Search

Background Due to unmet need for bone augmentation, our aim was to promote osteogenic differentiation of human adipose stem cells (hASCs) encapsulated in gellan gum (GG) or collagen type I (COL) hydrogels with bioactive glass (experimental glass 2-06 of composition [wt-%]: Na2O 12.1, K2O 14.0, CaO 19.8, P2O5 2.5, B2O3 1.6, SiO2 50.0) extract based osteogenic medium (BaG OM) for bone construct development. GG hydrogels were crosslinked with spermidine (GG-SPD) or BaG extract (GG-BaG). Methods Mechanical properties of cell-free GG-SPD, GG-BaG, and COL hydrogels were tested in osteogenic medium (OM) or BaG OM at 0, 14, and 21 d. Hydrogel embedded hASCs were cultured in OM or BaG OM for 3, 14, and 21 d, and analyzed for viability, cell number, osteogenic gene expression, osteocalcin production, and mineralization. Hydroxyapatite-stained GG-SPD samples were imaged with Optical Projection Tomography (OPT) and Selective Plane Illumination Microscopy (SPIM) in OM and BaG OM at 21 d. Furthermore, Raman spectroscopy was used to study the calcium phosphate (CaP) content of hASC-secreted ECM in GG-SPD, GG-BaG, and COL at 21 d in BaG OM. Results The results showed viable rounded cells in GG whereas hASCs were elongated in COL. Importantly, BaG OM induced significantly higher cell number and higher osteogenic gene expression in COL. In both hydrogels, BaG OM induced strong mineralization confirmed as CaP by Raman spectroscopy and significantly improved mechanical properties. GG-BaG hydrogels rescued hASC mineralization in OM. OPT and SPIM showed homogeneous 3D cell distribution with strong mineralization in BaG OM. Also, strong osteocalcin production was visible in COL. Conclusions Overall, we showed efficacious osteogenesis of hASCs in 3D hydrogels with BaG OM with potential for bone-like grafts.

Hops are a major component of beer that is added during brewing. In addition to its wide range of bioactivity, it exhibits neuroactive properties as a sedative and sleeping aid. The compounds responsible for this activity are yet to be revealed and understood in terms of their pharmacological properties. Here we evaluated the potential of several hops flavonoids in modulating the GABAergic activity and assessed their selectivity to GABAA receptors subtypes. GABA-potentiating effects were measured using [3H]ethynylbicycloorthobenzoate (EBOB) radioligand binding assay in native and recombinant α1β3γ2, α2β3γ2 and α6β3δ receptors expressed in HEK293 cells. Flumazenil sensitivity of GABA-potentiating effects and [3H]Ro 15-4513 binding assay were used to examine the flavonoids binding to benzodiazepine site. The prenylflavonoids xanthohumol (XN), isoxanthohumol (IXN) and 8-prenylnaringenin (8PN) potentiated GABA-induced displacement of [3H]EBOB binding in a concentration-dependent manner. The IC50 for this potentiation in native GABAA receptors were 29.7 µM, 11.6 µM, 7.3 µM, respectively. In recombinant receptors, the sensitivity to prenylflavonoid potentiation of GABA-induced displacement of [3H]EBOB binding followed the order α6β3δ > α2β3γ2 > α1β3γ2 with the strongest inhibition observed by 8PN in α6β3δ (IC50 = 3.6 μM). Flumazenil had no significant effect on the prenylflavonoid-induced displacement of [3H]EBOB binding and [3H]Ro 15-4513 displacement from native GABAA receptors was only detected at high micromolar concentrations (100 µM). We identified potent prenylflavonoids in hops that positively modulate GABA-induced responses in native and αβγ/δ recombinant GABAA receptors at low micromolar concentrations. These GABAergic modulatory effects were not mediated via the high-affinity benzodiazepine binding site.

Urinary tract infections (UTIs) are a common problem worldwide. The most prevalent causative pathogen of UTI is Escherichia coli, focus of this study. The current golden standard for detecting UTI is bacterial culture, creating a major workload for hospital laboratories - cost-effective and rapid mass screening of patient samples is needed. Here we present an alternative approach to screen patient samples with a single-step assay utilising time-resolved luminescence and luminescence modulating biosensing phages. Filamentous phage M13 was biopanned for binding luminescence quenching metal (copper) and further E. coli. The screening assay luminescence modulation was further enhanced by selecting right chemical environment for the functioning phage clones. Semi-specific interaction between phage, target bacteria and metal was detected by modulation in the signal of a weakly chelating, easily quenchable lanthanide complex. In the presence of the target pathogen, the phages collected quenching metal from solution to the bacterial surface changing the quenching effect on the lanthanide label and thus modulating the signal. Our method was compared with the bacterial culture data obtained from 70 patient samples. The developed proof-of-principle screening assay showed sensitivity and a specificity at the 90% mark when compared to culture method although some samples had high turbidity and even blood. The detection limit of E. coli was in the range of 1000–10 000 colony forming units/mL. Untreated urine sample was screened and time-resolved luminescence signal result was achieved within 10 min in a single incubation step.

Most clear cell renal cell carcinomas (ccRCCs) have inactivation of the von Hippel-Lindau tumor suppressor protein (pVHL), resulting in the accumulation of hypoxia-inducible factor -subunits (HIF-) and their downstream targets. HIF-2 expression is particularly high in ccRCC and is associated with increased ccRCC growth and aggressiveness. In the canonical HIF signaling pathway, HIF-prolyl hydroxylase 3 (PHD3) suppresses HIF-2 protein by post-translational hydroxylation under sufficient oxygen availability. Here, using immunoblotting and immunofluorescence staining, qRT-PCR, and siRNA-mediated gene silencing, we show that unlike in the canonical pathway, PHD3 silencing in ccRCC cells leads to down-regulation of HIF-2 protein and mRNA. Depletion of other PHD family members had no effect on HIF-2 expression, and PHD3 knockdown in non-RCC cells resulted in the expected increase in HIF-2 protein expression. Accordingly, PHD3 knockdown decreased HIF-2 target gene expression in ccRCC cells and expression was restored upon forced HIF-2 expression. The effect of PHD3 depletion was pinpointed to HIF2A mRNA stability. In line with these in vitro results, a strong positive correlation of PHD3 and HIF2A mRNA expression in ccRCC tumors was detected. Our results suggest that in contrast to the known negative regulation of HIF-2 in most cell types, high PHD3 expression in ccRCC cells maintains elevated HIF-2 expression and that of its target genes, which may enhance kidney cancer aggressiveness.

A rapid ultra-high performance liquid chromatography–atmospheric pressure chemical ionization tandem mass spectrometric (UHPLC-APCI-MS/MS) method was developed for the analysis of vitamin K compounds: phylloquinone (PK) and menaquinones (MK-n). Non-chlorinated mobile phase composition was optimized for separation of eight vitamin K compounds on a reversed phase column in 10 min. Sample treatment with liquid and solid phase extractions and by the use of MK-4 as an internal standard enabled the quantitation of microgram level of vitamin K compounds in food. The method was used to screen and quantitate vitamin K from 17 fermented food products. The highest amount of PK was detected in kimchi (42 µg/100 g), whereas the highest MK-7 content was detected in natto (902 µg/100 g). Some MK-9 was present in kefir (5 µg/100 g). Two Chinese fermented soybean pastes contained significant amount of MK-6 (5–36 µg/100 g), MK-7 (12–86 µg/100 g), and MK-8 (22–44 µg/100 g).

Gaining insights into the processes that transform dispersed biopolymers into well-ordered structures, such as soluble spidroin-proteins to spider silk threads, is essential for attempts to understand their biological function and to mimic their unique properties. One of these processes is liquid-liquid phase separation, which can act as an intermediate step for molecular assembly. We have shown that a self-coacervation step that occurs at a very high protein concentration (> 200 gl(-1)) is crucial for the fiber assembly of an engineered triblock silk-like molecule. In this study, we demonstrate that the addition of a crowding agent lowers the concentration at which coacervation occurs by almost two orders of magnitude. Coacervates induced by addition of a crowding agent are functional in terms of fiber formation, and the crowding agent appears to affect the process solely by increasing the effective concentration of the protein. Furthermore, induction at lower concentrations allows us to study the thermodynamics of the system, which provides insights into the coacervation mechanism. We suggest that this approach will be valuable for studies of biological coacervating systems in general.

Metal-phenolic network (MPN) coatings have generated increasing interest owing to their biologically inspired nature, facile fabrication, and near-universal adherence, especially for biomedical applications. However, a key issue in biomedicine is protein fouling, and the adsorption of proteins on tannic acid-based MPNs remains to be comprehensively studied. Herein, we investigate the interaction of specific biomedically relevant proteins in solution (e.g., bovine serum albumin (BSA), immunoglobulin G (IgG), fibrinogen) and complex biological media (serum) using layer-by-layer-assembled tannic acid/FeIII MPN films. When FeIII was the outermost layer, galloyl-modified poly(2-ethyl-2-oxazoline) (P(EtOx)-Gal) could be grafted to the films through coordination bonds. Protein fouling and bacterial adhesion were greatly suppressed after functionalization with P(EtOx)-Gal and the mass of adsorbed protein was reduced by 79%. Interestingly, larger proteins adsorbed more on both the MPNs and P(EtOx)-functionalized MPNs. This study provides fundamental information on the interactions of MPNs with single proteins, mixtures of proteins as encountered in serum, and the noncovalent, coordination-based, functionalization of MPN films.

The tetrapyrrole chromophore biliverdin IXα (BV) in the bacteriophytochrome from Deinococcus radiodurans (DrBphP) is usually assumed to be fully protonated, but this assumption has not been systematically validated by experiments or extensive computations. Here, we use force field molecular dynamics simulations and quantum mechanics/molecular mechanics calculations with density functional theory and XMCQDPT2 methods to investigate the effect of the five most probable protonation forms of BV on structural stability, binding pocket interactions, and absorption spectra in the two photochromic states of DrBphP. While agreement with X-ray structural data and measured UV/vis spectra suggest that in both states the protonated form of the chromophore dominates, we also find that a minor population with a deprotonated D-ring could contribute to the red-shifted tail in the absorption spectra.

In this research we introduce a plasmonic nanoparticle based optical biosensor for monitoring of molecular binding events. The sensor utilizes spotted gold nanoparticle arrays as sensing platform. The nanoparticle spots are functionalized with capture DNA sequences complementary to the analyte (target) DNA. Upon incubation with the target sequence, it will bind on the respectively complementary functionalized particle spot. This binding changes the local refractive index, which is detected spectroscopically as the resulting changes of the localized surface plasmon resonance (LSPR) peak wavelength. In order to increase the signal, a small gold nanoparticle label is introduced. The binding can be reversed using chemical means (10 mM HCl). It is demonstrated that multiplexed detection and identification of several fungal pathogen DNA sequences subsequently on one sensor array are possible by this approach.

The bark from fast-growing willow crops contains long and strong sclerenchyma fiber bundles, which could potentially replace the pulp and annual plant fibers currently used to reinforce green polymer composites. Here we successfully demonstrate the isolation of fiber bundles of willow bark with a simple alkali treatment under much milder conditions than what pulp fiber separation requires. The fiber bundles separated had hydrophobic surfaces, which made them compatible with polymers, in this case with polylactic acid, without using any additives. The most hydrophobic fiber bundles of willow bark provided the strongest and toughest composites, superior to the corresponding isotropic composites of pulp and flax fibers. Integration of the fiber bundle isolation with a prior recovery of hot water extractable aromatics from the bark and further processing of the debarked willow stems into bioethanol and lignin, for example, could make the full valorization of the willow biomass feasible.

Lack of synthetic enantiospecific triacylglycerols (TAGs) has hindered our understanding of the impact of TAG structure on the absorption and metabolic fate of fatty acids (FAs). In a five-day feeding trial with mildly (n-3) deficient rats, the bioavailability of docosahexaenoic acid [22:6(n-3), DHA] and stearic acid (18:0) from the two different enantiomers of TAG: sn-22:6(n-3)-18:0-18:0 and sn-18:0-18:0-22:6(n-3), and their regioisomeric TAG: sn-18:0-22:6(n-3)-18:0 was compared. Less secretion of fecal DHA was detected from the sn-2 position compared with the sn-1 and sn-3 positions, but no difference was found in DHA content of the fasting plasma or in the weight of the body or organs. 18:0 was lost to feces mainly as cleaved from the primary positions but also as glycerol-bound. The 5-day intervention in rats was long enough to modify the fatty acid profile of plasma phospholipids.

Although scientists have known about the umami taste for decades, it is only recently that it has gradually gained wider public recognition as the fifth primary taste. Umami is elicited by l -glutamate, some amino acids and purine nucleotides. Glutamate is found in a wide range of foods but despite that, this taste is not generally included in taste evaluation tests in European countries because it has been found to be hard to conceptualize by the population. To the best of our knowledge, a comprehensive survey on umami familiarity in European countries has not been conducted. Therefore, we chose three countries representative of northern (Finland), central (Germany) and southern Europe (Italy) for our study. Each group contained 300, 271 and 252 participants, respectively. We collected the categorical descriptors spontaneously expressed by volunteers after tasting an umami solution alone, next to the hedonic value perceived in comparison with a watery salt solution and pure water. A significant difference in the hedonic response by country was found. The Finnish sample group used the correct word “umami” at a higher rate (15%) than Italians and Germans (both at 2%). Finland also showed better discrimination between the monosodium glutamate (umami) and sodium chloride (cooking salt) solutions. Overall, the umami taste was rated less pleasant than the salt, females preferring umami more than males, while it was the other way around for salt. Interestingly, a similar percentage of individuals with very low sensitivity for both umami and salt was detected in the countries.

Insoluble residue (INS) is a lignin-rich fraction of brewer's spent grain (BSG) that also contains β-glucan and arabinoxylan, the major constituents of dietary fiber. We investigated the effects of INS in diet-induced obese mice in terms of lipid metabolism and metabolic diseases. Male mice (C57bl6) were fed a high-fat diet (HFD), a HFD + 20% INS, a HFD + 20% cellulose (CEL), a HFD with a combination of 20% INS-CEL (1:1), or a control diet for 14 weeks. Insulin and glucose tolerance tests were performed after 12 weeks. Fasting plasma lipids, bile acid, and fecal bile acid were measured after 14 weeks of feeding, and tissues were collected for gene expression analysis. Body weight gain was significantly reduced with all fibers, but only INS and INS-CEL decreased fasting plasma low-density lipoprotein cholesterol and total cholesterol compared to HFD. CEL and INS-CEL significantly improved insulin resistance. Fecal bile acids were significantly increased by all fibers, but there was no change in plasma bile acid. Clostridium leptum was increased with all fibers, but universal bacterial diversity was only with INS and INS-CEL. In addition, INS significantly increased the abundance of Bacteriodes, while CEL decreased Atopobium and Lactobacillus. INS feeding significantly upregulated various genes of cholesterol and bile acid metabolism, such as Srebp2, Hmgcr, Ldlr, Cyp7a1, Pparα, Fxr, and Pxr, in the liver. INS, INS-CEL, and CEL significantly attenuated liver steatosis. Our results suggest that INS from BSG induced beneficial systemic changes in mice via gut microbiota, bile acids, and gene expression in the liver.

Hydrothermal pretreatment is commonly used for enhancing enzymatic hydrolysis of lignocellulosics. Spruce and wheat straw were pretreated with increasing severity and lignin characteristics were analysed. The effect of enzymatically isolated lignin on the hydrolysis of Avicel and the adsorption of a cellobiohydrolase onto lignin was measured. Non-pretreated lignins had only a minor effect on Avicel hydrolysis. The structural changes in lignin accompanying hydrothermal pretreatment were associated with increased binding and inactivation of the cellulase on the lignin surface. The inhibitory effect was more pronounced in spruce than in wheat straw lignin. However, similar pretreatment severities caused similar levels of inhibition in Avicel hydrolysis for both biomass sources. The combined severity factor of the pretreatment correlated well with the inhibitory effect of lignin.

Polyacrylamides are widely applied for instance in wastewater treatment, papermaking, oil recovery and mining. Since they are bulk chemicals used in large quantities around the world, their fate in nature is of considerable interest. Both single microbial species as well as mixed populations have been investigated for degradation. Biodegradation of polyacrylamide begins with amidase catalysed deamination of polyacrylamide to ammonia and polyacrylate. The liberated ammonia is then used as a nitrogen source for growth by the microbes. The carbon backbone, polyacrylate, is more recalcitrant to biodegradation than the amide moieties. There are nevertheless reports on microbial growth with polyacrylamide and polyacrylate as the carbon sources. Several aerobic bacteria isolated from polyacrylamide and polyacrylate containing environments, as well as from soil, have been shown to catabolize these polymers. Although enzymology of the bacterial degradation is largely unknown, some hypothetical pathways have been suggested. With white-rot fungi degradation has been demonstrated to be initiated by an indirect radical mechanism catalysed by oxidative enzymes. Anaerobic decomposition has been reported for sulfur-reducing bacteria and mixed bacterial populations.

Advanced knowledge about the importance of gut microbiota for human health has increased the interest towards using new bacteria — besides the traditionally used lactobacilli and bifidobacteria — as probiotics. Currently candidates for next generation probiotics (NGP) have been searched amongst the gut bacteria that are associated with health, including strains from the genera Bacteroides, Clostridium, Faecalibacterium and Akkermansia. Unlike Lactobacillus spp. and Bifidobacterium spp., NGP, especially Bacteroides spp. and Clostridium spp., seem to be more ambivalent regarding their potential to cause infectious diseases. Therefore thorough safety assessment of NGP strains is vital. One important aspect of the safety assessment, which is sometimes ignored, is the potential to carry and spread antibiotic resistance genes. Because of the technological challenges in the production and formulation of many NGP species, it is likely that these will mainly be sold as supplements. The suitability and safety of novel probiotics to target consumer groups and conditions need to be carefully selected to ensure that the health promoting effects can be obtained.

openaire: EC/H2020/339478/EU//LAYERENG-HYBMAT Near-infrared to green and red up-conversion is reported for a new type of hybrid (Yb,Er)-IR-806 films prepared using the atomic/molecular layer deposition (ALD/MLD)thin-film fabrication technology. These amorphous films can be realized in controllable nanothicknesses and they show an exceptionally wide absorption band in the near-infrared region. The films exhibit a highly local up-conversion excitation process, which suggests strong potential for use as e.g. temperature sensors with Ångström scale surface resolution.

Neutral sulphite pulping of softwood as a potential biorefinery was investigated, by producing pulps at two main yield levels (55–65% and 75–85%) under various process conditions and by analysing the composition of the resulting fibre fractions and pulping spent liquors. It was found that both delignification rate and selectivity were enhanced by addition of anthraquinone (AQ 0.1% on o.d. wood) whereas delignification was accelerated by higher temperature (180 °C vs. 170 °C) and initial liquor pH (10 vs. 7). The established conditions preserved significant amount of carbohydrates, especially galactoglucomannan, in the fibres at the selected yield levels. At the yield level of 65%, still over 60% of galactoglucomannan was detected in the pulps whereas 50–70% of lignin was removed. In the spent liquors, lignosulphonates as the principal component represented the possible source of by-product in addition to acetic and formic acids.

Propofol is an amphiphilic small molecule that strongly influences the function of cell membranes, yet data regarding interfacial properties of propofol remain scarce. Here we consider propofol adsorption at the air/water interface as elucidated by means of vibrational sum frequency spectroscopy (VSFS), neutron reflectometry (NR), and surface tensiometry. VSFS data show that propofol adsorbed at the air/water interface interacts with water strongly in terms of hydrogen bonding and weakly in the proximity of the hydrocarbon parts of the molecule. In the concentration range studied there is almost no change in the orientation adopted at the interface. Data from NR show that propofol forms a dense monolayer with a thickness of 8.4 A and a limiting area per molecule of 40 A2, close to the value extracted from surface tensiometry. The possibility that islands or multilayers of propofol form at the air/water interface is therefore excluded as long as the solubility limit is not exceeded. Additionally, measurements of the 1H NMR chemical shifts demonstrate that propofol does not form dimers or multimers in bulk water up to the solubility limit.

The aim of this research was to study the potential of malolactic fermentation to modify the composition of the juices of sea buckthorn, chokeberry and lingonberry. Juices were prepared with and without pectinolytic enzyme treatment, followed by fermentation with commercially available strains of Lactobacillus plantarum, originally isolated from fermented plant materials. The juices before and after the fermentation were analyzed with GC-FID, HPLC-DAD, and HPLC-MS. Enzyme treatment significantly increased the phenolic content in the juices by 11-50%. None of strains showed ability to ferment lingonberry juice. On the other hand, L. plantarum DSM 10492 and DSM 20174 converted all malic acid to lactic acid in sea buckthorn and chokeberry juices, respectively. Fermentation with DSM 10492 reduced the content of flavonols by 9-14% and hydroxycinnamic acids by 20-24% in chokeberry juice. Flavonol glycosides and sugars in sea buckthorn as well as anthocyanins in chokeberry remained unaffected by the fermentation.

Here, an oxidation/acid dual-responsive nanohybrids/ark system was produced. The microfluidics-produced nanohybrids endow the system with an orchestrated cascade from wound detection, reactive oxygen species scavenging, drug release to hydrogel formation. The drug release behavior imitates the dynamic wound healing process, thus rendering an enhanced bio-mimetic regeneration.

The estimation of the correct binding mode and affinity of a ligand into a target protein using computational methods is challenging. However, docking can introduce poses from which the correct binding mode could be identified using other methods. Here, we analyzed the reliability of binding energy estimation using the molecular mechanics-generalized Born surface area (MMGBSA) method without and with energy minimization to identify the likely ligand binding modes within docking results. MMGBSA workflow (a) outperformed docking in recognizing the correct binding modes of androgen receptor ligands and (b) improved the correlation coefficient of computational and experimental results of rescored docking poses to phosphodiesterase 4B. Combined with stability and atomic distance analysis, MMGBSA helped to (c) identify the binding modes and sites of metabolism of cytochrome P450 2A6 substrates. The standard deviation of estimated binding energy within one simulation was lowered by minimization in all three example cases. Minimization improved the identification of the correct binding modes of androgen receptor ligands. Although only three case studies are shown, the results are analogous and indicate that these behaviors could be generalized. Such identified binding modes could be further used, for example, with free energy perturbation methods to understand binding energetics more accurately.

Myeloid cell leukemia 1 (Mcl1), is an antiapoptotic member of the Bcl-2 family proteins, has gained considerable importance due to its overexpression activity prevents the oncogenic cells to undergo apoptosis. This overexpression activity of Mcl1 eventually develops strong resistance to a wide variety of anticancer agents. Therefore, designing novel inhibitors with potentials to elicit higher binding affinity and specificity to inhibit Mcl1 activity is of greater importance. Thus, Mcl1 acts as an attractive cancer target. Despite recent experimental advancement in the identification and characterization of benzothiophene and benzofuran scaffold-merged compounds, the molecular mechanisms of their binding to Mcl1 are yet to be explored. The current study demonstrates an integrated approach – pharmacophore-based 3D-QSAR, docking, molecular dynamics (MD) simulation and free-energy estimation – to access the precise and comprehensive effects of current inhibitors targeting Mcl1 together with its known activity values. The pharmacophore – ANRRR.240 – based 3D-QSAR model from the current study provided high confidence (R2=0.9154, Q2=0.8736 and RMSE = 0.3533) values. Furthermore, the docking correctly predicted the binding mode of highly active compound 42. Additionally, the MD simulation for docked complex under explicit-solvent conditions together with free-energy estimation exhibited stable interaction and binding strength over the time period. Also, the decomposition analysis revealed potential energy contributing residues – M231, M250, V253, R265, L267 and F270 – to the complex stability. Overall, the current investigation might serve as a valuable insight, either to (i) improve the binding affinity of the current compounds or (ii) discover new generation anticancer agents that can effectively downregulate Mcl1 activity. Communicated by Ramaswamy H. Sarma

In this study, a semipreparative liquid chromatography method was developed for the isolation of chemically well-defined proanthocyanidin (PA) oligomer and polymer fractions. The aim was to achieve better separation than traditionally achieved for the PAs with other chromatographic methods. The method was tested with eleven PA rich Sephadex LH-20 fractions, which originated from eleven different plant species. The resulting semipreparative fractions were analyzed by both triple quadrupole and high-resolution mass spectrometry assisted by ultrahigh-performance liquid chromatography (UPLC) separation. The results showed remarkable differences in the procyanidin to prodelphinidin ratio, mean degree of polymerization, and specific oligomeric and polymeric content. However, some of these features indicated consistent patterns between species as the function of UPLC retention time. The developed method enables the production of tens of well-defined fractions of PA oligomers and polymers from the unresolved chromatographic PA hump. Accordingly, this allows researchers to explore the most bioactive parts of the complex PA humps of any plant species, which have not been possible earlier.

Analysing the structure and dynamics of biotic interaction networks and the processes shaping them is currently one of the key fields in ecology. In this paper, we develop a novel approach to gut content analysis, thereby deriving a new perspective on community interactions and their responses to environment. For this, we use an elevational gradient in the High Arctic, asking how the environment and species traits interact in shaping predator-prey interactions involving the wolf spider Pardosa glacialis. To characterize the community of potential prey available to this predator, we used pitfall trapping and vacuum sampling. To characterize the prey actually consumed, we applied molecular gut content analysis. Using joint species distribution models, we found elevation and vegetation mass to explain the most variance in the composition of the prey community locally available. However, such environmental variables had only a small effect on the prey community found in the spider's gut. These observations indicate that Pardosa exerts selective feeding on particular taxa irrespective of environmental constraints. By directly modelling the probability of predation based on gut content data, we found that neither trait matching in terms of predator and prey body size nor phylogenetic or environmental constraints modified interaction probability. Our results indicate that taxonomic identity may be more important for predator-prey interactions than environmental constraints or prey traits. The impact of environmental change on predator-prey interactions thus appears to be indirect and mediated by its imprint on the community of available prey.

Hydrolysis of nucleoside triphosphate (NTP) plays a key role for the function of many biomolecular systems. However, the chemistry of the catalytic reaction in terms of an atomic-level understanding of the structural, dynamic, and free energy changes associated with it often remains unknown. Here, we report the molecular mechanism of adenosine triphosphate (ATP) hydrolysis in the ATP-binding cassette (ABC) transporter BtuCD-F. Free energy profiles obtained from hybrid quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) simulations show that the hydrolysis reaction proceeds in a stepwise manner. First, nucleophilic attack of an activated lytic water molecule at the ATP γ-phosphate yields ADP + HPO42– as intermediate product. A conserved glutamate that is located very close to the γ-phosphate transiently accepts a proton and thus acts as catalytic base. In the second step, the proton is transferred back from the catalytic base to the γ-phosphate, yielding ADP + H2PO4–. These two chemical reaction steps are followed by rearrangements of the hydrogen bond network and the coordination of the Mg2+ ion. The rate constant estimated from the computed free energy barriers is in very good agreement with experiments. The overall free energy change of the reaction is close to zero, suggesting that phosphate bond cleavage itself does not provide a power stroke for conformational changes. Instead, ATP binding is essential for tight dimerization of the nucleotide-binding domains and the transition of the transmembrane domains from inward- to outward-facing, whereas ATP hydrolysis resets the conformational cycle. The mechanism is likely relevant for all ABC transporters and might have implications also for other NTPases, as many residues involved in nucleotide binding and hydrolysis are strictly conserved., ATP hydrolysis in ABC transporter BtuCD follows a three-step mechanism. The reaction free energy in the protein is close to zero and thus cannot provide a power stroke for conformational transitions.

Background Viral infection is implicated in development of autoimmunity. Parvovirus B19 (B19V) nonstructural protein, NS1, a helicase, covalently modifies self double-stranded deoxyribonucleic acid (dsDNA) and induces apoptosis. This study tested whether resulting apoptotic bodies (ApoBods) containing virally modified dsDNA could induce autoimmunity in an animal model. Methods BALB/c mice were inoculated with (1) pristane-induced, (2) B19V NS1-induced, or (3) staurosporine-induced ApoBods. Serum was tested for dsDNA autoantibodies by Crithidia luciliae staining and enzyme-linked immunosorbent assay. Brain, heart, liver, and kidney pathology was examined. Deposition of self-antigens in glomeruli was examined by staining with antibodies to dsDNA, histones H1 and H4, and TATA-binding protein. Results The B19V NS1-induced ApoBod inoculation induced dsDNA autoantibodies in a dose-dependent fashion. Histopathological features of immune-mediated organ damage were evident in pristane-induced and NS1-induced ApoBod groups; severity scores were higher in these groups than in staurosporine-treated groups. Tissue damage was dependent on NS1-induced ApoBod dose. Nucleosomal antigens were deposited in target tissue from pristane-induced and NS1-induced ApoBod inoculated groups, but not in the staurosporine-induced ApoBod inoculated group. Conclusions This study demonstrated proof of principle in an animal model that virally modified dsDNA in apoptotic bodies could break tolerance to self dsDNA and induce dsDNA autoantibodies and end-organ damage., Inoculation of normal mice with apoptotic bodies, generated by parvovirus B19V helicase NS1 expression, induces dsDNA antibodies and organ damage. This study provides proof of concept that viral modification of host cell DNA can lead to autoimmunity.

Human blood is a self-regenerating lipid-rich biological fluid that is routinely collected in hospital settings. The inventory of lipid molecules found in blood plasma (plasma lipidome) offers insights into individual metabolism and physiology in health and disease. Disturbances in the plasma lipidome also occur in conditions that are not directly linked to lipid metabolism; therefore, plasma lipidomics based on MS is an emerging tool in an array of clinical diagnostics and disease management. However, challenges exist in the translation of such lipidomic data to clinical applications. These relate to the reproducibility, accuracy, and precision of lipid quantitation, study design, sample handling, and data sharing. This position paper emerged from a workshop that initiated a community-led process to elaborate and define a set of generally accepted guidelines for quantitative MS-based lipidomics of blood plasma or serum, with harmonization of data acquired on different instrumentation platforms across independent laboratories as an ultimate goal. We hope that other fields may benefit from and follow such a precedent.

Objectives Chronic stress, also associated with climacteric-related symptoms, may influence cortisol secretion. We studied cortisol metabolism in peri- and postmenopausal women with diverse climacteric-related symptoms. Study design and main outcome measures The study population was 35 women, aged 45–70 years. Plasma cortisol levels were measured from blood samples collected every 20 min over 24 h. Urinary cortisol was analysed from 24-hour urine collections. Climacteric-related symptoms (vasomotor, sleep, depressive, anxiety, cognitive, sexual, menstrual, and somatic) were evaluated with the Women’s Health Questionnaire (WHQ). Associations between cortisol variables (24-hour, night, day, maximum, minimum, morning baseline, cortisol awakening response (CAR), area under the curve, slope, and 24-hour urinary cortisol) and the symptoms were first examined with a correlation analysis. Then, the women were divided into two groups according to their climacteric symptomatology, and differences in cortisol variables between the groups were investigated. Diurnal cortisol curves by symptomatology were also analyzed visually. Results In the correlation analysis, more frequent vasomotor symptoms were associated with a higher CAR (rs = 0.37, p = 0.039) and lower 24-hour urinary cortisol excretion (rs= -0.45, p = 0.012), and more frequent depressive symptoms were associated with a higher minimum cortisol level (rs = 0.33, p = 0.0498). When the women were divided into two groups, women with more frequent vasomotor (p = 0.012) or somatic symptoms (p = 0.021) had a lower 24-hour urinary cortisol excretion than less symptomatic women. Conclusions Although previous studies have reported associations between climacteric-related symptoms and cortisol secretion, these two factors were not substantially interrelated in our study.

Microbial natural products are a tremendous source of new bioactive chemical entities for drug discovery. Next generation sequencing has revealed an unprecedented genomic potential for production of secondary metabolites by diverse micro-organisms found in the environment and in the microbiota. Genome mining has further led to the discovery of numerous uncharacterized ‘cryptic’ metabolic pathways in the classical producers of natural products such as Actinobacteria and fungi. These biosynthetic gene clusters may code for improved biologically active metabolites, but harnessing the full genetic potential has been hindered by the observation that many of the pathways are ‘silent’ under laboratory conditions. Here we provide an overview of the various biotechnological methodologies, which can be divided to pleiotropic, biosynthetic gene cluster specific, and targeted genome-wide approaches that have been developed for the awakening of microbial secondary metabolic pathways. Keywords: Biosynthesis, Chemical diversity, Genome mining, Metabolic engineering, Synthetic biology

Carbohydrate moieties are essential for the biological activity of anthracycline anticancer agents such as nogalamycin, which contains l-nogalose and l-nogalamine units. The former of these is attached through a canonical O-glycosidic linkage, but the latter is connected via an unusual dual linkage composed of C-C and O-glycosidic bonds. In this work, we have utilized enzyme immobilization techniques and synthesized l-rhodosamine-thymidine diphosphate (TDP) from α-d-glucose-1-TDP using seven enzymes. In a second step, we assembled the dual linkage system by attaching the aminosugar to an anthracycline aglycone acceptor using the glycosyl transferase SnogD and the α-ketoglutarate dependent oxygenase SnoK. Furthermore, our work indicates that the auxiliary P450-type protein SnogN facilitating glycosylation is surprisingly associated with attachment of the neutral sugar l-nogalose rather than the aminosugar l-nogalamine in nogalamycin biosynthesis.

Obesity is closely associated to several diseases such as type 2 diabetes, cardiovascular disease, hepatic steatosis, airway disease, neurodegeneration, biliary diseases and certain cancers. It is, therefore, of importance to assess the role of nutrition in disease prevention as well as its effect in the course of such pathologies. In the present study, we addressed the impact of the exposure to different obesogenic diets in the mice brains phosphoproteome. To analyze if the obesity could be able to modify the protein pattern expression of brain neurons, obesity was induced in two different groups of mice. One group of mice was fed with hyperglycemic diet (HGD) and the other one was fed with high-fat diet (HFD), both for 12 weeks. A control group of lean mice was fed with a standard diet (SD). Metabolic parameters were measured before sacrifice, and brains were harvested for label-free phosphoproteomic analysis. Mice brains were analyzed to find differences, if any, in protein phosphorylation. Interestingly, the changes were independent of the obesogenic diet as no changes were detected between the two obese groups. Dephosphorylation of proteins involved in neuronal development (among others SYNGAP1 and PPP1R9B), in vesicle trafficking (for example SNAP91 and AMPH) and in cytoskeletal functions (for example, CLASP2 and GSK3B) was identified, while increased phosphorylation was detected for microtubule proteins (such as MAP2 and MAPT). Phospho site analysis of the mouse brain proteome reveals important changes that point to a connection between diet-induced obesity and impairment of neuronal functions and signaling.

Predicting the effects of global increase in temperatures on disease virulence is challenging, especially for environmental opportunistic bacteria, because pathogen fitness may be differentially affected by temperature within and outside host environment. So far, there is very little empirical evidence on the connections between optimal temperature range and virulence in environmentally growing pathogens. Here, we explored whether the virulence of an environmentally growing opportunistic fish pathogen, Flavobacterium columnare, is malleable to evolutionary changes via correlated selection on thermal tolerance. To this end, we experimentally quantified the thermal performance curves (TPCs) for maximum biomass yield of 49 F. columnare isolates from eight different geographic locations in Finland over ten years (2003–2012). We also characterized virulence profiles of these strains in a zebra fish (Danio rerio) infection model. We show that virulence among the strains increased over the years, but thermal generalism, and in particular tolerance to higher temperatures, was negatively associated with virulence. Our data suggest that temperature has a strong effect on the pathogen genetic diversity and therefore presumably also on disease dynamics. However, the observed increase in frequency and severity of F. columnare epidemics over the last decade cannot be directly linked to bacterial evolution due to increased mean temperature, but is most likely associated with factors related to increased length of growing season, or other time‐dependent change in environment. Our study demonstrates that complex interactions between the host, the pathogen and the environment influence disease virulence of an environmentally growing opportunistic pathogen. peerReviewed

Edible mushrooms are a global food with a history of consumption spanning several millennia. However, studies utilizing modern sensory methods on mushrooms are still scarce. In this study, the sensory properties of Nordic edible mushrooms were analyzed by two methods. In the sensory profile, sous vide processed wild mushroom species Cantharellus cibarius, Craterellus tubaeformis, Boletus edulis, and Lactarius camphoratus were studied with cultivated Agaricus bisporus as a control species. The sensory profile consisted of 18 descriptors, and the 5 mushrooms differed from each other in all of them. Only B. edulis and A. bisporus were linked to typical mushroom-like odor. In projective mapping, consumers evaluated blanched wild C. cibarius, C. tubaeformis and Suillus variegatus as well as cultivated Lentinula edodes and both blanched and fresh A. bisporus based on odor and on flavor. The consumers intuitively grouped the samples into three groups: wild, fresh cultivated and processed cultivated mushrooms. Wild mushrooms had a high odor intensity and various odor descriptions but a low flavor intensity. Cultivated mushrooms had opposite descriptions. Both tests showed differences in the sensory descriptors between the cultivated and wild mushrooms with the former linked to typical ‘mushroom’, indicating the importance and need for descriptive profiles for different mushroom types.