Different effects of carbohydrate binding modules on the viscoelasticity of nanocellulose gels

Many cellulose degrading and modifying enzymes have distinct parts called carbohydrate binding modules (CBMs). The CBMs have been shown to increase the concentration of enzymes on the insoluble substrate and thereby enhance catalytic activity. It has been suggested that CBMs also have a role in disrupting or dispersing the insoluble cellulose substrate, but dispute remains and explicit evidence of such a mechanism is lacking. We produced the isolated CBMs from two major cellulases (Cel6A and Cel7A) from Trichoderma reesei as recombinant proteins in Escherichia coli. We then studied the viscoelastic properties of native unmodified cellulose nanofibrils (CNF) in combination with the highly purified CBMs to detect possible functional effects of the CBMs on the CNF. The two CBMs showed clearly different effects on the viscoelastic properties of CNF. The difference in effects is noteworthy, yet it was not possible to conclude for example disruptive effects. We discuss here the alternative explanations for viscoelastic effects on CNF caused by CBMs, including the effect of ionic cosolutes.     

Defining RNA motif–aminoglycoside interactions via two-dimensional combinatorial screening and structure–activity relationships through sequencing

RNA is an extremely important target for the development of chemical probes of function or small molecule therapeutics. Aminoglycosides are the most well studied class of small molecules to target RNA. However, the RNA motifs outside of the bacterial rRNA A-site that are likely to be bound by these compounds in biological systems is largely unknown. If such information were known, it could allow for aminoglycosides to be exploited to target other RNAs and, in addition, could provide invaluable insights into potential bystander targets of these clinically used drugs. We utilized two-dimensional combinatorial screening (2DCS), a library-versus-library screening approach, to select the motifs displayed in a 3 × 3 nucleotide internal loop library and in a 6-nucleotide hairpin library that bind with high affinity and selectivity to six aminoglycoside derivatives. The selected RNA motifs were then analyzed using structure–activity relationships through sequencing (StARTS), a statistical approach that defines the privileged RNA motif space that binds a small molecule. StARTS allowed for the facile annotation of the selected RNA motif–aminoglycoside interactions in terms of affinity and selectivity. The interactions selected by 2DCS generally have nanomolar affinities, which is higher affinity than the binding of aminoglycosides to a mimic of their therapeutic target, the bacterial rRNA A-site.     

S6K1 is a multifaceted regulator of Mdm2 that connects nutrient status and DNA damage response

p53 mediates DNA damage‐induced cell‐cycle arrest, apoptosis, or senescence, and it is controlled by Mdm2, which mainly ubiquitinates p53 in the nucleus and promotes p53 nuclear export and degradation. By searching for the kinases responsible for Mdm2 S163 phosphorylation under genotoxic stress, we identified S6K1 as a multifaceted regulator of Mdm2. DNA damage activates mTOR‐S6K1 through p38α MAPK. The activated S6K1 forms a tighter complex with Mdm2, inhibits Mdm2‐mediated p53 ubiquitination, and promotes p53 induction, in addition to phosphorylating Mdm2 on S163. Deactivation of mTOR‐S6K1 signalling leads to Mdm2 nuclear translocation, which is facilitated by S163 phosphorylation, a reduction in p53 induction, and an alteration in p53‐dependent cell death. These findings thus establish mTOR‐S6K1 as a novel regulator of p53 in DNA damage response and likely in tumorigenesis. S6K1–Mdm2 interaction presents a route for cells to incorporate the metabolic/energy cues into DNA damage response and links the aging‐controlling Mdm2–p53 and mTOR‐S6K pathways.     

Phylogenetic, genomic organization and expression analysis of hydrophobin genes in the ectomycorrhizal basidiomycete Laccaria bicolor

Hydrophobins are morphogenetic, small secreted hydrophobic fungal proteins produced in response to changing development and environmental conditions. These proteins are important in the interaction between certain fungi and their hosts. In mutualistic ectomycorrhizal fungi several hydrophobins form a subclass of mycorrhizal-induced small secreted proteins that are likely to be critical in the formation of the symbiotic interface with host root cells. In this study, two genomes of the ectomycorrhizal basidiomycete Laccaria bicolor strains S238N-H82 (from North America) and 81306 (from Europe) were surveyed to construct a comprehensive genome-wide inventory of hydrophobins and to explore their characteristics and roles during host colonization. The S238N-H82 L. bicolor hydrophobin gene family is composed of 12 genes while the 81306 strain encodes nine hydrophobins, all corresponding to class I hydrophobins. The three extra hydrophobin genes encoded by the S238N-H82 genome likely arose via gene duplication and are bordered by transposon rich regions. Expression profiles of the hydrophobin genes of L. bicolor varied greatly depending on life stage (e.g. free living mycelium vs. root colonization) and on the host root environment. We conclude from this study that the complex diversity and range of expression profiles of the Laccaria hydrophobin multi-gene family have likely been a selective advantage for this mutualist in colonizing a wide range of host plants.     

Desiccation and hypertonicity of the airway surface fluid and thermally induced asthma.

To determine whether drying and hypertonicity of the airway surface fluid (ASF) are involved in thermally induced asthma, nine subjects performed isocapnic hyperventilation (HV) (minute ventilation 62.2 +/- 8.3 l/min) of frigid air (-8.9 +/- 3.3 degrees C) while periciliary fluid was collected endoscopically from the trachea. Osmolality was measured by freezing-point depression. The baseline 1-s forced expiratory volume was 73 +/- 4% of predicted and fell 26.4% 10 min postchallenge (P > 0.0001). The volume of ASF collected was 11.0 +/- 2.2 microl at rest and remained constant during and after HV as the airways narrowed (HV 10.6 +/- 1.9, recovery 6.5 +/- 1.7 microl; P = 0.18). The osmolality also remained stable throughout (rest 336 +/- 16, HV 339 +/- 16, and recovery 352 +/- 19 mosmol/kgH(2)O, P = 0.76). These data demonstrate that airway desiccation and hypertonicity of the ASF do not develop during hyperpnea in asthma; therefore, other mechanisms must cause exercise- and hyperventilation-induced airflow limitation.     

Studies on the production of nigerloxin using agro-industrial residues by solid-state fermentation

Nigerloxin, a new and potent lipoxygenase inhibitor, was discovered in our laboratory through solid-state fermentation of wheat bran by Aspergillus niger V. Teigh (MTCC-5166). The aim of this study is to investigate the possibility of using different agro-industrial residues as nutritional supplements along with wheat bran to enhance the production of nigerloxin. Nigerloxin produced by SSF was quantified spectrophotometrically at 292 nm. The results indicate that the inhibitor production was influenced by the type of solid substrate supplemented, moisture content, pH and size of the inoculum. Individually optimized supplements were tested in different combinations to determine their effects on nigerloxin production. A twofold increase in the production of nigerloxin (4.9 ± 0.3 mg gds⁻¹) was achieved by supplementing wheat bran with 10% w/w sweet lemon peel and 5% v/w methanol at optimized process parameters, that is, an initial moisture content of 65% v/w and incubation period of 6 days with an initial inoculum size of 2 ml (8 × 10⁵ spores gds⁻¹). Nigerloxin production was stable between pH of 4 and 5.     

Somatic embryogenesis in clonal neem, <Emphasis Type="Italic">Azadirachta indica</Emphasis> A. Juss. and analysis for <Emphasis Type="Italic">in vitro</Emphasis> azadirachtin production

Summary Efficient and highly reproducible induction of somatic embryogenesis was obtained in four out of seven selected clones of neem, Azadirachta indica A. Juss. This was achieved either directly from root and nodal explants or indirectly from callus cultures initiated from leaf explants excised from 1-yr-old axenic plants. Direct induction of somatic embryogenesis was achieved both from nodal and root segments within 8 wk of culture on MS1 medium without growth regulators. However, the addition of 2.3–4.5 μM thidiazuron and 0.5 μM 2,4-dichlorophenoxyacetic acid into the medium were necessary to induce somatic embryogenesis via callus phase from leaf explants. Repetitive embryogenesis was observed within 3–4 wk following transfer of somatic embryos to a plant growth regulator-free medium. When somatic embryos of nodal and root segments were left on the induction medium without subculturing, approximately 15% of the somatic embryos developed into whole plantlets after passing through a series of developmental stages. Plantlets thus produced were hardy, lush green, and acclimatized casily under greenhouse conditions. However, somatic embryos derived from leaf explants showed low conversion rates (<5%). HPLC analysis revealed no detectable levels of azadirachtin in somatic embryos.     

Laboratory evaluation of grain protectant efficacy of different foliar extracts against maize weevil – Sitophilus zeamais (Motschulsky)

The present study was aimed to evaluate six indigenous plant extracts for their ability to protect maize from infestation by maize weevil Sitophilus zeamais (M). The plant extracts were tested for its toxicity by fumigation and grain surface coating methods. Further, the effects of botanical treatments on F1 progeny, seed emergence and antifungal activities were studied. The results indicated that the extracts of Eicchornia crassipes controlled the test insect in grain surface coating method (100%, P = 0.05). Further, the same plant extract had significantly reduced the development of F1 progeny (97 ± 0.7%). In germination experiments, T + UEx samples showed normal seed germination of maize. However, in T + Ex samples, the seed germination depended on the grain protectant efficacy of the tested botanical extracts. Always higher percantage seed germination was recorded in those treatments where there was less infestation. Extracts of Carica papaya had significantly reduced the incidence of seed-borne fungus Aspergillus flavus (4 ± 2.4%) and did not affect the seed germination when compared to other treatments and untreated control.