Mechanistic insights into allosteric regulation of the A2A adenosine G protein-coupled receptor by physiological cations.

Cations play key roles in regulating G-protein-coupled receptors (GPCRs), although their mechanisms are poorly understood. Here, 19F NMR is used to delineate the effects of cations on functional states of the adenosine A2A GPCR. While Na+ reinforces an inactive ensemble and a partial-agonist stabilized state, Ca2+ and Mg2+ shift the equilibrium toward active states. Positive allosteric effects of divalent cations are more pronounced with agonist and a G-protein-derived peptide. In cell membranes, divalent cations enhance both the affinity and fraction of the high affinity agonist-bound state. Molecular dynamics simulations suggest high concentrations of divalent cations bridge specific extracellular acidic residues, bringing TM5 and TM6 together at the extracellular surface and allosterically driving open the G-protein-binding cleft as shown by rigidity-transmission allostery theory. An understanding of cation allostery should enable the design of allosteric agents and enhance our understanding of GPCR regulation in the cellular milieu

NMR Study of the Dissolution of Core-Crystalline Micelles

Short fragments of the core-crystalline micelles formed by a sample of poly­(ferrocenyl­dimethylsilane)-<i>block</i>-poly­(isoprene) (PFS-<i>b</i>-PI) block copolymer (BCP) underwent self-seeding in decane when heated above its dissolution temperature. Variable temperature (VT) ¹H NMR and diffusion-ordered pulsed-gradient spin–echo (DOSY) NMR were used to monitor the behavior of micelles that dissolved as a function of increasing temperature. We examined a sample of micelle fragments of PFS₆₅-<i>b</i>-PI₆₃₇ characterized by <i>L</i>_n = 39 nm and <i>L</i>_w/<i>L</i>_n = 1.13. The PI corona had high mobility and gave a ¹H NMR signal in both micellar and unimer forms. In contrast, the PFS component could only be detected for the dissolved unimer. We found from ¹H NMR that essentially all the BCP molecules were incorporated into the micelles at temperatures up to and including 50 °C, at the limit of NMR detection. Both PFS and PI resonances could be detected between 70 and 100 °C, and the integration ratio of the PFS-to-PI peaks increased with temperature. DOSY NMR measured the self-diffusion coefficients (<i>D</i>_s) of the micelle fragments and unimer at these temperatures. The hydrodynamic radii (<i>R</i>_h) for these species were calculated from <i>D</i>_s using the Stokes–Einstein equation. The PFS signals gave <i>R</i>_h values in the range of 5–6 nm at temperatures between 80 and 100 °C, consistent with unimer diffusion. PI signals were fitted by an exponential decay at 25 °C with <i>R</i>_h = 38 nm characteristic of the micelle fragments and at 90, 95, and 100 °C with <i>R</i>_h ≈ 6 nm, corresponding to unimer. At intermediate temperatures (70–85 °C), PI signals were fitted to a sum of two exponential terms, consistent with a fast diffusing species and a slow diffusing species. Interestingly, we noticed that the size of the micelle fragments at elevated temperatures (80 and 85 °C) was sensitive to sample history; samples heated directly to the elevated temperatures were found to be shorter than those heated stepwise

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

Oxidative Kinetic Resolution of Aromatic Alcohols Using Iron Nanoparticles

The final publication is available at Springer via http://dx.doi.org/10.1007/s11244-013-0086-2Using our transfer hydrogenation (TH) pre-catalyst trans-(R,R)-[Fe(NCMe)CO(PPh2C6H4CH=NCHPh-)2][BF4]2 (1) we investigated the reverse reaction, oxidative kinetic resolution, and were able to achieve turn-over frequencies up to 335 h−1 and s-values in favour of the (R)-alcohol up to 10.2. Using racemic 1-phenylethanol we optimized reaction conditions to maximize enantioselectivity and turn-over frequency (TOF) and studied the effect of different proton/hydride acceptors, temperatures, and bases. Using KOtBu as the base and benzophenone in THF as the solvent and acceptor at 45 °C, we tested a series of substrates with varying electronic and steric factors. By increasing the steric bulk at the alcohol, the enantioselectivity increased, however the TOF decreased dramatically. Varying the electronics of the substrates using electron withdrawing and donating substituents showed a less significant effect. We propose that the active species in catalysis is zero-valent iron nanoparticles (Fe NPs), a postulate that we support with microscopy imaging, sub-stoichiometric poisoning experiments, and analysis of the reaction profile. Further support is given in terms of a polymer-supported substrate experiment whereby the active species in catalysis is too large to permeate the pores of a functionalized polymer. To the best of our knowledge, we propose that this is the first reported example of using a nanoparticle surface for oxidative kinetic resolution

View to the U: An eye on UTM Research

This is an audio recording from the podcast series "View to the U: An eye on UTM research".On this episode of VIEW to the U, Drs. Katie Harris-Howard, Elizabeth Parke, Dmitry Pichugin, and Vera Velasco from the Office of the Vice-Principal, Research at UofT Mississauga, talk about their roles as Senior Research Associates or SRA. So, what is an SRA? At their core, they are educators and academics in their own right, with established programs of research and expertise in their respective disciplines – in this case, cellular neurobiology, visual culture, biophysics, and plant physiology. And as part of their roles at UTM, they oversee a Core Facility. Find out more on this episode of VIEW to the U

Fuel interchangeability for lean premixed combustion in cylindrical radiant burner operated in the internal combustion mode

The trend towards a transition from fossil to renewable energy makes urgent the development of fuel-flexible burners that provides interchangeability between natural gas and a wide range of renewable gases. This study examines the fuel interchangeability potential of a novel annular cylindrical burner made of a Ni-Al intermetallic with advanced high-temperature properties. The burner is designed to operate in the internal combustion mode when the reactions are aerodynamically stabilized in the volume of the cylindrical cavity. The stability limits, radiation efficiency, and CO/NOX emissions of the burner were studied within the firing rate range of 160–420 kW/m2 and the equivalence ratio range of 0.5–1.0. Experiments were carried out for the natural gas and its blends with CO2, H2 and H2-CO, whose flame speeds differ by a factor of three. The addition of H2 and CO to natural gas was found to reduce emissions and expands the radiant operation mode to a lean region without loss in radiation efficiency. The addition of CO2 to natural gas also reduces NOX, but CO emission increases, radiation efficiency decreases and the infrared range is narrowed. For all studied fuels, within the ranges of firing rates of 260–420 kW/m2 and equivalence ratios of 0.70–0.95, the burner operates in stable radiant mode with a radiation efficiency up to 30–45%, NOX emission of 10–50 ppm, and CO emission of 0–40 ppm. In general, the results confirm the fuel flexibility of the new radiant burner to be much the same as for other radiant burners; however, the radiation efficiency is substantially higher than that for cylindrical radiant burners of other designs

Site-Specific Labeling of Protein Lysine Residues and N‑Terminal Amino Groups with Indoles and Indole-Derivatives

Indoles and indole-derivatives can be used to site-specifically label proteins on lysine and N-terminal amino groups under mild, nondenaturing reaction conditions. Hen egg white lysozyme (HEWL) and α-lactalbumin were labeled with indole, fluoroindole, or fluoroindole-2-carboxylate via electrophilic aromatic substitutions to lysine side chain Nε- and N-terminal amino imines, formed in situ in the presence of formaldehyde. The reaction is highly site-selective, easily controlled by temperature, and does not eliminate the native charge of the protein, unlike many other common lysine-specific labeling strategies. ¹⁹F NMR was used to monitor reaction progression, and in the case of HEWL, unique resonances for each labeled side chain could be resolved. We demonstrate that the indole tags are highly selective for primary amino groups. ¹⁹F NMR demonstrates that each lysine exhibits a different rate of conjugation to indoles making it possible to employ these tags as a means of probing surface topology by NMR or mass spectrometry. Given the site-specificity of this tagging method, the mildness of the reaction conditions (aqueous, buffered, or unbuffered) and the low stoichiometry required for the reaction, indole-derivatives should serve as a valuable addition to the bioconjugation toolkit. We propose that labeling lysine side chains and N-terminal amino groups with indoles is a versatile and general strategy for bioconjugations with substituted indoles having broad implications for protein functionalization

Effect of Concentration on the Dissolution of One-Dimensional Polymer Crystals:A TEM and NMR Study

We report a study of the dissolution of core-crystalline polyferrocenyl­dimethylsilane-<i>block</i>-polyisoprene (PFS₅₃-<i>b</i>-PI₆₃₇, where the subscripts are the degrees of polymerization of the two blocks) micelle fragments in decane for different concentrations (ranging from 0.01 to 6 mg mL^–1) by a combination of transmission electron microscopy (TEM) and high-temperature ¹H NMR. We used self-seeding experiments at different temperatures as an efficient, although indirect, way to evaluate the dissolution of these micelles fragments. We annealed micelle fragment solutions at five different temperatures (50, 60, 65, 70, and 75 °C) for 30 min and cooled them to room temperature to regrow the micelles. The amount of micelle fragments that dissolved at the annealing temperature was then evaluated by comparing the length of the regrown micelles with that of the starting micelle fragments. We show that seed crystallites are less prone to dissolution as their concentration increases. In addition, by combining results of self-seeding experiments and ¹H NMR measurements at 75 °C, we evaluated the percentage of unimer released upon the partial dissolution of seed fragments at 75 °C and established that the mechanism of seed fragment dissolution is also concentration dependent: at low concentrations, they dissolve in a cooperative process, whereas at high concentrations, they dissolve partially from both ends