Polarizability rescaling and atom-based Thole scaling in the CHARMM Drude polarizable force field for ethers

Within the CHARMM polarizable force field based on the classical Drude oscillator, atomic polarizabilities are derived via fitting to ab initio calculated data on isolated gas phase molecules, with an empirical scaling factor applied to account for differences between the gas and condensed phases. In the development of polarizable models for the ethers, a polarizability scaling factor of 0.7 was previously applied [Vorobyov et al. J Comput Chem 3:1120–1133, 2007]. While the resulting force field models gave good agreement with a variety of experimental data, they systematically underestimated the liquid phase dielectric constants. Here, a new CHARMM polarizable model is developed for the ethers, employing a polarizability scaling factor of 0.85 and including atom-based Thole scale factors recently introduced into the CHARMM Drude polarizable force field [Harder et al. J Phys Chem B 112:3509-3521, 2008]. The new model offers a significant improvement in the reproduction of liquid phase dielectric constants, while maintaining the good agreement of the previous model with all other experimental and quantum mechanical data, highlighting the sensitivity of liquid phase properties to the choice of atomic polarizability parameters.     

Reduction potential variations in azurin through secondary coordination sphere phenylalanine incorporations

Recent evidence has shown that the properties of metal binding sites can be tuned by more than the ligands in the primary coordination sphere. We investigated the incorporation of four phenylalanine residues into the secondary coordination sphere of the small soluble blue copper protein azurin. The locations for placement of these residues in azurin were based on the structure of the highly hydrophobic blue copper protein rusticyanin, which is known to have a significantly higher reduction potential than azurin. Using site-directed mutagenesis, these residues in close proximity to the copper binding site were mutated to large hydrophobic phenylalanine residues individually and in combination. We also added the Met121Leu mutation on top of the Phe mutations to construct a total of 13 variants. We found little change in the UV-visible absorption and EPR data for these proteins, however modest increases in reduction potential were observed with increases by as much as 30 mV per Phe residue. Furthermore, we observed the increases in potential to be additive.     

The role of the voltage-dependent anion channels in the outer membrane of mitochondria in the regulation of cellular metabolism

The role of the voltage-dependent anion channels (VDAC) harbored in the outer membrane of mitochondria in the regulation of cellular metabolism was investigated using an experimental model of ethanol toxicity in cultured hepatocytes. It was demonstrated that ethanol inhibits State 3 and uncoupled mitochondrial respirations, decreases the accessibility of mitochondrial adenylate kinase localized in the intermembrane space of mitochondria, and suppresses ureagenic respiration and synthesis of urea in cultured hepatocytes. Increasing the permeability of the outer mitochondrial membrane with closed VDAC with high concentrations of digitonin (> 80 microM), which creates pores in the membrane, allowing the alternative bypass of closed VDAC, and restores all reactions suppressed with ethanol. It is concluded that the effect of ethanol in hepatocytes leads to global loss of mitochondrial functions due to the closure of VDAC, which limits the free diffusion of metabolites into the intermembrane space of mitochondria. Our studies demonstrated that ethanol affects the main mitochondrial functions and revealed the role of VDAC channels in the outer mitochondrial membrane in the regulation of liver specific intracellular processes such as ureagenesis. The data obtained can be used for the development of pharmaceutical drugs that prevent the closure of VDAC in mitochondria of ethanol oxidizing liver, thus protecting liver tissue from the hepatotoxic action of alcohol.     

Tracing conformational changes in proteins

Background

Many proteins undergo extensive conformational changes as part of their functionality. Tracing these changes is important for understanding the way these proteins function. Traditional biophysics-based conformational search methods require a large number of calculations and are hard to apply to large-scale conformational motions.

Results

In this work we investigate the application of a robotics-inspired method, using backbone and limited side chain representation and a coarse grained energy function to trace large-scale conformational motions. We tested the algorithm on four well known medium to large proteins and we show that even with relatively little information we are able to trace low-energy conformational pathways efficiently. The conformational pathways produced by our methods can be further filtered and refined to produce more useful information on the way proteins function under physiological conditions.

Conclusions

The proposed method effectively captures large-scale conformational changes and produces pathways that are consistent with experimental data and other computational studies. The method represents an important first step towards a larger scale modeling of more complex biological systems.

     

Sea urchin egg fertilization studied with a fluorescent probe (ANS)

The rates of intracellular DNA synthesis at various temperatures between 39 ° and 31 °C were determined in hamster fibroblasts and HeLa cells by measuring average amounts of ³H-thymidine incorporated per cell in S phase per unit of time. The energy of activation and Q₁₀ for intracellular DNA synthesis were calculated from the slopes of the relative rates of DNA synthesis in HeLa cells and hamster fibroblasts vs. time, plotted on Arrhenius coordinates. In both cell types the incorporation of thymidine into DNA is characterized by an energy of activation of 21 000 calories/mole and a Q₁₀ of 2.94. The absolute rates of DNA synthesis were determined in hamster cells at various temperatures, with values ranging from 1.44 to 0.60 × 10^?14 g DNA/ min/cell at 39 ° to 31 °C, respectively. The length of the S phase of the hamster cell was calculated over a 39 ° to 31 °C range, and found to be 5.0 to 11.9 h, respectively. It is concluded that the S phase length is partly determined by the rate of temperature-dependent DNA synthesis.     

A new neural framework for visuospatial processing

The division of cortical visual processing into distinct dorsal and ventral streams is a key framework that has guided visual neuroscience. The characterization of the ventral stream as a 'What' pathway is relatively uncontroversial, but the nature of dorsal stream processing is less clear. Originally proposed as mediating spatial perception ('Where'), more recent accounts suggest it primarily serves non-conscious visually guided action ('How'). Here, we identify three pathways emerging from the dorsal stream that consist of projections to the prefrontal and premotor cortices, and a major projection to the medial temporal lobe that courses both directly and indirectly through the posterior cingulate and retrosplenial cortices. These three pathways support both conscious and non-conscious visuospatial processing, including spatial working memory, visually guided action and navigation, respectively.     

Stable isotopes of Lithosiini and lichens in Hong Kong show the biodindicator potential of lichenivorous moths

Urban landscapes provide unique environments for a wide variety of plants and animals, but their suitability may be limited by anthropogenic impacts such as pollution. We examined the potential utility of lichen and lichen-feeding moths as biodindicators of air pollution in Hong Kong by comparing carbon (C) and nitrogen (N) stable isotope values in lichens, lichenivorous and non-lichenivorous moths (Lepidoptera: Erebidae) and a moth outgroup (Lepidoptera: Geometridae). Our results show that stable isotope values for C and N were similar for lichens and lichen feeding moths, while non-lichen feeding moths formed a distinct group. In addition, we found consistent δ¹³C and δ¹⁵N values across moth body parts, indicating that any portion of the specimen is suitable for isotopic fingerprinting. Our results highlight that lichen feeding moths may be useful for integrating signals of atmospheric nitrogen pollution and could therefore have utility in monitoring and quantifying air quality over time and space.     

Deletion of glyceraldehyde-3-phosphate dehydrogenase (gapN) in Clostridium saccharoperbutylacetonicum N1-4(HMT) using CLEAVE™ increases the ATP pool and accelerates solvent production

The development and advent of mutagenesis tools for solventogenic clostridial species in recent years has allowed for the increased refinement of industrially relevant strains. In this study we have utilised CLEAVE™, a CRISPR/Cas genome editing system developed by Green Biologics Ltd., to engineer a strain of Clostridium saccharoperbutylacetonicum N1-4(HMT) with potentially useful solvents titres and energy metabolism. As one of two enzymes responsible for the conversion of glyceraldehyde-3-phosphate (GAP) to 3-phosphoglyceric acid in glycolysis, it was hypothesised that deletion of gapN would increase ATP and NADH production that could in turn improve solvent production. Herein, whole genome sequencing has been used to evaluate CLEAVE™ and the successful knockout of gapN, demonstrating a clean knockout with no other detectable variations from the wild type sequence. Elevated solvent levels were detected during the first 24 h of batch fermentation, indicating an earlier shift to solventogenesis. A 2.4-fold increase in ATP concentration was observed, and quantitation of NAD(P)H derivatives revealed a more reducing cytoplasm for the gapN strain. These findings expand our understanding of clostridium carbon metabolism and report a new approach to optimising biofuel production.     

Predicted sweat rates for group water planning in sport: accuracy and application

This study tested the accuracy of a novel, limited-availability web application (H₂Q™) for predicting sweat rates in a variety of sports using estimates of energy expenditure and air temperature only. The application of predictions for group water planning was investigated for soccer match play. Fourteen open literature studies were identified where group sweat rates were reported (n = 20 group means comprising 230 individual observations from 179 athletes) with fidelity. Sports represented included: walking, cycling, swimming, and soccer match play. The accuracy of H₂Q™ sweat rates was tested by comparing to measured group sweat rates using the concordance correlation coefficient (CCC) with 95% confidence interval [CI]. The relative absolute error (RAE) with 95% [CI] was also assessed, whereby the mean absolute error was expressed relative to an acceptance limit of 0.250 L/h. The CCC was 0.98 [0.95, 0.99] and the RAE was 0.449 [0.279, 0.620], indicating that the prediction error was on average 0.112 L/h. The RAE was < 1.0 for 19/20 observations (95%). Drink volumes modeled as a proxy for sweat losses during soccer match play prevented dehydration (< 1% loss of body mass). The H₂Q™ web application demonstrated high group sweat prediction accuracy for the variety of sports activities tested. Water planning for soccer match play suggests the feasibility of easily and accurately predicting sweat rates to plan group water needs and promote optimal hydration in training and/or competition.