Relationship between stability and flexibility in the most flexible region of Photinus pyralis luciferase

Firefly luciferase is a protein with a large N-terminal and a small C-terminal domain. B-factor analysis shows that its C-terminal is much more flexible than its N-terminal. Studies on hyperthermophile proteins have been shown that the increased thermal stability of hyperthermophile proteins is due to their enhanced conformational rigidity and the relationship between flexibility, stability and function in most of proteins is on debate. Two mutations (D474K and D476N) in the most flexible region of firefly luciferase were designed. Thermostability analysis shows that D476N mutation doesn't have any significant effect but D474K mutation destabilized protein. On the other hand, flexibility analysis using dynamic quenching and limited proteolysis demonstrates that D474K mutation became much more flexible than wild type although D476N doesn't have any significant difference. Intrinsic and ANS fluorescence studies demonstrate that D476N mutation is brought about by structural changes without significant effect on thermostability and flexibility. Molecular modeling reveals that disruption of a salt bridge between D(474) and K(445) accompanying with some H-bond deletion may be involved in destabilization of D474K mutant.     

On Comparison Between Fungal and Bacterial Pretreatment of Coal for Enhanced Biogenic Methane Generation

Owing to better understanding of subsurface geochemical carbon recycling and real-time active methanogenesis in major coal basins around the globe, substantial share of subsurface methane generation is attributed to biogenic origin. Since coal, being complex geopolymer, does not appear to be a favorable microbial substrate, enhancement in biogenic methane yield depends on its degradation into simpler organic substrates. This review puts forward a comparative analysis of fungal and bacterial pretreatment for determining the extent of facilitation in initial degradation of coal, which is still rate limiting step in overall conversion of coal into methane. Primarily, the initial fungal degradation of coal differs from bacterial pretreatment of coal in terms of the nature of released organics. On the basis of previous reports, fungal pretreatment of coal yields, majorly, polyaromatic hydrocarbons, however, bacterial pretreatment results in the generation of mixed organics pool of aromatics and aliphatics. The presence of aliphatics may be prospected for achieving greater conversion rates of coal conversion into methane. Considering the criticality of preliminary degradation of coal and associated issues, the fate of commercial biogenic methane generation would be dictated by the factors pertaining to geological considerations and reservoir geology, chemistry of coal and associated water tables, geomicrobial considerations and economic viability.     

Increased Thymic Cell Turnover under Boron Stress May Bypass TLR3/4 Pathway in African Ostrich

Previous studies revealed that thymus is a targeted immune organ in malnutrition, and high-boron stress is harmful for immune organs. African ostrich is the living fossil of ancient birds and the food animals in modern life. There is no report about the effect of boron intake on thymus of ostrich. The purpose of present study was to evaluate the effect of excessive boron stress on ostrich thymus and the potential role of TLR3/4 signals in this process. Histological analysis demonstrated that long-term boron stress (640 mg/L for 90 days) did not disrupt ostrich thymic structure during postnatal development. However, the numbers of apoptotic cells showed an increased tendency, and the expression of autophagy and proliferation markers increased significantly in ostrich thymus after boron treatment. Next, we examined the expression of TLR3 and TLR4 with their downstream molecular in thymus under boron stress. Since ostrich genome was not available when we started the research, we first cloned ostrich TLR3 TLR4 cDNA from thymus. Ostrich TLR4 was close to white-throated Tinamou. Whole avian TLR4 codons were under purify selection during evolution, whereas 80 codons were under positive selection. TLR3 and TLR4 were expressed in ostrich thymus and bursa of fabricius as was revealed by quantitative real-time PCR (qRT-PCR). TLR4 expression increased with age but significantly decreased after boron treatment, whereas TLR3 expression showed the similar tendency. Their downstream molecular factors (IRF1, JNK, ERK, p38, IL-6 and IFN) did not change significantly in thymus, except that p100 was significantly increased under boron stress when analyzed by qRT-PCR or western blot. Taken together, these results suggest that ostrich thymus developed resistance against long-term excessive boron stress, possibly by accelerating intrathymic cell death and proliferation, which may bypass the TLR3/4 pathway. In addition, attenuated TLRs activity may explain the reduced inflammatory response to pathogens under boron stress.     

Density,Destinations or Both? A Comparison of Measures of Walkability in Relation to Transportation Behaviors,Obesity and Diabetes in Toronto,Canada

The design of suburban communities encourages car dependency and discourages walking, characteristics that have been implicated in the rise of obesity. Walkability measures have been developed to capture these features of urban built environments. Our objective was to examine the individual and combined associations of residential density and the presence of walkable destinations, two of the most commonly used and potentially modifiable components of walkability measures, with transportation, overweight, obesity, and diabetes. We examined associations between a previously published walkability measure and transportation behaviors and health outcomes in Toronto, Canada, a city of 2.6 million people in 2011. Data sources included the Canada census, a transportation survey, a national health survey and a validated administrative diabetes database. We depicted interactions between residential density and the availability of walkable destinations graphically and examined them statistically using general linear modeling. Individuals living in more walkable areas were more than twice as likely to walk, bicycle or use public transit and were significantly less likely to drive or own a vehicle compared with those living in less walkable areas. Individuals in less walkable areas were up to one-third more likely to be obese or to have diabetes. Residential density and the availability of walkable destinations were each significantly associated with transportation and health outcomes. The combination of high levels of both measures was associated with the highest levels of walking or bicycling (p<0.0001) and public transit use (p<0.0026) and the lowest levels of automobile trips (p<0.0001), and diabetes prevalence (p<0.0001). We conclude that both residential density and the availability of walkable destinations are good measures of urban walkability and can be recommended for use by policy-makers, planners and public health officials. In our setting, the combination of both factors provided additional explanatory power.     

DNA Tetrominoes: The Construction of DNA Nanostructures Using Self-Organised Heterogeneous Deoxyribonucleic Acids Shapes

The unique programmability of nucleic acids offers alternative in constructing excitable and functional nanostructures. This work introduces an autonomous protocol to construct DNA Tetris shapes (L-Shape, B-Shape, T-Shape and I-Shape) using modular DNA blocks. The protocol exploits the rich number of sequence combinations available from the nucleic acid alphabets, thus allowing for diversity to be applied in designing various DNA nanostructures. Instead of a deterministic set of sequences corresponding to a particular design, the protocol promotes a large pool of DNA shapes that can assemble to conform to any desired structures. By utilising evolutionary programming in the design stage, DNA blocks are subjected to processes such as sequence insertion, deletion and base shifting in order to enrich the diversity of the resulting shapes based on a set of cascading filters. The optimisation algorithm allows mutation to be exerted indefinitely on the candidate sequences until these sequences complied with all the four fitness criteria. Generated candidates from the protocol are in agreement with the filter cascades and thermodynamic simulation. Further validation using gel electrophoresis indicated the formation of the designed shapes. Thus, supporting the plausibility of constructing DNA nanostructures in a more hierarchical, modular, and interchangeable manner.     

Top-Down Nanofabrication and Characterization of 20 nm Silicon Nanowires for Biosensing Applications

A top-down nanofabrication approach is used to develop silicon nanowires from silicon-on-insulator (SOI) wafers and involves direct-write electron beam lithography (EBL), inductively coupled plasma-reactive ion etching (ICP-RIE) and a size reduction process. To achieve nanometer scale size, the crucial factors contributing to the EBL and size reduction processes are highlighted. The resulting silicon nanowires, which are 20 nm in width and 30 nm in height (with a triangular shape) and have a straight structure over the length of 400 μm, are fabricated precisely at the designed location on the device. The device is applied in biomolecule detection based on the changes in drain current (I_ds), electrical resistance and conductance of the silicon nanowires upon hybridization to complementary target deoxyribonucleic acid (DNA). In this context, the scaled-down device exhibited superior performances in terms of good specificity and high sensitivity, with a limit of detection (LOD) of 10 fM, enables for efficient label-free, direct and higher-accuracy DNA molecules detection. Thus, this silicon nanowire can be used as an improved transducer and serves as novel biosensor for future biomedical diagnostic applications.