Comparison of Segger and other methods for segmentation and rigid-body docking of molecular components in cryo-EM density maps

Segmentation and docking are useful methods for the discovery of molecular components in electron cryo-microscopy (cryo-EM) density maps of macromolecular complexes. In this article, we describe the segmentation and docking methods implemented in Segger. For 11 targets posted in the 2010 cryo-EM challenge, we segmented the regions corresponding to individual molecular components using Segger. We then used the segmented regions to guide rigid-body docking of individual components. Docking results were evaluated by comparing the docked components with published structures, and by calculation of several scores, such as atom inclusion, density occupancy, and geometry clash. The accuracy of the component segmentation using Segger and other methods was assessed by comparing segmented regions with docked components.     

Magnesium Sulfate Protects Against the Bioenergetic Consequences of Chronic Glutamate Receptor Stimulation

Extracellular glutamate is elevated following brain ischemia or trauma and contributes to neuronal injury. We tested the hypothesis that magnesium sulfate (MgSO₄, 3 mM) protects against metabolic failure caused by excitotoxic glutamate exposure. Rat cortical neuron preparations treated in medium already containing a physiological concentration of Mg²⁺ (1 mM) could be segregated based on their response to glutamate (100 µM). Type I preparations responded with a decrease or small transient increase in oxygen consumption rate (OCR). Type II neurons responded with >50% stimulation in OCR, indicating a robust response to increased energy demand without immediate toxicity. Pre-treatment with MgSO₄ improved the initial bioenergetic response to glutamate and ameliorated subsequent loss of spare respiratory capacity, measured following addition of the uncoupler FCCP, in Type I but not Type II neurons. Spare respiratory capacity in Type I neurons was also improved by incubation with MgSO₄ or NMDA receptor antagonist MK801 in the absence of glutamate treatment. This finding indicates that the major difference between Type I and Type II preparations is the amount of endogenous glutamate receptor activity. Incubation of Type II neurons with 5 µM glutamate prior to excitotoxic (100 µM) glutamate exposure recapitulated a Type I phenotype. MgSO₄ protected against an excitotoxic glutamate-induced drop in neuronal ATP both with and without prior 5 µM glutamate exposure. Results indicate that MgSO₄ protects against chronic moderate glutamate receptor stimulation and preserves cellular ATP following treatment with excitotoxic glutamate.     

An unusual brachiopod assemblage in a Late Jurassic (Kimmeridgian) stromatactis mud-mound of the Eastern Carpathians (Hăghimaş Mountains), Romania

An unusual fossil assemblage dominated by superabundant rhynchonellid brachiopods in a stromatactis mud-mound is recorded from the Hăghimaş Mountains (Eastern Carpathians), Romania. The mound mainly consists of bioclastic wackestones to packstones with a very rich macrofauna including crinoids, sponges, juvenile ammonites, and echinoids. The brachiopods represent a low-diversity but high-abundance association, dominated by the rhynchonellids Lacunosella and Septaliphoria. The taphonomical features of the fossil assemblage indicate an autochthonous fauna, with successive generations of brachiopods in life position and complete well-preserved individuals in different growth stage alongside an accessory population of crinoids and sponges. Brachiopod-brachiopod endosymbiotic life strategy is documented for the first time from a post-Paleozoic brachiopod assemblage. The mound reveals abundant stromatactis, filled by radiaxial fibrous or drusy calcite cement and internal polymud sediments. This is the first Late Jurassic (Kimmeridgian) stromatactis mud-mound identified in the Eastern Carpathians.     

Methodology optimization and diversification for the investigation of virulence potential in Haemophilus influenzae clinical strains

Ten Haemophilus influenzae strains were isolated from patients aged between 1.6 - 24 years, with various diagnoses (acute meningitis, acute upper respiratory infection, otitis media and acute sinusitis). Identification was based on phenotypic and molecular characteristics; antibiotic susceptibility testing was performed by diffusion method according to CLSI standards 2011 for seven antibiotics. The results of molecular testing showed that all the studied strains produced an amplicon of 1000 bp with ompP2 primers indicating that all strains were H. influenzae. For six strains, the PCR amplicon obtained with bexA specific primers, proving that the strains were capsulated. The results of phenotypic testing showed that four strains were ampicillin nonsusceptible and (beta-lactamase-positive. The virulence potential of H. influenzae clinical strains was investigated by phenotypic methods, including the assessment of the soluble virulence factors on specific media containing the biochemical substratum for the investigated enzymatic factor, as well as the adherence and invasion capacity to HeLa cells monolayer using Cravioto modified method. The studied strains exhibited mainly a diffuse adherence pattern and different adherence indexes. Interestingly, two strains isolated from the same pacient (blood and CSF) showed a different degree of invasiveness, the strain isolated from blood being 20 times more invasive than the one isolated from CSF.     

Association between OLR1 K167N SNP and intima media thickness of the common carotid artery in the general population

Background and Purpose

The lectin-like oxidised LDL receptor-1 (OLR1) gene encodes a scavenger receptor implicated in the pathogenesis of atherosclerosis. Although functional roles have been suggested for two variants, epidemiological studies on OLR1 have been inconsistent. Methods - We tested the association between the non-synonymous substitution K167N (rs11053646) and intima media thickness of the common carotid artery (CCA-IMT) in 2,141 samples from the Progression of Lesions in the Intima of the Carotid (PLIC) study (a prospective population-based study).

Results

Significantly increased IMT was observed in male carriers of the minor C (N) allele compared to GC and GG (KN and KK) genotype. Functional analysis on macrophages suggested a decreased association to Ox-LDL in NN carriers compared to KN and KK carriers which is also associated with a reduced OLR1 mRNA expression. Macrophages from NN carriers present also a specific inflammatory gene expression pattern compared to cells from KN and KK carriers.

Conclusions

These data suggest that the 167N variant of LOX-1 receptor affects the atherogenic process in the carotid artery prior to evidence of disease through an inflammatory process.     

In vitro susceptibility of staphylococci to linezolid and other antimicrobial agents

Linezolid is a member of the new class of antibacterial agents called oxazolidinones that are active against Gram positive organisms and exert their action by protein synthesis inhibition. In this study we investigated the in vitro activity of linezolid versus the other agent against clinical strains of staphylococci: Staphylococcus aureus (n = 82) and S. epidermidis (n = 32) collected in 2002 from hospitalized patients and healthy individuals, isolated from different biological samples. Agar dilution minimum inhibitory concentrations (MICs) were determined by using Mueller-Hinton agar according to the guidelines established by the National Committee for Clinical Laboratory Standards. Linezolid demonstrated excellent in vitro activity against all isolates tested, with MICs values in the range of susceptibility (< or = 8 microg/ml). No associated resistance between linezolid and other agents tested was observed. The resistance among Gram positive bacteria continues to spread and for many patients infected with these resistant organisms antimicrobial therapy is ineffective and linezolid may be a new alternative treatment.     

Stability and instability properties of aggregation of single chain amphiphiles into binary mixtures

In a lysophospholipid binary mixture, there are three ways of association between the mixture components of single-chain amphiphiles: (a) between two identical molecules each of the first and second component (self-association process) and (b) between two different molecules (cross-association process). Association probabilities for three binary mixtures were analysed as functions depending on the electric dipole moments of the polar head groups. A 3-D view representation is most suitable for this analysis. The most important finding is that for certain values of the electric dipole moments there are molecular couples which have a maximum stability to the changes in the external electrolytic medium. This fact confirms the formation of clusters and their stability, which is equivalent to the existence of micro-heterogeneities within the lipid bilayers. On the other hand, there are unstable molecular associations, and this fact influences the appearance of some phase transitions. Generally, the increase of the electric dipole moment or the increase of the acyl-chain length of one component from a binary lipid mixture decreases the self-association probability between its own molecules, but it increases the self-association probability of the other mixture components. Furthermore, the cross-association probability has high values for any binary lipid mixture of single-chain amphiphiles.     

Electron Cryo-microscopy Structure of Ebola Virus Nucleoprotein Reveals a Mechanism for Nucleocapsid-like Assembly

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Comparative In Vivo Effects of Hemoglobin-Based Oxygen Carriers (HBOC) with Varying Prooxidant and Physiological Reactivity

A series of hemoglobin-based oxygen carrier candidates (HBOC), previously noted for their differences in prooxidative and physiological reactivity, were compared in terms of the negative effects displayed upon injection in Wistar rats. At the concentrations tested, antioxidant strategies based on albumin as well as based on rubrerythrin appear to offer observable physiological advantages.