark natural habitat with white, green and red light, and measured the effects on life-history decisions and fitness in two free-living songbird species, the great tit (Parus major) and pied flycatcher (Ficedula hypoleuca) in two consecutive years. In 2013, but not in 2014, we found an effect of light treatment on lay date, and of the interaction of treatment and distance to the nearest lamp post on chick mass in great tits but not in pied flycatchers. We did not find an effect in either species of light treatment on breeding densities, clutch size, probability of brood failure, number of fledglings and adult survival. The finding that light colour may have differential effects opens up the possibility to mitigate negative ecological effects of nocturnal illumination by using different light spectra.
Authentication of Geographical Origin and Crop System of Grape Juices by Phenolic Compounds and Antioxidant Activity Using Chemometrics
Granato, D. ; Koot, A.H. ; Schnitzler, E. ; Ruth, S.M. van - \ 2015
Journal of Food Science 80 (2015)3. - ISSN 0022-1147 - p. C584 - C593.
in-vitro - red wines - oxidative stress - fruit juices - rats - capacity - vivo - mechanism - profile
The main goal of this work was to propose an authentication model based on the phenolic composition and antioxidant and metal chelating capacities of purple grape juices produced in Brazil and Europe in order to assess their typicality. For this purpose, organic, conventional, and biodynamic grape juices produced in Brazil (n = 65) and in Europe (n = 31) were analyzed and different multivariate class-modeling and classification statistical techniques were employed to differentiate juices based on the geographical origin and crop system. Overall, Brazilian juices, regardless of the crop system adopted, presented higher contents of total phenolic compounds and flavonoids, total monomeric anthocyanins, proanthocyanidins, flavonols, flavanols, cyanidin-3-glucoside, delphinidin-3-glucoside, and malvidin-3,5-glucoside. No differences were observed for trans-resveratrol, malvidin-3-glucoside, and pelargonidin-3-glucoside between countries and among crop systems. A total of 91% of Brazilian and 97% of European juices were adroitly classified using partial least squares discriminant analysis when the producing region was considered (92% efficiency), in which the free-radical scavenging activity toward 2,2-diphenyl-1-picrylhydrazyl, content of total phenolic compounds, gallic acid, and malvidin-3-glucoside were the variables responsible for the classification. Intraregional models based on soft independent modeling of class analogy were able to differentiate organic from conventional Brazilian juices as well as conventional and organic/biodynamic European juices.
Natural loss-of-function mutation of EDR1 conferring resistance to tomato powdery mildew in Arabidopsis thaliana accession C24
Gao, D. ; Appiano, M. ; Huibers, R.P. ; Loonen, A.E.H.M. ; Visser, R.G.F. ; Wolters, A.M.A. ; Bai, Y. - \ 2015
Molecular Plant Pathology 16 (2015)1. - ISSN 1464-6722 - p. 71 - 82.
salicylic-acid - downy mildew - gene - defense - plants - microsatellites - mechanism - evolution - cloning - kinase
To screen for potentially novel types of resistance to tomato powdery mildew Oidium neolycopersici, a disease assay was performed on 123 Arabidopsis thaliana accessions. Forty accessions were fully resistant, and one, C24, was analysed in detail. By quantitative trait locus (QTL) analysis of an F2 population derived from C24 × Sha (susceptible accession), two QTLs associated with resistance were identified in C24. Fine mapping of QTL-1 on chromosome 1 delimited the region to an interval of 58¿kb encompassing 15 candidate genes. One of these was Enhanced Disease Resistance 1 (EDR1). Evaluation of the previously obtained edr1 mutant of Arabidopsis accession Col-0, which was identified because of its resistance to powdery mildew Golovinomyces cichoracearum, showed that it also displayed resistance to O.¿neolycopersici. Sequencing of EDR1 in our C24 germplasm (referred to as C24-W) revealed two missing nucleotides in the second exon of EDR1 resulting in a premature stop codon. Remarkably, C24 obtained from other laboratories does not contain the EDR1 mutation. To verify the identity of C24-W, a DNA region containing a single nucleotide polymorphism (SNP) unique to C24 was sequenced showing that C24-W contains the C24-specific nucleotide. C24-W showed enhanced resistance to O.¿neolycopersici compared with C24 not containing the edr1 mutation. Furthermore, C24-W displayed a dwarf phenotype, which was not associated with the mutation in EDR1 and was not caused by the differential accumulation of pathogenesis-related genes. In conclusion, we identified a natural edr1 mutant in the background of C24.
The impact of elevated water nitrite concentration on physiology, growth and feed intake of African catfish Clarias gariepinus (Burchell 1822)
Roques, J.A.C. ; Schram, E. ; Spanings, T. ; Schaik, T. van; Abbink, W. ; Boerrigter, J. ; Vries, P. de; Vis, J.W. van de; Flik, G. - \ 2015
Aquaculture Research 46 (2015)6. - ISSN 1355-557X - p. 1384 - 1395.
channel catfish - rainbow-trout - fresh-water - ictalurus-punctatus - oncorhynchus-mykiss - na+/k+-atpase - carpio l. - toxicity - chloride - mechanism
The nitrite threshold concentration in rearing water of African catfish (Clarias gariepinus) was assessed. African catfish with an initial mean (SD) weight of 219.7 (57.8) g were exposed to an increasing range of water nitrite from 6 (Control) to 928 µM nitrite for 28 days. Mean (SD) plasma nitrite concentrations increased from 5.0 (3.6) to 32.5 (12.6) µM at 928 µM ambient nitrite. The increase in nitrite was accompanied by gradual increase in plasma nitrate from 41.6 (28.4) µM to 420.2 (106.4) µM. Haematocrit, haemoglobin, methemoglobin, plasma concentrations of cortisol, glucose, lactate, osmolality, gill morphology and branchial Na+/K+-ATPase activity were not affected. Feed intake, final weight, SGR, FCR and mortality were not affected. We advise not to exceed a water nitrite concentration of 43 µM (0.6 mg L-1 NO2--N) to prevent the risk of reduced growth and feed intake in African catfish aquaculture.
One-step synthesis of delta-MnO2 nanoparticles using ascorbic acid and their scavenging properties to Pb(II), Zn(II) and methylene blue
Wang, M.X. ; Pang, P. ; Koopal, L.K. ; Qiu, G.H. ; Wang, Y. ; Liu, F. - \ 2014
Materials Chemistry and Physics 148 (2014)3. - ISSN 0254-0584 - p. 1149 - 1156.
high-temperature decomposition - manganese oxide - structural evolution - oxidation-state - layered mno2 - birnessite - adsorption - mechanism - nanobelts - dissolution
To obtain delta-MnO2 particles with a large specific surface area, MnO2 was synthesized in an ice-water bath using ascorbic acid (AA) to reduce KMnO4. At pH 3 and 5 and KMnO4/AA molar ratios of 8/1 and 10/1, nanoparticles of delta-MnO2 were produced. The specific surface areas (SSAs) of the samples ranged from 163 to 207 m(2)/g. The Mn average oxidation state of the samples ranged from 3.88 to 3.98 and increased with the KMnO4/AA ratio and pH. The adsorption of the samples with respect to metal ion revealed pseudo adsorption capacities of 3425 mmol Pb2+/kg and 1789 mmol Zn2+/kg. The decolorization behaviors of sample S10-5 (produced at pH 5 and KMnO4/AA molar ratios of 10/1) to methylene blue (MB) were compared at different pH values and temperatures. After 120 min at room temperature, 97% of the MB was adsorbed, and approximately 68% was oxidized. The adsorbed amount and the level of oxidation increased with increasing temperature and decreased with increasing pH. (C) 2014 Elsevier B.V. All rights reserved.
Effect of Selected Plant Extracts and D- and L-Lysine on the Cyanobacterium Microcystis aeruginosa
Lurling, M.F.L.L.W. ; Oosterhout, J.F.X. - \ 2014
Water 6 (2014)6. - ISSN 2073-4441 - p. 1807 - 1825.
moringa-oleifera seeds - controlling eutrophication - fresh-water - amino-acid - blooms - coagulation - inhibition - mechanism - substances - phosphorus
We tested extracts from Fructus mume, Salvia miltiorrhiza and Moringa oleifera as well as L-lysine and D-Lysine as curative measures to rapidly suppress the cyanobacterium Microcystis aeruginosa NIVA-CYA 43. We tested these compounds under similar conditions to facilitate comparisons. We hypothesized that for each compound, relatively low concentrations—i.e., 5–50 mg L-1, would reduce M. aeruginosa biomass. At these low concentrations, only L-lysine caused a decline in M. aeruginosa biomass at =4.3 mg L-1. F. mume extract was effective to do so at high concentrations, i.e., at =240 mg L-1, but the others were virtually non-effective. Low pH caused by organic acids is a probable explanation for the effect of F. mume extract. No complete wipe-outs of the experimental population were achieved as Photosystem II efficiency showed a recovery after six days. L-lysine may be effective at low concentrations—meaning low material costs. However, the effect of L-lysine seems relatively short-lived. Overall, the results of our study did not support the use of the tested plant extracts and amino-acid as promising candidates for curative application in M. aeruginosa bloom control.
Probing the relation between protein–protein interactions and DNA binding for a linker mutant of the bacterial nucleoid protein H-NS
Giangrossi, M. ; Wintraecken, K. ; Spurio, R. ; Vries, R.J. de - \ 2014
Biochimica et Biophysica Acta. Proteins & Proteomics 1844 (2014)2. - ISSN 1570-9639 - p. 339 - 345.
in-vivo oligomerization - virulence gene icsa - escherichia-coli - curved dna - structuring protein - shigella-flexneri - organization - domain - mechanism - transcription
We have investigated the relationship between oligomerization in solution and DNA binding for the bacterial nucleoid protein H-NS. This was done by comparing oligomerization and DNA binding of H-NS with that of a H-NS D68V-D71V linker mutant. The double linker mutation D68V-D71V, that makes the linker significantly more hydrophobic, leads to a dramatically enhanced and strongly temperature-dependent H-NS oligomerization in solution, as detected by dynamic light scattering. The DNA binding affinity of H-NS D68V-D71V for the hns promoter region is lower and has stronger temperature dependence than that of H-NS. DNase I footprinting experiments show that at high concentrations, regions protected by H-NS D68V-D71V are larger and less defined than for H-NS. In vitro transcription assays show that the enhanced protection also leads to enhanced transcriptional repression. Whereas the lower affinity of the H-NS D68V-D71V for DNA could be caused by competition between oligomerization in solution and oligomerization on DNA, the larger size of protected regions clearly confirms the notion that cooperative binding of H-NS to DNA is related to protein–protein interactions. These results emphasize the relative contributions of protein–protein interactions and substrate-dependent oligomerization in the control of gene repression operated by H-NS.
Influence of water availability on the enzymatic hydrolysis of proteins
Butré, C.I. ; Wierenga, P.A. ; Gruppen, H. - \ 2014
Process Biochemistry 49 (2014)11. - ISSN 1359-5113 - p. 1903 - 1912.
substrate-inhibition - functional-properties - ionic-strength - amino-acid - hydration - nmr - macromolecules - mechanism - kinetics - protease
The overall rate of enzymatic protein hydrolysis decreases with increasing protein concentration (0.1–30% (w/v)) at constant enzyme/substrate ratio. To understand the role of water, the amount of available water was expressed as the ratio between free and bound water and experimentally determined from water activity and T2 relaxation time (NMR) measurements. At low protein concentrations a large excess of water is present (1.5 × 106 water molecules per protein molecule at 0.1% (w/v) whey protein isolate (WPI), but only 3984 at 30% (w/v) WPI. Assuming that 357 molecules of water are needed for full hydration of the protein, these values correspond to a 4280 and 11 times excess of water, showing that at 30% (w/v) WPI the amount of water becomes limited. At the same time, only a small decrease was observed in water activity (1.00–0.997 for 0.1–30% (w/v) WPI), and an increase of bound water measured by NMR (
Quantification of variability in trichome patterns
Greese, B. ; Huelskamp, M. ; Fleck, C. - \ 2014
Frontiers in Plant Science 5 (2014). - ISSN 1664-462X
reaction-diffusion systems - to-cell variability - gene-expression - spatial-pattern - biological-systems - lateral inhibition - voronoi diagrams - differentiation - arabidopsis - mechanism
While pattern formation is studied in various areas of biology, little is known about the intrinsic noise leading to variations between individual realizations of the pattern. One prominent example for de novo pattern formation in plants is the patterning of trichomes on Arabidopsis leaves, which involves genetic regulation and cell-to-cell communication. These processes are potentially variable due to , e.g., the abundance of cell components or environmental conditions. To elevate the understanding of the regulatory processes underlying the pattern formation it is crucial to quantitatively analyze the variability in naturally occurring patterns. Here, we review recent approaches towards characterization of noise on trichome initiation. We present methods for the quantification of spatial patterns, which are the basis for data-driven mathematical modeling and enable the analysis of noise from different sources. Besides the insight gained on trichome formation, the examination of observed trichome patterns also shows that highly regulated biological processes can be substantially affected by variability.
Shear structuring as a new method to make anisotropic structures from soy-gluten blends
Grabowska, K.J. ; Tekidou, S. ; Boom, R.M. ; Goot, A.J. van der - \ 2014
Food Research International 64 (2014). - ISSN 0963-9969 - p. 743 - 751.
starch-zein blends - wheat-flour - electron-microscopy - globular-proteins - extrusion-cooking - behavior - microstructure - dispersions - consumption - mechanism
The concept of shear-induced structuring was applied to concentrated blends of soy protein isolate (SPI) and wheat gluten (WG) to create novel semi-solid food textures. Concurrent simple shear deformation and heating (95 °C) of the protein blends generated original structures consisting of fibers or layers. The ratio of SPI to vital WG and the final concentration determined the morphology of the structure. It is hypothesized that the spatial distribution of the SPI-rich phase and the WG-rich phase in a blend was altered by the shear flow. When both phases became aligned horizontally in the shear cell, a fibrous structure was formed; when they became aligned vertically in the shear cell, a layered structure was formed. The structures obtained were analyzed visually and using texture analysis and scanning electron microscopy.
Plasma Micro-Nanotextured, Scratch, Water and Hexadecane Resistant, Superhydrophobic, and Superamphiphobic Polymeric Surfaces with Perfluorinated Monolayers
Ellinas, K. ; Pujari, S.P. ; Dragatogiannis, D.A. ; Charitidis, C.A. ; Tserepi, A. ; Zuilhof, H. ; Gogolides, E. - \ 2014
ACS Applied Materials and Interfaces 6 (2014)9. - ISSN 1944-8244 - p. 6510 - 6524.
self-assembled monolayers - contact-angle hysteresis - superoleophobic surfaces - superomniphobic surfaces - fabrication - friction - design - wettability - mechanism - adhesive
Superhydrophobic and superamphiphobic toward superoleophobic polymeric surfaces of polymethyl methacrylate (PMMA), polyether ether ketone (PEEK), and polydimethyl siloxane (PDMS) are fabricated in a two-step process: (1) plasma texturing (i.e., ion-enhanced plasma etching with simultaneous roughening), with varying plasma chemistry depending on the polymer, and subsequently (2) grafting of self-assembled perfluorododecyltrichlorosilane monolayers (SAMs). Depending on the absence or not of an etch mask (i.e., colloidal microparticle self-assembly on it), random or ordered hierarchical micro-nanotexturing can be obtained. We demonstrate that stable organic monolayers can be grafted onto all these textured polymeric surfaces. After the monolayer deposition, the initially hydrophilic polymeric surfaces become superamphiphobic with static contact angles for water and oils >153°, for hexadecane >142°, and hysteresis
Potato and Mushroom Polyphenol Oxidase Activities Are Differently Modulated by Natural Plant Extracts
Kuijpers, T.F.M. ; Herk, T. van; Vincken, J.P. ; Janssen, R.H. ; Narh, D.L. ; Berkel, W.J.H. van; Gruppen, H. - \ 2014
Journal of Agricultural and Food Chemistry 62 (2014)1. - ISSN 0021-8561 - p. 214 - 221.
tyrosinase inhibitors - chlorogenic acid - ilex-paraguariensis - constituents - activation - licorice - identification - mechanism - agents - sds
Enzymatic browning is a major quality issue in fruit and vegetable processing and can be counteracted by different natural inhibitors. Often, model systems containing a single polyphenol oxidase (PPO) are used to screen for new inhibitors. To investigate the impact of the source of PPO on the outcome of such screening, this study compared the effect of 60 plant extracts on the activity of PPO from mushroom (Agaricus bisporus, AbPPO) and PPO from potato (Solanum tuberosum, StPPO). Some plant extracts had different effects on the two PPOs: an extract that inhibited one PPO could be an activator for the other. As an example of this, the mate (Ilex paraguariensis) extract was investigated in more detail. In the presence of mate extract, oxygen consumption by AbPPO was found to be reduced >5-fold compared to a control reaction, whereas that of StPPO was increased >9-fold. RP-UHPLC-MS analysis showed that the mate extract contained a mixture of phenolic compounds and saponins. Upon incubation of mate extract with StPPO, phenolic compounds disappeared completely and saponins remained. Flash chromatography was used to separate saponins and phenolic compounds. It was found that the phenolic fraction was mainly responsible for inhibition of AbPPO and activation of StPPO. Activation of StPPO was probably caused by activation of latent StPPO by chlorogenic acid quinones.
Kinetic and structural analysis of two transferase domains inPasteurella multocida hyaluronan synthase
Kooy, F.K. ; Beeftink, H.H. ; Eppink, M.H.M. ; Tramper, J. ; Eggink, G. ; Boeriu, C.G. - \ 2014
Journal of Molecular Catalysis. B, Enzymatic 102 (2014). - ISSN 1381-1177 - p. 138 - 145.
blood-group-b - enzymological characterization - conformational-changes - n-acetylglucosamine - crystal-structure - group-a - glycosyltransferase - polypeptide - mechanism - substrate
Pasteurella multocida hyaluronan synthase (PmHAS) encompasses two transferase domains that elongatea growing hyaluronan (HA) oligosaccharide chain by addition of either GlcNAc or GlcUA residues froma corresponding UDP-sugar. Initial velocity studies of single-step elongations were conducted for bothdomains by independently varying the concentrations of the HA oligosaccharide and the UDP-sugar.Two-substrate models were discriminated by their goodness-of-fit parameters and by dead-end inhi-bition studies. A mechanistic shift from a steady-state ordered bi-bi to rapid equilibrium ordered bi-bimechanism was observed at the NAc-site between the HA6and HA8elongation. This shift was invokedby a minor reduction in turnover number kcat. Both NAc- and UA-transferase domains follow a sequentialkinetic mechanism, most likely an ordered one in which the UDP-sugar donor binds first, followed bythe HA oligosaccharide. After transfer of the sugar moiety, both products are released, first the elongatedHA oligosaccharide and then the UDP sugar. This mechanism was visualized with a structural model ofPmHAS that presented two flexible loops, one in each transferase domain; these loops form a bridgeabove the active site.
Use of dynamic membranes for the preparation of vitamin E-loaded lipid particles: An alternative to prevent fouling observed in classical cross-flow emulsification
Lauoini, A. ; Charcosset, C. ; Fessi, H. ; Schroën, C.G.P.H. - \ 2014
Chemical Engineering Journal 236 (2014). - ISSN 1385-8947 - p. 498 - 505.
droplet break-up - multiple emulsions - process parameters - drug-delivery - encapsulation - homogenization - clearance - contactor - mechanism - beds
Solid lipid particles (SLP) were introduced at the beginning of the 1990s as an alternative to encapsulation systems such as emulsions and liposomes used in cosmetic and pharmaceutical preparations. The present paper investigated for the first time the preparation of SLP based on premix emulsification with packed beds of micron-sized glass beads. A coarse pre-emulsion was prepared by mixing the aqueous phase (water and Tween 80) and the lipid phase (Precirol and vitamin E) under magnetic stirring at 1200 rpm during 15 min, followed by passing the premix through the glass beads layer. SLP were formed by cooling to room temperature of the final emulsion. SLP were successfully produced under various conditions, but was most optimally carried out by extruding a coarse O/W emulsion 6 times under a pressure of 2 bar through a dynamic membrane. For example, when a 2 mm layer of glass beads sized 63 lm was used, the premix size of 5 lm was reduced to 1.5 lm. It was found that particle size tended to decrease with increasing feed pressure, increasing number of passes, decreasing glass bead size and decreasing bed height. Even more importantly, the dynamic membrane was hardly prone to fouling compared to the membranes used in traditional cross-flow emulsification which typically need small pore size for the production of particles of similar size. In addition, the small beads could be easily cleaned by disintegrating the bed. The preparation process developed was easy to use, easy to scale-up, and the particle size could be controlled by appropriate choice of process parameters.
Rooting plant development
Scheres, B. - \ 2013
Development 140 (2013)5. - ISSN 0950-1991 - p. 939 - 941.
arabidopsis root - cell fate - meristem - differentiation - mechanism - shoot - framework - epidermis - division - pattern
In 1993, we published a paper in Development detailing the anatomical structure of the Arabidopsis root. The paper described how root growth was maintained by the precisely tuned activity of a small set of 'initials', which acted as the source of dividing and differentiating cells, and how these stem cell-like cells surrounded a few infrequently dividing cells. This work underpinned subsequent research on root developmental biology and sparked a detailed molecular analysis of how stem cell groups are positioned and maintained in plants.
Cell death of gamma interferon-stimulated human fibroblasts upon toxoplasma gondii infection induces early parasite egress and limits parasite replication
Niedelman, W. ; Sprokholt, J.K. ; Clough, B. ; Frickel, E. ; Saeij, J.P.J. - \ 2013
Infection and Immunity 81 (2013)12. - ISSN 0019-9567 - p. 4341 - 4349.
ifn-gamma - indoleamine 2,3-dioxygenase - intracellular pathogen - autophagy - iron - macrophages - tryptophan - resistance - mechanism - triggers
The intracellular protozoan parasite Toxoplasma gondii is a major food-borne illness and opportunistic infection for the immunosuppressed. Resistance to Toxoplasma is dependent on gamma interferon (IFN-¿) activation of both hematopoietic and nonhematopoietic cells. Although IFN-¿-induced innate immunity in nonhematopoietic cells has been extensively studied in mice, it remains unclear what resistance mechanisms are relied on in nonhematopoietic human cells. Here, we report an IFN-¿-induced mechanism of resistance to Toxoplasma in primary human foreskin fibroblasts (HFFs) that does not depend on the deprivation of tryptophan or iron. In addition, infection is still controlled in HFFs deficient in the p65 guanylate binding proteins GBP1 or GBP2 and the autophagic protein ATG5. Resistance is coincident with host cell death that is not dependent on the necroptosis mediator RIPK3 or caspases and is correlated with early egress of the parasite before replication. This IFN-¿-induced cell death and early egress limits replication in HFFs and could promote clearance of the parasite by immune cells.