High pressure simulations of biomolecules

Pressure is a thermodynamic variable which is particularly suitable for exploration of the properties of biological macromolecules. For proteins, in particular, denaturation induced by pressure is different from that induced by temperature or denaturants. The response of proteins to pressure changes can provide information on properties of their native and non-native states. This review focuses on molecular dynamics studies of the effect of pressure on detailed atomic models of proteins. It also reports on other theoretical approaches, such as Monte Carlo simulations, which have been used to study simplified models. Another purpose of this review is to try to point out potential future studies that may be both interesting and feasible, with constantly increasing computing power.     

Combined effects of an oxidative enzyme and dissolved humic substances on 13C-labelled 2,4-D herbicide as revealed by high-resolution 13C NMR spectroscopy

Phenoxyalkanoic acids are a widely used class of herbicides. This work employed high-resolution ¹³C NMR to study the structural changes induced by humic substances and horseradish perodixase on 2,4-dichorophenoxyacetic acid (2,4-D) ¹³C-labelled in the side chain. NMR spectra showed that humic substances chemically catalyze abiotic splitting of [¹³C]2,4-D into 2,4-dichlorophenol and [¹³C]acetic acid at pH 7 but not at pH 4.7. Peroxidase did not catalyze the oxidative degradation of [¹³C]2,4-D at any pH tested and inhibited the effect of humic substances. Catalytic degradation by humic substances was attributed to free-radical reactions enhanced by the stereochemical contribution of large conformational structures formed by heterogeneous humic molecules at neutral pHs. Inhibition of 2,4-D degradation when humic substances were combined with peroxidase was explained by modification of both chemical and conformational humic structure due to peroxidase-promoted oxidative cross-coupling among humic molecules. Our findings show for the first time that the abiotic degradation of 2,4-D is catalyzed by dissolved humic substances at neutral pH. Journal of Industrial Microbiology & Biotechnology (2001) 26, 70–76. Received 09 February 2000/ Accepted in revised form 22 May 2000     

Interaction between Wheat-Germ Agglutinin and acid phosphatase present in dry de-embryonated wheat grains

Abstract

Different hydrolases were found to be present in the crude extract of dry de-embryonated wheat grains, including acid phosphatase. When the crude extract was chromatographed on a Wheat-Germ Agglutinin-Sepharose affinity column, an aliquot of acid phosphatase bound to the column and could be eluted specifically by N-acetyl-D-glucosamine solution. Some properties of the WGA-binding acid phosphatase (pH optimum, heat stability, Triton X-100 sensitivity, inhibition by some ions, molecular weight and K_m) were studied. WGA appeared to have a stabilizing effect on the enzyme, while it was ineffective on the K_m.     

Semi-Supervised Multi-View Learning for Gene Network Reconstruction

The task of gene regulatory network reconstruction from high-throughput data is receiving increasing attention in recent years. As a consequence, many inference methods for solving this task have been proposed in the literature. It has been recently observed, however, that no single inference method performs optimally across all datasets. It has also been shown that the integration of predictions from multiple inference methods is more robust and shows high performance across diverse datasets. Inspired by this research, in this paper, we propose a machine learning solution which learns to combine predictions from multiple inference methods. While this approach adds additional complexity to the inference process, we expect it would also carry substantial benefits. These would come from the automatic adaptation to patterns on the outputs of individual inference methods, so that it is possible to identify regulatory interactions more reliably when these patterns occur. This article demonstrates the benefits (in terms of accuracy of the reconstructed networks) of the proposed method, which exploits an iterative, semi-supervised ensemble-based algorithm. The algorithm learns to combine the interactions predicted by many different inference methods in the multi-view learning setting. The empirical evaluation of the proposed algorithm on a prokaryotic model organism (E. coli) and on a eukaryotic model organism (S. cerevisiae) clearly shows improved performance over the state of the art methods. The results indicate that gene regulatory network reconstruction for the real datasets is more difficult for S. cerevisiae than for E. coli. The software, all the datasets used in the experiments and all the results are available for download at the following link: http://figshare.com/articles/Semi_supervised_Multi_View_Learning_for_Gene_Network_Reconstruction/1604827.     

Computational Model of Ca2+ Wave Propagation in Human Retinal Pigment Epithelial ARPE-19 Cells

Objective

Computational models of calcium (Ca²⁺) signaling have been constructed for several cell types. There are, however, no such models for retinal pigment epithelium (RPE). Our aim was to construct a Ca²⁺ signaling model for RPE based on our experimental data of mechanically induced Ca²⁺ wave in the in vitro model of RPE, the ARPE-19 monolayer.

Methods

We combined six essential Ca²⁺ signaling components into a model: stretch-sensitive Ca²⁺ channels (SSCCs), P₂Y₂ receptors, IP₃ receptors, ryanodine receptors, Ca²⁺ pumps, and gap junctions. The cells in our epithelial model are connected to each other to enable transport of signaling molecules. Parameterization was done by tuning the above model components so that the simulated Ca²⁺ waves reproduced our control experimental data and data where gap junctions were blocked.

Results

Our model was able to explain Ca²⁺ signaling in ARPE-19 cells, and the basic mechanism was found to be as follows: 1) Cells near the stimulus site are likely to conduct Ca²⁺ through plasma membrane SSCCs and gap junctions conduct the Ca²⁺ and IP³ between cells further away. 2) Most likely the stimulated cell secretes ligand to the extracellular space where the ligand diffusion mediates the Ca²⁺ signal so that the ligand concentration decreases with distance. 3) The phosphorylation of the IP₃ receptor defines the cell’s sensitivity to the extracellular ligand attenuating the Ca²⁺ signal in the distance.

Conclusions

The developed model was able to simulate an array of experimental data including drug effects. Furthermore, our simulations predict that suramin may interfere ligand binding on P₂Y₂ receptors or accelerate P₂Y₂ receptor phosphorylation, which may partially be the reason for Ca²⁺ wave attenuation by suramin. Being the first RPE Ca²⁺ signaling model created based on experimental data on ARPE-19 cell line, the model offers a platform for further modeling of native RPE functions.     

MAPK14/p38α-dependent modulation of glucose metabolism affects ROS levels and autophagy during starvation

Increased glycolytic flux is a common feature of many cancer cells, which have adapted their metabolism to maximize glucose incorporation and catabolism to generate ATP and substrates for biosynthetic reactions. Indeed, glycolysis allows a rapid production of ATP and provides metabolic intermediates required for cancer cells growth. Moreover, it makes cancer cells less sensitive to fluctuations of oxygen tension, a condition usually occurring in a newly established tumor environment. Here, we provide evidence for a dual role of MAPK14 in driving a rearrangement of glucose metabolism that contributes to limiting reactive oxygen species (ROS) production and autophagy activation in condition of nutrient deprivation. We demonstrate that MAPK14 is phosphoactivated during nutrient deprivation and affects glucose metabolism at 2 different levels: on the one hand, it increases SLC2A3 mRNA and protein levels, resulting in a higher incorporation of glucose within the cell. This event involves the MAPK14-mediated enhancement of HIF1A protein stability. On the other hand, MAPK14 mediates a metabolic shift from glycolysis to the pentose phosphate pathway (PPP) through the modulation of PFKFB3 (6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase 3) degradation by the proteasome. This event requires the presence of 2 distinct degradation sequences, KEN box and DSG motif Ser273, which are recognized by 2 different E3 ligase complexes. The mutation of either motif increases PFKFB3 resistance to starvation-induced degradation. The MAPK14-driven metabolic reprogramming sustains the production of NADPH, an important cofactor for many reduction reactions and for the maintenance of the proper intracellular redox environment, resulting in reduced levels of ROS. The final effect is a reduced activation of autophagy and an increased resistance to nutrient deprivation.     

Una Iscrizione Commeaiorativa Già Esistente All'Orto Botanico di Padova

Abstract

The use of the mini-plug system for the production of container seedlings is relatively new, so there is little information on the potential impact of method on the quality of planting stock. The objective in this study was to evaluate the impact of mini-plug growing method on quality of Pinus brutia seedlings, and compare the performance of this stock type with that of standard container nursery stock. Seedling survival, growth and physiological status (root growth potential, shoot electrolyte leakage) were measured after pre-cultivation in mini-plugs, at the end of the first growing season in standard containers and after field transplanting. Our results showed that mini-plug transplants of P. brutia seedlings performed as well as the standard planting stock currently used in nursery operation in Greece. For the pre-cultivation of P. brutia seedlings in mini-plugs, the use of peat and a density of 2000 mini-plugs m^?2 are recommended.