Modular and scalable load-wall sled buck for pure-lateral and oblique side impact tests

A considerable majority of side impact sled tests using different types of human surrogates has used a load-wall design not specific to subject anthropometry. The use of one load-wall configuration cannot accurately isolate and evaluate regional responses for the same load-wall geometry. As the anatomy and biomechanical responses of the human torso depends on the region, and anthropomorphic test devices continue to advance and accommodate regional differences, it is important to obtain specific data from sled tests. To achieve this goal, the present study designed a scalable modular load-wall consisting of the shoulder, thorax, abdomen, and superior and inferior pelvis, and lower limb plates. The first five plates were connected to a vertical fixture and the limb plate was connected to another fixture. The width, height, and thickness, and the gap between plates were modular. Independent adjustments in the coronal and sagittal planes allowed region-specific positioning depending on surrogate anthropometry, example pelvis width and seated height. Two tri-axial load cells were fixed on the contralateral face of each plate of the load-wall to record impact force-time histories. The load-wall and vertical fixture design can be used to conduct side impact tests with varying vectors, pure-lateral to anterior and posterior oblique, by appropriately orienting the load-wall with respect to the surrogate. The feasibility of the design to extract region-specific biomechanical data was demonstrated by conducting pure-lateral and anterior oblique sled tests using two different surrogates at a velocity of 6.7m/s. Uses of this design are discussed for different applications.     

Transglycosylation by Chitinase D from Serratia proteamaculans Improved through Altered Substrate Interactions

We describe the improvement of transglycosylation (TG) by chitinase D from Serratia proteamaculans (SpChiD). The SpChiD produced a smaller quantity of TG products for up to 90 min with 2 mm chitotetraose as the substrate and subsequently produced only hydrolytic products. Of the five residues targeted at the catalytic center, E159D resulted in substantial loss of both hydrolytic and TG activities. Y160A resulted in a product profile similar to SpChiD and a rapid turnover of substrate with slightly increased TG activity. The rest of the three mutants, M226A, Y228A, and R284A, displayed improved TG and decreased hydrolytic ability. Four of the five amino acid substitutions, F64W, F125A, G119S, and S116G, at the catalytic groove increased TG activity, whereas W120A completely lost the TG activity with a concomitant increase in hydrolysis. Mutation of Trp-247 at the solvent-accessible region significantly reduced the hydrolytic activity with increased TG activity. The mutants M226A, Y228A, F125A, S116G, F64W, G119S, R284A, and W247A accumulated approximately double the concentration of TG products like chitopentaose and chitohexaose, compared with SpChiD. The double mutant E159D/F64W regained the activity with accumulation of 6.0% chitopentaose at 6 h, similar to SpChiD at 30 min. Loss of chitobiase activity was unique to Y228A. Substitution of amino acids at the catalytic center and/or groove substantially improved the TG activity of SpChiD, both in terms of the quantity of TG products produced and the extended duration of TG activity.     

Characterization of temperature inducible promoters from a novel rolling circle replicating plasmid of Enterococcus faecium DJ1

Limited studies have been performed on the characterization of small size plasmids of Enterococcus faecium with the intention of evaluating the strength of their promoters in Escherichia coli. The complete nucleotide sequence (3.825 Kb) and structural organization of E. faecium DJ1 cryptic plasmid pNJAKD is presented. Seven promoter sequences from the pNJAKD plasmid of E. faecium have been identified. The regions coding for the putative promoters were either amplified using PCR based techniques or chemically synthesized as oligonucleotides of different sizes. These were subsequently cloned in the pEGFP vector at the Pvu II site. The efficiency of putative promoter fragments were measured using the intensity of eGFP fluorescence in E. coli JM101, DH5α and BL21(DE3), among which AKD3 exhibited moderate to strongest promoter activity at temperatures of 30, 37, and 42°C.     

Photosynthesis and yield in cotton (Gossypium hirsutum L.) Var. Vikram after exclusion of ambient solar UV-B/A

The impact of ambient solar UV was studied on the photosynthesis and yield of cotton (Gossypium hirsutum) var. Vikram in a field experiment by excluding either UV-B (<315 nm) or UV-B/A (<400 nm) components of solar spectrum. Cotton plants were grown in cages covered with polyester filters that could specifically cut off UV-B or UV-B/A part of the solar spectrum. The control plants were grown under a filter transmissible to UV. Exclusion of UV enhanced plant height, leaf area, total biomass, and the yield parameters (number and weight of bolls, length of fiber and number of seeds) of cotton. Enhancement in the vegetative growth and yield of the plants could be related to enhanced rate of photosynthesis in the leaves. Polyphasic chlorophyll a fluorescence (OJIP) transients from UV excluded plants gave a higher fluorescence yield at I–P phase. Fluorescence measurements indicated enhanced F _v/F _m ratio and reduction capacity after exclusion of solar UV. Exclusion also enhanced stomatal conductance and intercellular CO₂ concentration and reduced the stomatal resistance. Total soluble proteins were higher after UV exclusion, and in SDS–PAGE analysis, bands corresponding to smaller subunits (14 kDa) of Rubisco were more intensely stained. Experiments indicated suppressive action of ambient UV on carbon fixation and yield of cotton plants. Exclusion of solar UV proved to be beneficial in enhancing the yield of cotton plants.     

Spermidine and abscisic acid-mediated phosphorylation of a cytoplasmic protein from rice root in response to salinity stress

Importance of higher polyamines, spermidine, and spermine, in relation to the mechanism and adaptation to combat abiotic stress has been well established in cereals. Owing to their polycationic nature at physiological pH, polyamines bind strongly to negative charges in cellular components such as nucleic acids, various proteins, and phospholipids. To study the physiological role of polyamine during salinity stress, phosphorylation study was carried out in cytosolic soluble protein fraction isolated from the roots of salt tolerant (Nonabokra) and salt sensitive (M-1-48) rice cultivars treated with none (control), NaCl (150 mM, 16 h), spermidine (1 mM, 16 h) or with abscisic acid (100 μM, 16 h). A calcium independent auto regulatory 42 kDa protein kinase was found to phosphorylate myelin basic protein and casein but not histone. Interestingly, this was the only protein to be phosphorylated in root cytosolic fraction during NaCl/abscisic acid/spermidine treatment indicating its importance in salinity mediated signal transduction. This is the first report of polyamine as well as abscisic acid induced protein kinase activity in rice root in response to salinity stress.     

Antitumor potential of Castanopsis indica (Roxb. ex Lindl.) A. DC. leaf extract against Ehrlich's ascites carcinoma cell

Methanol extract of C. indica (MECI) leaves showed direct cytotoxicity on Ehrlich ascites carcinoma (EAC) cell in a dose dependant manner and there was significant decrease in the tumor volume, viable cell count, tumor weight and elevated the life span of EAC tumor bearing mice. Hematological profile and biochemical estimations were significantly restored to normal levels in MECI treated as compared to EAC control mice. MECI treatment significantly modulated the tissue antioxidant assay parameters as compared to the EAC control mice. The results revealed that MECI possesses significant dose dependent antitumor potential which may be due to its cytotoxicity and antioxidant properties.     

Solvent-induced tuning of internal structure in a protein amyloid protofibril

An important goal in studies of protein aggregation is to obtain an understanding of the structural diversity that is characteristic of amyloid fibril and protofibril structures at the molecular level. In this study, what to our knowledge are novel assays based on time-resolved fluorescence anisotropy decay and dynamic quenching measurements of a fluorophore placed at different specific locations in the primary structure of a small protein, barstar, have been used to determine the extent to which the protein sequence participates in the structural core of protofibrils. The fluorescence measurements reveal the structural basis of how modulating solvent polarity results in the tuning of the protofibril conformation from a pair of parallel β-sheets in heat-induced protofibrils to a single parallel β-sheet in trifluorethanol-induced protofibrils. In trifluorethanol-induced protofibrils, the single β-sheet is shown to be built up from in-register β-strands formed by nearly the entire protein sequence, while in heat-induced protofibrils, the pair of β-sheets motif is built up from β-strands formed by only the last two-third of the protein sequence.     

Transcriptional activation by Oct4 is sufficient for the maintenance and induction of pluripotency

Oct4 is an essential regulator of pluripotency in vivo and in vitro in embryonic stem cells, as well as a key mediator of the reprogramming of somatic cells into induced pluripotent stem cells. It is not known whether activation and/or repression of specific genes by Oct4 is relevant to these functions. Here, we show that fusion proteins containing the coding sequence of Oct4 or Xlpou91 (the Xenopus homolog of Oct4) fused to activating regions, but not those fused to repressing regions, behave as Oct4, suppressing differentiation and promoting maintenance of undifferentiated phenotypes in vivo and in vitro. An Oct4 activation domain fusion supported embryonic stem cell self-renewal in vitro at lower concentrations than that required for Oct4 while alleviating the ordinary requirement for the cytokine LIF. At still lower levels of the fusion, LIF dependence was restored. We conclude that the necessary and sufficient function of Oct4 in promoting pluripotency is to activate specific target genes.     

Unlocking the potential of metagenomics through replicated experimental design

Metagenomics holds enormous promise for discovering novel enzymes and organisms that are biomarkers or drivers of processes relevant to disease, industry and the environment. In the past two years, we have seen a paradigm shift in metagenomics to the application of cross-sectional and longitudinal studies enabled by advances in DNA sequencing and high-performance computing. These technologies now make it possible to broadly assess microbial diversity and function, allowing systematic investigation of the largely unexplored frontier of microbial life. To achieve this aim, the global scientific community must collaborate and agree upon common objectives and data standards to enable comparative research across the Earth's microbiome. Improvements in comparability of data will facilitate the study of biotechnologically relevant processes, such as bioprospecting for new glycoside hydrolases or identifying novel energy sources.