Structural and dynamic changes associated with beneficial engineered single-amino-acid deletion mutations in enhanced green fluorescent protein.

Single-amino-acid deletions are a common part of the natural evolutionary landscape but are rarely sampled during protein engineering owing to limited and prejudiced molecular understanding of mutations that shorten the protein backbone. Single-amino-acid deletion variants of enhanced green fluorescent protein (EGFP) have been identified by directed evolution with the beneficial effect of imparting increased cellular fluorescence. Biophysical characterization revealed that increased functional protein production and not changes to the fluorescence parameters was the mechanism that was likely to be responsible. The structure EGFP(D190Δ) containing a deletion within a loop revealed propagated changes only after the deleted residue. The structure of EGFP(A227Δ) revealed that a `flipping mechanism was used to adjust for residue deletion at the end of a β-strand, with amino acids C-terminal to the deletion site repositioning to take the place of the deleted amino acid. In both variants new networks of short-range and long-range interactions are generated while maintaining the integrity of the hydrophobic core. Both deletion variants also displayed significant local and long-range changes in dynamics, as evident by changes in B factors compared with EGFP. Rather than being detrimental, deletion mutations can introduce beneficial structural effects through altering core protein properties, folding and dynamics, as well as function

Better together: building protein oligomers naturally and by design

This is the final version. Available on open access from Portland Press via the DOI in this recordProtein oligomers are more common in nature than monomers, with dimers being the most prevalent final structural state observed in known structures. From a biological perspective, this makes sense as it conserves vital molecular resources that may be wasted simply by generating larger single polypeptide units, and allows new features such as cooperativity to emerge. Taking inspiration from nature, protein designers and engineers are now building artificial oligomeric complexes using a variety of approaches to generate new and useful supramolecular protein structures. Oligomerisation is thus offering a new approach to sample structure and function space not accessible through simply tinkering with monomeric proteins

DNA-Directed Assembly of Carbon Nanotube-Protein Hybrids.

Here, we report the controlled assembly of SWCNT-GFP hybrids employing DNA as a linker. Two distinct, enriched SWCNTs chiralities, (6,5), (7,6), and an unsorted SWCNT solution, were selectively functionalized with DNA and hybridized to a complementary GFPDNA conjugate. Atomic force microscopy images confirmed that GFP attachment occurred predominantly at the terminal ends of the nanotubes, as designed. The electronic coupling of the proteins to the nanotubes was confirmed via in-solution fluorescence spectroscopy, that revealed an increase in the emission intensity of GFP when linked to the CNTs

Fit Fans: perspectives of a practitioner and understanding participant health needs within a health promotion programme for older men delivered within an English Premier League Football Club

Fit Fans was a men’s health promotion intervention delivered within an English Premier League Football Club (2010–2011), which aimed to support the local community dwelling older men in lifestyle promotion (physical activity [PA], diet and well-being). The purpose of this study was to provide a reflexive account of a practitioner and the needs of participants. Seven men (mean age 58 years) attended weekly PA and lifestyle sessions over an eight-month period. Baseline physiological measurements included body mass index, resting blood pressure and abdominal girth. Principles of ethnography and observational research (i.e. field notes, reflective diary) were adopted by the practitioner. Unexpectedly, the cohort exhibited a range of serious diagnosed illnesses that challenged the practitioner’s skill base and experience in the delivery of the intervention. Reflections of the practitioner and the stories of the progression that participants made add insight to future football in the community programmes

T2{}^2K2{}^2: The Twitter Top-K Keywords Benchmark

Information retrieval from textual data focuses on the construction of vocabularies that contain weighted term tuples. Such vocabularies can then be exploited by various text analysis algorithms to extract new knowledge, e.g., top-k keywords, top-k documents, etc. Top-k keywords are casually used for various purposes, are often computed on-the-fly, and thus must be efficiently computed. To compare competing weighting schemes and database implementations, benchmarking is customary. To the best of our knowledge, no benchmark currently addresses these problems. Hence, in this paper, we present a top-k keywords benchmark, T${}^2$K${}^2$, which features a real tweet dataset and queries with various complexities and selectivities. T${}^2$K${}^2$ helps evaluate weighting schemes and database implementations in terms of computing performance. To illustrate T${}^2$K${}^2$'s relevance and genericity, we successfully performed tests on the TF-IDF and Okapi BM25 weighting schemes, on one hand, and on different relational (Oracle, PostgreSQL) and document-oriented (MongoDB) database implementations, on the other hand

Optimal client recommendation for market makers in illiquid financial products

The process of liquidity provision in financial markets can result in prolonged exposure to illiquid instruments for market makers. In this case, where a proprietary position is not desired, pro-actively targeting the right client who is likely to be interested can be an effective means to offset this position, rather than relying on commensurate interest arising through natural demand. In this paper, we consider the inference of a client profile for the purpose of corporate bond recommendation, based on typical recorded information available to the market maker. Given a historical record of corporate bond transactions and bond meta-data, we use a topic-modelling analogy to develop a probabilistic technique for compiling a curated list of client recommendations for a particular bond that needs to be traded, ranked by probability of interest. We show that a model based on Latent Dirichlet Allocation offers promising performance to deliver relevant recommendations for sales traders.Comment: 12 pages, 3 figures, 1 tabl

Quantitative Imaging of B1 Cyclin Expression Across the Cell Cycle Using Green Fluorescent Protein Tagging and Epifluorescence

In this article, we report the number of cyclin B1 proteins tagged with enhanced green fluorescent protein (eGFP) in fixed U‐2 OS cells across the cell cycle. We use a quantitative analysis of epifluorescence to determine the number of eGFP molecules in a nondestructive way, and integrated over the cell we find 104 to 105 molecules. Based on the measured number of eGFP tagged cyclin B1 proteins, knowledge of cyclin B1 dynamics through the cell cycle, and the cell morphology, we identify the stages of cells in the cell cycle

Palynology, Phytoliths and Diatoms in and adjacent to the Great Cave: stratigraphic and taphonomic studies of Late Quaternary vegetation history

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Relationship between FEV1 change and patient-reported outcomes in randomised trials of inhaled bronchodilators for stable COPD: a systematic review.

BACKGROUND: Interactions between spirometry and patient-reported outcomes in COPD are not well understood. This systematic review and study-level analysis investigated the relationship between changes in FEV1 and changes in health status with bronchodilator therapy. METHODS: Six databases (to October 2009) were searched to identify studies with long-acting bronchodilator therapy reporting FEV1 and health status, dyspnoea or exacerbations. Mean and standard deviations of treatment effects were extracted for each arm of each study. Relationships between changes in trough FEV1 and outcomes were assessed using correlations and random-effects regression modelling. The primary outcome was St George's Respiratory Questionnaire (SGRQ) total score. RESULTS: Thirty-six studies (≥ 3 months) were included. Twenty-two studies (23,654 patients) with 49 treatment arms each contributing one data point provided SGRQ data. Change in trough FEV1 and change in SGRQ total score were negatively correlated (r = -0.46, p < 0.001); greater increases in FEV1 were associated with greater reductions (improvements) in SGRQ. The correlation strengthened with increasing study duration from 3 to 12 months. Regression modelling indicated that 100 mL increase in FEV1 (change at which patients are more likely to report improvement) was associated with a statistically significant reduction in SGRQ of 2.5 (95% CI 1.9, 3.1), while a clinically relevant SGRQ change (4.0) was associated with 160.6 (95% CI 129.0, 211.6) mL increase in FEV1. The association between change in FEV1 and other patient-reported outcomes was generally weak. CONCLUSIONS: Our analyses indicate, at a study level, that improvement in mean trough FEV1 is associated with proportional improvements in health status