13C-Methyl isocyanide as an NMR probe for cytochrome P450 active site

The cytochromes P450 (CYPs) play a central role in many biologically important oxidation reactions, including the metabolism of drugs and other xenobiotic compounds. Because they are often assayed as both drug targets and anti-targets, any tools that provide: (a) confirmation of active site binding and (b) structural data, would be of great utility, especially if data could be obtained in reasonably high throughput. To this end, we have developed an analog of the promiscuous heme ligand, cyanide,with a 13CH3-reporter attached. This 13C-methyl isocyanide ligand binds to bacterial (P450cam) and membrane-bound mammalian (CYP2B4) CYPs. It can be used in a rapid 1D experiment to identify binders, and provides a qualitative measure of structural changes in the active site

Classification of Facial Expressions based on Transitions Derived from Third Order Neighborhood LBP

The present paper extended the LBP transitions derived from second-order neighbourhood on to third order neighbourhood LBP (TN-LBP) and derived transitions on Trapezoid patterns for facial expression classification. The TN-LBP forms four Trapezoid Patterns (TP) i.e. top left, bottom right and top right, bottom left. So far no researcher carried out work on classification problem based on transitions on third-order neighborhood LBP. The present paper derived transitions on the two reciprocal 201C;Trapezoids of TN-LBP (T-TN-LBP) i.e. top left vs. bottom right. Each of these Trapezoids on TN-LBP will have five pixies and each of them will have 25 i.e 32 patterns. The present paper derived transitions on two symmetric T-TN-LBP. Based on this, facial expression recognition algorithm is built. The proposed approach is compared with the existing methods

Stone Image Classification Based on Overlapped 5-bit T-Patterns occurrence on 5-by-5 Sub Images

Texture classification is widely used in understanding the visual patterns and has wide range of applications. The present paper derived a novel approach to classify the stone textures based on the patterns occurrence on each sub window. The present approach identifies overlapped nine 5 bit T-patterns (O5TP) on each 5×5 sub window stone image. Based the number of occurrence of T-patterns count the present paper classify the stone images into any of the four classes i.e. brick, granite, marble and mosaic stone images.  The novelty of the present approach is that no standard classification algorithm is used for the classification of stone images. The proposed method is experimented on Mayang texture images, Brodatz textures, Paul Bourke color images, VisTex database, Google color stone texture images and also original photo images taken by digital camera. The outcome of the results indicates that the proposed approach percentage of grouping performance is higher to that of many existing approaches

Experimental study and analysis of lubricants dispersed with nano Cu and TiO2 in a four-stroke two wheeler

The present investigation summarizes detailed experimental studies with standard lubricants of commercial quality known as Racer-4 of Hindustan Petroleum Corporation (India) dispersed with different mass concentrations of nanoparticles of Cu and TiO2. The test bench is fabricated with a four-stroke Hero-Honda motorbike hydraulically loaded at the rear wheel with proper instrumentation to record the fuel consumption, the load on the rear wheel, and the linear velocity. The whole range of data obtained on a stationery bike is subjected to regression analysis to arrive at various relationships between fuel consumption as a function of brake power, linear velocity, and percentage mass concentration of nanoparticles in the lubricant. The empirical relation correlates with the observed data with reasonable accuracy. Further, extension of the analysis by developing a mathematical model has revealed a definite improvement in brake thermal efficiency which ultimately affects the fuel economy by diminishing frictional power in the system with the introduction of nanoparticles into the lubricant. The performance of the engine seems to be better with nano Cu-Racer-4 combination than the one with nano TiO2

Moderate Traumatic Brain Injury Causes Acute Dendritic and Synaptic Degeneration in the Hippocampal Dentate Gyrus

Hippocampal injury-associated learning and memory deficits are frequent hallmarks of brain trauma and are the most enduring and devastating consequences following traumatic brain injury (TBI). Several reports, including our recent paper, showed that TBI brought on by a moderate level of controlled cortical impact (CCI) induces immature newborn neuron death in the hippocampal dentate gyrus. In contrast, the majority of mature neurons are spared. Less research has been focused on these spared neurons, which may also be injured or compromised by TBI. Here we examined the dendrite morphologies, dendritic spines, and synaptic structures using a genetic approach in combination with immunohistochemistry and Golgi staining. We found that although most of the mature granular neurons were spared following TBI at a moderate level of impact, they exhibited dramatic dendritic beading and fragmentation, decreased number of dendritic branches, and a lower density of dendritic spines, particularly the mushroom-shaped mature spines. Further studies showed that the density of synapses in the molecular layer of the hippocampal dentate gyrus was significantly reduced. The electrophysiological activity of neurons was impaired as well. These results indicate that TBI not only induces cell death in immature granular neurons, it also causes significant dendritic and synaptic degeneration in pathohistology. TBI also impairs the function of the spared mature granular neurons in the hippocampal dentate gyrus. These observations point to a potential anatomic substrate to explain, in part, the development of posttraumatic memory deficits. They also indicate that dendritic damage in the hippocampal dentate gyrus may serve as a therapeutic target following TBI

A method for increasing electroporation competence of Gram-negative clinical isolates by polymyxin B nonapeptide

The study of clinically relevant bacterial pathogens relies on molecular and genetic approaches. However, the generally low transformation frequency among natural isolates poses technical hurdles to widely applying common methods in molecular biology, including transformation of large constructs, chromosomal genetic manipulation, and dense mutant library construction. Here we demonstrate that culturing clinical isolates in the presence of polymyxin B nonapeptide (PMBN) improves their transformation frequency via electroporation by up to 100-fold in a dose-dependent and reversible manner. The effect was observed for PMBN-binding uropathogenic Escherichia coli (UPEC) and Salmonella enterica strains but not naturally polymyxin resistant Proteus mirabilis. Using our PMBN electroporation method we show efficient delivery of large plasmid constructs into UPEC, which otherwise failed using a conventional electroporation protocol. Moreover, we show a fivefold increase in the yield of engineered mutant colonies obtained in S. enterica with the widely used lambda-Red recombineering method, when cells are cultured in the presence of PMBN. Lastly, we demonstrate that PMBN treatment can enhance the delivery of DNA-transposase complexes into UPEC and increase transposon mutant yield by eightfold when constructing Transposon Insertion Sequencing (TIS) libraries. Therefore, PMBN can be used as a powerful electropermeabilisation adjuvant to aid the delivery of DNA and DNA–protein complexes into clinically important bacteria.</p

Transposon-Directed Insertion-Site Sequencing Reveals Glycolysis Gene gpmA as Part of the H2O2 Defense Mechanisms in Escherichia coli

Hydrogen peroxide (H2O2) is a common effector of defense mechanisms against pathogenic infections. However, bacterial factors involved in H2O2tolerance remain unclear. Here we used transposon-directed insertion-site sequencing (TraDIS), a technique allowing the screening of the whole genome, to identify genes implicated in H2O2tolerance inEscherichia coli. Our TraDIS analysis identified 10 mutants with fitness defect upon H2O2exposure, among which previously H2O2-associated genes (oxyR,dps,dksA,rpoS,hfqandpolA) and other genes with no known association with H2O2tolerance inE. coli(corA,rbsR,nhaAandgpmA). This is the first description of the impact ofgpmA, a gene involved in glycolysis, on the susceptibility ofE. colito H2O2. Indeed, confirmatory experiments showed that the deletion ofgpmAled to a specific hypersensitivity to H2O2comparable to the deletion of the major H2O2scavenger genekatG. This hypersensitivity was not due to an alteration of catalase function and was independent of the carbon source or the presence of oxygen. Transcription ofgpmAwas upregulated under H2O2exposure, highlighting its role under oxidative stress. In summary, our TraDIS approach identifiedgpmAas a member of the oxidative stress defense mechanism inE. coli