Endolysin of bacteriophage BFK20: evidence of a catalytic and a cell wall binding domain

A gene product of ORF24' was identified on the genome of corynephage BFK20 as a putative phage endolysin. The protein of endolysin BFK20 (gp24') has a modular structure consisting of an N-terminal amidase_2 domain (gp24CD) and a C-terminal cell wall binding domain (gp24BD). The C-terminal domain is unrelated to any of the known cell wall binding domains of phage endolysins. The whole endolysin gene and the sequences of its N-terminal and C-terminal domains were cloned; proteins were expressed in Escherichia coli and purified to homogeneity. The lytic activities of endolysin and its catalytic domain were demonstrated on corynebacteria and bacillus substrates. The binding activity of cell wall binding domain alone and in fusion with green fluorescent protein (gp24BD-GFP) were shown by specific binding assays to the cell surface of BFK20 host Brevibacterium flavum CCM 251 as well as those of other corynebacteria.     

The "far-west" of Anopheles gambiae molecular forms

The main Afrotropical malaria vector, Anopheles gambiae sensu stricto, is undergoing a process of sympatric ecological diversification leading to at least two incipient species (the M and S molecular forms) showing heterogeneous levels of divergence across the genome. The physically unlinked centromeric regions on all three chromosomes of these closely related taxa contain fixed nucleotide differences which have been found in nearly complete linkage disequilibrium in geographic areas of no or low M-S hybridization. Assays diagnostic for SNP and structural differences between M and S forms in the three centromeric regions were applied in samples from the western extreme of their range of sympatry, the only area where high frequencies of putative M/S hybrids have been reported. The results reveal a level of admixture not observed in the rest of the range. In particular, we found: i) heterozygous genotypes at each marker, although at frequencies lower than expected under panmixia; ii) virtually all possible genotypic combinations between markers on different chromosomes, although genetic association was nevertheless detected; iii) discordant M and S genotypes at two X-linked markers near the centromere, suggestive of introgression and inter-locus recombination. These results could be indicative either of a secondary contact zone between M and S, or of the maintenance of ancestral polymorphisms. This issue and the perspectives opened by these results in the study of the M and S incipient speciation process are discussed.     

Lipid mapping in human dystrophic muscle by cluster-time-of-flight secondary ion mass spectrometry imaging

Human striated muscle samples, from male control and Duchenne muscular dystrophy-affected children, were subjected to cluster-time-of-flight secondary ion mass spectrometry (cluster-ToF-SIMS) imaging using a 25 keV Bi(3)(+) liquid metal ion gun under static SIMS conditions. Spectra and ion density maps, or secondary ion images, were acquired in both positive and negative ion mode over several areas of 500 x 500 microm(2) (image resolution, 256 x 256 pixels). Characteristic distributions of various lipids were observed. Vitamin E and phosphatidylinositols were found to concentrate within the cells, whereas intact phosphocholines accumulated over the most damaged areas of the dystrophic muscles, together with cholesterol and sphingomyelin species. Fatty acyl chain composition varied depending on the region, allowing estimation of the local damage extent.     

Pyrimidinone-peptoid hybrid molecules with distinct effects on molecular chaperone function and cell proliferation

The Hsp70 molecular chaperones are ATPases that play critical roles in the pathogenesis of many human diseases, including breast cancer. Hsp70 ATP hydrolysis is relatively weak but is stimulated by J domain-containing proteins. We identified pyrimidinone-peptoid hybrid molecules that inhibit cell proliferation with greater potency than previously described Hsp70 modulators. In many cases, anti-proliferative activity correlated with inhibition of J domain stimulation of Hsp70.     

Development and characterization of a FIA system for selective assay of l-ascorbic acid in food samples

An amperometric detector and an enzymatic reaction were combined for the measurement of l-ascorbic acid. The enzyme cell (containing immobilized ascorbate oxidase) was connected to a flow injection analyzer (FIA) system with a glassy carbon electrode as an amperometric detector. During optimization and measurements two sample injectors were used, one before and one after the enzyme cell, thus eliminating the background interferences. Subtraction of the signal area given in the presence of enzyme from the one given in the absence of enzyme was applied for measuring analyte concentrations and calibration at 400 mV. Analysis capacity of system is 25 samples/hour. The relative standard deviation (RSD) was below 5% (5 times repeated, 400 μmol/L conc.), linearity up to 400 μmol/L, limit of detection (LOD) 5 μmol/L, fitting of calibration curve in 25–400 μmol/L range was R ² = 0.99.     

Addiction: Decreased reward sensitivity and increased expectation sensitivity conspire to overwhelm the brain's control circuit

Based on brain imaging findings, we present a model according to which addiction emerges as an imbalance in the information processing and integration among various brain circuits and functions. The dysfunctions reflect (a) decreased sensitivity of reward circuits, (b) enhanced sensitivity of memory circuits to conditioned expectations to drugs and drug cues, stress reactivity, and (c) negative mood, and a weakened control circuit. Although initial experimentation with a drug of abuse is largely a voluntary behavior, continued drug use can eventually impair neuronal circuits in the brain that are involved in free will, turning drug use into an automatic compulsive behavior. The ability of addictive drugs to co‐opt neurotransmitter signals between neurons (including dopamine, glutamate, and GABA) modifies the function of different neuronal circuits, which begin to falter at different stages of an addiction trajectory. Upon exposure to the drug, drug cues or stress this results in unrestrained hyperactivation of the motivation/drive circuit that results in the compulsive drug intake that characterizes addiction.     

Mu Opioid Receptor Binding Correlates with Nicotine Dependence and Reward in Smokers

The rewarding effects of nicotine are associated with activation of nicotine receptors. However, there is increasing evidence that the endogenous opioid system is involved in nicotine''s rewarding effects. We employed PET imaging with [¹¹C]carfentanil to test the hypotheses that acute cigarette smoking increases release of endogenous opioids in the human brain and that smokers have an upregulation of mu opioid receptors (MORs) when compared to nonsmokers. We found no significant changes in binding potential (BP_ND) of [¹¹C]carfentanil between the placebo and the active cigarette sessions, nor did we observe differences in MOR binding between smokers and nonsmokers. Interestingly, we showed that in smokers MOR availability in bilateral superior temporal cortices during the placebo condition was negatively correlated with scores on the Fagerström Test for Nicotine Dependence (FTND). Also in smokers, smoking-induced decreases in [¹¹C]carfentanil binding in frontal cortical regions were associated with self-reports of cigarette liking and wanting. Although we did not show differences between smokers and nonsmokers, the negative correlation with FTND corroborates the role of MORs in superior temporal cortices in nicotine addiction and provides preliminary evidence of a role of endogenous opioid signaling in frontal cortex in nicotine reward.     

Detection of Atomic Scale Changes in the Free Volume Void Size of Three-Dimensional Colorectal Cancer Cell Culture Using Positron Annihilation Lifetime Spectroscopy

Positron annihilation lifetime spectroscopy (PALS) provides a direct measurement of the free volume void sizes in polymers and biological systems. This free volume is critical in explaining and understanding physical and mechanical properties of polymers. Moreover, PALS has been recently proposed as a potential tool in detecting cancer at early stages, probing the differences in the subnanometer scale free volume voids between cancerous/healthy skin samples of the same patient. Despite several investigations on free volume in complex cancerous tissues, no positron annihilation studies of living cancer cell cultures have been reported. We demonstrate that PALS can be applied to the study in human living 3D cell cultures. The technique is also capable to detect atomic scale changes in the size of the free volume voids due to the biological responses to TGF-β. PALS may be developed to characterize the effect of different culture conditions in the free volume voids of cells grown in vitro.     

Relationships between Degree of Polymerization and Antioxidant Activities: A Study on Proanthocyanidins from the Leaves of a Medicinal Mangrove Plant Ceriops tagal

Tannins from the leaves of a medicinal mangrove plant, Ceriops tagal, were purified and fractionated on Sephadex LH-20 columns. ¹³C nuclear magnetic resonance (¹³C-NMR), reversed/normal high performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI MS) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDT-TOF MS) analysis showed that the tannins were predominantly B-type procyanidins with minor A-type linkages, galloyl and glucosyl substitutions, and a degree of polymerization (DP) up to 33. Thirteen subfractions of the procyanidins were successfully obtained by a modified fractionation method, and their antioxidant activities were investigated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging capacity and ferric reducing antioxidant power (FRAP) method. All these subfractions exhibited potent antioxidant activities, and eleven of them showed significantly different mean DP (mDP) ranging from 1.43±0.04 to 31.77±1.15. Regression analysis demonstrated that antioxidant activities were positively correlative with mDP when around mDP <10, while dropped and then remained at a level similar to mDP = 5 with around 95 µg ml⁻¹ for DPPH scavenging activity and 4 mmol AAE g⁻¹ for FRAP value.