Quantitative analysis of DNA-protein interactions using double-labeled native gel electrophoresis and fluorescence-based imaging

We have developed a sensitive, non-radioactive method to assess the interaction of transcription factors/DNA-binding proteins with DNA. We have modified the traditional radiolabeled DNA gel mobility shift assay to incorporate a DNA probe end-labeled with a Texas-red fluorophore and a DNA-binding protein tagged with the green fluorescent protein to monitor precisely DNA-protein complexation by native gel electrophoresis. We have applied this method to the DNA-binding proteins telomere release factor-1 and the sex-determining region-Y, demonstrating that the method is sensitive (able to detect 100 fmol of fluorescently labeled DNA), permits direct visualization of both the DNA probe and the DNA-binding protein, and enables quantitative analysis of DNA and protein complexation, and thereby an estimation of the stoichiometry of protein-DNA binding.

Quantum effects on adsorption and diffusion of hydrogen and deuterium in microporous materials

Monte Carlo and molecular dynamics simulations and neutron scattering experiments are used to study the adsorption and diffusion of hydrogen and deuterium in zeolite Rho in the temperature range of 30-150 K. In the molecular simulations, quantum effects are incorporated via the Feynman-Hibbs variational approach. We suggest a new set of potential parameters for hydrogen, which can be used when Feynman-Hibbs variational approach is used for quantum corrections. The dynamic properties obtained from molecular dynamics simulations are in excellent agreement with the experimental results and show significant quantum effects on the transport at very low temperature. The molecular dynamics simulation results show that the quantum effect is very sensitive to pore dimensions and under suitable conditions can lead to a reverse kinetic molecular sieving with deuterium diffusing faster than hydrogen.

Recent developments in C5/C5a inhibitors

Complement factor 5a (C5a) is formed upon complement system activation in response to infection, injury or disease. Whilst C5a is a potent mediator of immune and inflammatory processes, excessive production or inadequate regulation of C5a has been implicated in the pathogenesis of numerous immuno-inflammatory diseases, predominantly through experimental studies utilising animal models of disease. Both acute and chronic conditions may benefit from C5a inhibition, including rheumatoid arthritis, inflammatory bowel disease, asthma, psoriasis, haemorrhagic shock and neurodegenerative conditions. The potentially broad clinical application for treatments that inhibit the activity of C5a at C5a receptors and the large global market for anti-inflammatory therapeutics have made C5a and the C5a receptor attractive targets for academic and commercial drug development programmes. in the past 5 years, interest in C5a as a drug target has grown substantially, and this activity has resulted in a collection of patents and scientific papers reporting novel C5a and C5a receptor inhibitors and antagonists, and generated a secondary stream of patent applications broadly claiming the use of C5/C5a inhibitors as a method of treating various immune and inflammatory conditions. This paper will review the physiology and pathophysiology of C5a and discuss the development of C5a and C5a receptor inhibitors in light of the recent scientific and patent literature.

Structure of saccharose-based carbon and transport of confined fluids: hybrid reverse Monte Carlo reconstruction and simulation studies

We present results of the reconstruction of a saccharose-based activated carbon (CS1000a) using hybrid reverse Monte Carlo (HRMC) simulation, recently proposed by Opletal et al. [1]. Interaction between carbon atoms in the simulation is modeled by an environment dependent interaction potential (EDIP) [2,3]. The reconstructed structure shows predominance of sp(2) over sp bonding, while a significant proportion of sp(3) hybrid bonding is also observed. We also calculated a ring distribution and geometrical pore size distribution of the model developed. The latter is compared with that obtained from argon adsorption at 87 K using our recently proposed characterization procedure [4], the finite wall thickness (FWT) model. Further, we determine self-diffusivities of argon and nitrogen in the constructed carbon as functions of loading. It is found that while there is a maximum in the diffusivity with respect to loading, as previously observed by Pikunic et al. [5], diffusivities in the present work are 10 times larger than those obtained in the prior work, consistent with the larger pore size as well as higher porosity of the activated saccharose carbon studied here.

The 2.1 angstrom crystal structure of copGFP, a representative member of the copepod clade within the green fluorescent protein superfamily

The green fluorescent protein (avGFP), its variants, and the closely related GFP-like proteins are characterized structurally by a cyclic tri-peptide chromophore located centrally within a conserved beta-can fold. Traditionally, these GFP family members have been isolated from the Cnidaria although recently, distantly related GFP-like proteins from the Bilateria, a sister group of the Cnidaria have been described, although no representative structure from this phylum has been reported to date. We have determined to 2.1 angstrom resolution the crystal structure of copGFP, a representative GFP-like protein from a copepod, a member of the Bilateria. The structure of copGFP revealed that, despite sharing only 19% sequence identity with GFP, the tri-peptide chromophore (Gly57-Tyr58-Gly59) of copGFP adopted a cis coplanar conformation within the conserved beta-can fold. However, the immediate environment surrounding the chromophore of copGFP was markedly atypical when compared to other members of the GFP-superfamily, with a large network of bulky residues observed to surround the chromophore. Arg87 and Glu222 (GFP numbering 96 and 222), the only two residues conserved between copGFP, GFP and GFP-like proteins are involved in autocatalytic genesis of the chromophore. Accordingly, the copGFP structure provides an alternative platform for the development of a new suite of fluorescent protein tools. Moreover, the structure suggests that the autocatalytic genesis of the chromophore is remarkably tolerant to a high degree of sequence and structural variation within the beta-can fold of the GFP superfamily. (c) 2006 Elsevier Ltd . All rights reserved.

The effect of auditory and visual distracters on the useful field of view: Implications for the driving task

PURPOSE. The driving environment is becoming increasingly complex, including both visual and auditory distractions within the in- vehicle and external driving environments. This study was designed to investigate the effect of visual and auditory distractions on a performance measure that has been shown to be related to driving safety, the useful field of view. METHODS. A laboratory study recorded the useful field of view in 28 young visually normal adults (mean 22.6 +/- 2.2 years). The useful field of view was measured in the presence and absence of visual distracters (of the same angular subtense as the target) and with three levels of auditory distraction (none, listening only, listening and responding). RESULTS. Central errors increased significantly (P < 0.05) in the presence of auditory but not visual distracters, while peripheral errors increased in the presence of both visual and auditory distracters. Peripheral errors increased with eccentricity and were greatest in the inferior region in the presence of distracters. CONCLUSIONS. Visual and auditory distracters reduce the extent of the useful field of view, and these effects are exacerbated in inferior and peripheral locations. This result has significant ramifications for road safety in an increasingly complex in-vehicle and driving environment.

The use of hypertonic saline for treating intracranial hypertension after traumatic brain injury

The past decade has witnessed a resurgence of interest in the use of hypertonic saline for low-volume resuscitation after trauma. Preliminary studies suggested that benefits are limited to a subgroup of trauma patients with brain injury, but a recent study of prehospital administration of hypertonic saline to patients with traumatic brain injury failed to confirm a benefit. Animal and human studies have demonstrated that hypertonic saline has clinically desirable physiological effects on cerebral blood flow, intracranial pressure, and inflammatory responses in models of neurotrauma. There are few clinical studies in traumatic brain injury with patient survival as an end point. In this review, we examined the experimental and clinical knowledge of hypertonic saline as an osmotherapeutic agent in neurotrauma.

Ultrastructure of the spermatozoon of Myrmecocichla formicivora (Vieillot, 1881) and Philetairus socius (Latham, 1790) (Aves; Passeriformes), with a new interpretation of the passeridan acrosome

Passerine spermatozoa exhibit apomorphies that distinguish them from non-passerine neognaths and palaeognaths. The acrosome is longer than the nucleus (excepting the suboscines, most Corvida, and a few Passerida). A perforatorium and endonuclear canals are absent. The proximal centriole is absent (except in the suboscines). The distal centriole is secondarily short, contrasting with its elongate condition in palaeognaths and Galloanserae. In the Passerida a single mitochondrial strand winds extensively along the axoneme (restricted to the anterior axoneme in suboscines and Corvida). A fibrous, or amorphous, periaxonemal sheath, seen in palaeognaths and many non-passerines, respectively, is absent. The acrosome in Myrmecocichla formicivora and Philetairus socius is bipartite: an acrosome core is surmounted by an acrosome crest; the core is ensheathed by a layer which is a posterior extension of the crest. The acrosome helix is a lateral extension of the crest and the crest layer with (Myrmecocichla) or without (Philetairus) protrusion of material of the acrosome core into it. In M. formicivora, as in other muscicapoids, a fibrous helix is intertwined with at least the more proximal region of the mitochondrial helix. The fibrous helix is absent at maturity in Philetairus and other described passeroid spermatozoa with the possible exception of Passer italiae. In Philetairus a granular helix precedes the mitochondrial helix.

Extending a model of shift-work tolerance

The present study contributes to theory and practice through the development of a model of shift-work tolerance with the potential to indicate interventions that reduce nurses' intention toward turnover and increase job satisfaction in hospital-based settings. Survey data from 1257 nurses were used to conduct structural equation modeling that examine the direct and indirect effects of supervisor and colleague support, team identity, team climate, and control over working environment on time-based work/life conflict, psychological well-being, physical symptoms, job satisfaction, and turnover intention. The analysis of the proposed model revealed a good fit The chi-square difference test was non-significant (χ2(26)=338.56), the fit indices were high (CFI=.923, NFI=.918, and NNFI=.868), the distribution of residuals was symmetric and approached zero, the average standardized residual was low (AASR=.04), and the standardized RMR was .072. In terms of the predictor variable, the final model explained 48% of the variance in turnover intention. The data revealed considerable evidence of both direct effects on adjustment and complex indirect links between levels of adjustment and work-related social support, team identity, team climate, and control. Nurses with high supervisor and coworker support experienced more positive team climates, identified more strongly with their team, and increased their perceptions of control over their work environment. This in turn lowered their appraisals of their time-based work/life conflict, which consequently increased their psychological well-being and job satisfaction and reduced their physical health symptoms and turnover intention. The type of shift schedule worked by the nurses influenced levels of turnover intention, control over work environment, time-based work/life conflict, and physical symptoms.