Genome mining of cyanide-degrading nitrilases from filamentous fungi

A variety of fungal species are known to degrade cyanide through the action of cyanide hydratases, a specialized subset of nitrilases which hydrolyze cyanide to formamide. In this paper, we report on two previously unknown and uncharacterized cyanide hydratases from Neurospora crassa and Aspergillus nidulans. Recombinant forms of four cyanide hydratases from N. crassa, A. nidulans, Gibberella zeae, and Gloeocercospora sorghi were prepared after their genes were cloned with N-terminal hexahistidine purification tags, expressed in Escherichia coli, and purified using immobilized metal affinity chromatography. These enzymes were compared according to their relative specific activity, pH activity profiles, thermal stability, and ability to remediate cyanide contaminated waste water from silver and copper electroplating baths. Although all four were similar, the N. crassa cyanide hydratase (CHT) has the greatest thermal stability and widest pH range of >50% activity. N. crassa also demonstrated the highest rate of cyanide degradation in the presence of both heavy metals. The CHT of A. nidulans has the highest reaction rate of the four fungal nitrilases evaluated in this work. These data will help determine optimization procedures for the possible use of these enzymes in the bioremediation of cyanide-containing waste. Similar to known plant pathogenic fungi, both N. crassa and A. nidulans were induced to express CHT by growth in the presence of KCN.     

New uses for old drugs. Auranofin, a clinically established antiarthritic metallodrug, exhibits potent antimalarial effects in vitro: Mechanistic and pharmacological implications

The clinically established gold-based antiarthritic drug auranofin (AF) manifests a pronounced reactivity toward thiol and selenol groups of proteins. In particular, AF behaves as a potent inhibitor of mammalian thioredoxin reductases causing severe intracellular oxidative stress. Given the high sensitivity of Plasmodium falciparum to oxidative stress, we thought that auranofin might act as an effective antimalarial agent. Thus, we report here new experimental results showing that auranofin and a few related gold complexes strongly inhibit P. falciparum growth in vitro. The observed antiplasmodial effects probably arise from direct inhibition of P. falciparum thioredoxin reductase. The above findings and the safe toxicity profile of auranofin warrant rapid evaluation of AF for malaria treatment in animal models.     

Phytochemical analysis and in vitro evaluation of the biological activity against herpes simplex virus type 1 (HSV-1) of Cedrus libani A. Rich.

Cedrus libani are widely used as traditional medicine in Lebanon for treatment of different infection diseases. In the present study we reported the phytochemical composition analyzed by GC-MS of wood essential oil and cones and leaves ethanol extracts. The main components of wood essential oil were himachalol (22.50%), beta-himachalene (21.90%), and alpha-himachalene (10.50%). Leaves ethanol extract was characterized by a high content of germacrene d (29.40%). The same extract obtained from cones essentially contained alpha-pinene (51.0%) and beta-myrcene (13.0%). Moreover, we investigated extracts, essential oil, and identified compounds for their in vitro antiviral activities against herpes simplex virus type 1 (HSV-1). Cytotoxicity was evaluated by MTT assay in Vero cells. Cones and leaves ethanol extracts exhibited an interesting activity with IC50 of 0.50 and 0.66 mg/ml, respectively, at non-cytotoxic concentration. A comparable activity was found when essential oil was tested (IC50 of 0.44 mg/ml).     

Identification by time-resolved EPR of the peridinins directly involved in chlorophyll triplet quenching in the peridinin-chlorophyll a-protein from Amphidinium carterae

The mechanism of triplet-triplet energy transfer in the peridinin-chlorophyll-protein (PCP) from Amphidinium carterae was investigated by time-resolved EPR (TR-EPR). The approach exploits the concept of spin conservation during triplet-triplet energy transfer, which leads to spin polarization conservation in the observed TR-EPR spectra. The acceptor (peridinin) inherits the polarization of the donor (chlorophyll) in a way which depends on the relative geometrical arrangement of the donor-acceptor couple. Starting from the initially populated chlorophyll triplet state and taking the relative positions among Chls and peridinins from the X-ray structure of PCP, we calculated the expected triplet state polarization of any peridinin in the complex. Comparison with the experimental data allowed us to propose a path for triplet quenching in the protein. The peridinin-chlorophyll pair directly involved in the triplet-triplet energy transfer coincides with the one having the shortest center to center distance. A water molecule, which is coordinated to the central Mg atom of the Chl, is also placed in close contact with the peridinin. The results support the concept of localization of the triplet state mainly in one specific peridinin in each of the two pigment subclusters related by a pseudo C2 symmetry.     

The response of cortical neurons to in vivo-like input current: theory and experiment: II. Time-varying and spatially distributed inputs

The response of a population of neurons to time-varying synaptic inputs can show a rich phenomenology, hardly predictable from the dynamical properties of the membrane’s inherent time constants. For example, a network of neurons in a state of spontaneous activity can respond significantly more rapidly than each single neuron taken individually. Under the assumption that the statistics of the synaptic input is the same for a population of similarly behaving neurons (mean field approximation), it is possible to greatly simplify the study of neural circuits, both in the case in which the statistics of the input are stationary (reviewed in La Camera et al. in Biol Cybern, 2008) and in the case in which they are time varying and unevenly distributed over the dendritic tree. Here, we review theoretical and experimental results on the single-neuron properties that are relevant for the dynamical collective behavior of a population of neurons. We focus on the response of integrate-and-fire neurons and real cortical neurons to long-lasting, noisy, in vivo-like stationary inputs and show how the theory can predict the observed rhythmic activity of cultures of neurons. We then show how cortical neurons adapt on multiple time scales in response to input with stationary statistics in vitro. Next, we review how it is possible to study the general response properties of a neural circuit to time-varying inputs by estimating the response of single neurons to noisy sinusoidal currents. Finally, we address the dendrite–soma interactions in cortical neurons leading to gain modulation and spike bursts, and show how these effects can be captured by a two-compartment integrate-and-fire neuron. Most of the experimental results reviewed in this article have been successfully reproduced by simple integrate-and-fire model neurons.     

Modelling metapopulations with stochastic membrane systems

Metapopulations, or multi-patch systems, are models describing the interactions and the behavior of populations living in fragmented habitats. Dispersal, persistence and extinction are some of the characteristics of interest in ecological studies of metapopulations. In this paper, we propose a novel method to analyze metapopulations, which is based on a discrete and stochastic modelling framework in the area of Membrane Computing. New structural features of membrane systems, necessary to appropriately describe a multi-patch system, are introduced, such as the reduction of the maximal parallel consumption of objects, the spatial arrangement of membranes and the stochastic creation of objects. The role of the additional features, their meaning for a metapopulation model and the emergence of relevant behaviors are then investigated by means of stochastic simulations. Conclusive remarks and ideas for future research are finally presented.     

Immune suppression blocks sodium-sensitive hypertension following recovery from ischemic acute renal failure

The present study determined the effect of immune suppression with mycophenolate mofetil (MMF) on sodium-sensitive hypertension following recovery from ischemia reperfusion (I/R)-induced acute renal failure. Male Sprague-Dawley rats fed 0.4% NaCl chow were subjected to 40 min bilateral I/R or control sham surgery. After 35 days of recovery, when plasma creatinine levels had returned to normal, the rats were switched to 4.0% NaCl chow for 28 days and administered vehicle or MMF (20 mg.kg(-1).day(-1) ip). High-salt mean arterial pressure was significantly higher in I/R rats (144 +/- 16 mmHg) compared with vehicle-treated sham rats (122 +/- 2 mmHg). Treatment of I/R rats with MMF during the period of high salt intake prevented the salt-induced increase in arterial pressure (114 +/- 3 mmHg). Conscious creatinine clearance was lower in I/R rats (0.27 +/- 0.07 ml.min(-1).100 g body wt(-1)) compared with vehicle-treated sham rats (0.58 +/- 0.04 ml.min(-1).100 g body wt(-1)); MMF treatment prevented the decrease in creatinine clearance in I/R rats (0.64 +/- 0.07 ml.min(-1).100 g body wt(-1)). I/R injury also significantly increased glomerular tissue damage and increased the presence of ED-1 positive (macrophages) and S100A4 positive cells (fibroblasts) in the renal interstitium. The I/R rats treated with MMF exhibited a significant reduction in infiltrating macrophages and fibroblasts and decreased histological damage. The present data indicate that infiltrating immune cells mediate or participate in the development of sodium-sensitive hypertension and renal damage in rats apparently recovered from renal I/R injury.     

Degradation of the Retinoblastoma Tumor Suppressor by the Human Papillomavirus Type 16 E7 Oncoprotein Is Important for Functional Inactivation and Is Separable from Proteasomal Degradation of E7

The steady-state level and metabolic half-life of retinoblastoma tumor suppressor protein pRB are decreased in cells that express high-risk human papillomavirus (HPV) E7 proteins. Here we show that pRB degradation is a direct activity of E7 and does not reflect a property of cell lines acquired during the selection process for E7 expression. An amino-terminal domain of E7 that does not directly contribute to pRB binding but is required for transformation is also necessary for E7-mediated pRB degradation. Treatment with inhibitors of the 26S proteasome not only blocks E7-mediated pRB degradation but also causes the stabilization of E7. Mutagenic analyses, however, reveal that the processes of proteasomal degradation of E7 and pRB are not linked processes. HPV type 16 E7 also targets the pRB-related proteins p107 and p130 for destabilization by a proteasome-dependent mechanism. Using the SAOS2 flat-cell assay as a biological indicator for pRB function, we demonstrate that pRB degradation, not solely binding, is important for the E7-induced inactivation of pRB.