The EPR and LMR spectra of the DO2 radical: Determination of ground-state parameters

The detection of lines in both the gas phase EPR spectrum at 9 GHz and the far-infrared LMR spectrum of the DO₂ radical is reported. The measurements are fitted with an appropriate Hamiltonian and several parameters for the molecule in the $\text{X}\text{?}{}^2\text{A″}$ state are determined. The majority of the transitions in the EPR spectrum are K-type doubling transitions and involve the a-component of the electric dipole moment. However the observation of one b-type transition (5₀₅-4₁₄) permits the determination of the off-diagonal component of the spin-rotation tensor, ?_ab, and an estimate of the relative magnitudes of the a- and b-components of the dipole moment. A combination of the parameters for HO₂ with those for DO₂ leads to a better understanding of the properties of the molecule. In particular, the r₀ molecular geometry has been determined more reliably than previously to be $\text{r}{}_0\text{(OH) = 0.9774}\text{A}\text{?}$, $\text{r}{}_0\text{(OO) = 1.3339}\text{A}\text{?}$, ∠HOO = 104.15°.     

Far infrared laser magnetic resonance spectra of the ClSO and FSO radicals in the gas phase

The detection of the free radicals ClSO and FSO by the technique of far infrared laser magnetic resonance is reported. Observations of the former have been made with several laser lines; the spectra are complicated and have not been assigned but are attributed to ClSO on the basis of several chemical tests. The FSO radical was generated by the reaction of carbon disulphide with the products of a discharge through CF₄ + O₂. Spectra have been observed with laser lines at 394, 419 and 513 μm. Using the results of a subsequent microwave study of FSO, the rotational transitions involved in these spectra have been identified. The spectrum at 513 μm has been studied in detail and is shown to arise through level anticrossings between the levels 14_5,Kc (the lower levels involved in the transition) and 15_4,Kc+2. All aspects of this spectrum are reproduced quantitatively by the parameters determined from the microwave spectrum. It is thought that the other two FSO spectra arise in a similar manner. Such anticrossing signals are likely to be widespread in LMR experiments, causing intense narrow lines to occur at high magnetic fields even for molecules with weakly coupled electron spin.     

Markov Chain Sampling Methods for Dirichlet Process Mixture Models

Abstract

This article reviews Markov chain methods for sampling from the posterior distribution of a Dirichlet process mixture model and presents two new classes of methods. One new approach is to make Metropolis—Hastings updates of the indicators specifying which mixture component is associated with each observation, perhaps supplemented with a partial form of Gibbs sampling. The other new approach extends Gibbs sampling for these indicators by using a set of auxiliary parameters. These methods are simple to implement and are more efficient than previous ways of handling general Dirichlet process mixture models with non-conjugate priors.     

Darobactin B Stabilises a Lateral-Closed Conformation of the BAM Complex in E. coli Cells

The β-barrel assembly machinery (BAM complex) is essential for outer membrane protein (OMP) folding in Gram-negative bacteria, and represents a promising antimicrobial target. Several conformational states of BAM have been reported, but all have been obtained under conditions which lack the unique features and complexity of the outer membrane (OM). Here, we use Pulsed Electron-Electron Double Resonance (PELDOR, or DEER) spectroscopy distance measurements to interrogate the conformational ensemble of the BAM complex in E. coli cells. We show that BAM adopts a broad ensemble of conformations in the OM, while in the presence of the antibiotic darobactin B (DAR-B), BAM′s conformational equilibrium shifts to a restricted ensemble consistent with the lateral closed state. Our in-cell PELDOR findings are supported by new cryoEM structures of BAM in the presence and absence of DAR-B. This work demonstrates the utility of PELDOR to map conformational changes in BAM within its native cellular environment.     

Monitoring copopulated conformational states during protein folding events using electrospray ionization-ion mobility spectrometry-mass spectrometry

The precise mechanism of protein folding remains elusive and there is a deficiency of biophysical techniques that are capable of monitoring the individual behavior of copopulated protein conformers during the folding process. Herein, an ion mobility spectrometry (IMS) device integrated with electrospray ionization mass spectrometry (ESI-MS) has been used to successfully separate and analyze protein conformers differing in cross section and/or charge state. In an initial test, an ensemble of folded and partially folded conformers of the protein cytochrome c was separated. A detailed study undertaken on the amyloidogenic protein beta(2)-microglobulin (beta(2)m), which forms fibrils by protein unfolding followed by self-aggregation and is responsible for the disease dialysis-related amyloidosis, has generated important insights into its folding landscape. Initially, a systematic titration of beta(2)m over the pH range 2 to 7 using ESI-IMS-MS allowed individual conformers to be monitored and quantified throughout the acid denaturation process. Furthermore, a comparison of wild-type beta(2)m with single and double amino acid variants with a range of folding stabilities and propensities for amyloid fibril formation has provided illuminating evidence of the role of different conformers in protein stability and amyloidogenic aggregation. The ESI-IMS-MS data presented here not only demonstrate an important and informative further dimension to ESI-MS, but also illustrate the potential of the ESI-IMS-MS technique for unravelling protein folding enigmas in general and studying protein misfolding diseases in particular.     

Controlled/living radical polymerization of isoprene and butadiene in emulsion

A process for RAFT-controlled radical polymerization in emulsion [36] has been applied to the polymerizations of isoprene and of butadiene in emulsion systems, with the goal of producing latex particles containing block copolymers of acrylic acid (stabilizer and starting polymer), styrene (second polymer) and isoprene or butadiene (third polymer). The microstructure of the polymer chains was examined using dual-detection size-exclusion chromatography, and the nanostructure of the materials was investigated by differential scanning calorimetry and solid-state nuclear magnetic resonance. Reactions were always slow (although faster than the corresponding processes in solution), and exhibited limited reinitiation by isoprene when in emulsion. The materials containing isoprene exhibit a nanostructure with a phase separation into high-T_g polystyrene-rich domains and low-T_g polyisoprene-rich domains, revealed by DSC and NMR. This has the potential to lead to barrier materials with novel physical properties.     

Investigating the structural properties of amyloid-like fibrils formed in vitro from beta2-microglobulin using limited proteolysis and electrospray ionisation mass spectrometry

The protein beta(2)-microglobulin (beta(2)m) aggregates to form classical amyloid fibrils in patients undergoing long-term haemodialysis. Amyloid-like fibrils with a cross-beta fold can also be formed from wild-type beta(2)m under acidic conditions in vitro. The morphology of such fibrils depends critically on the conditions used: incubation of beta(2)m in low ionic strength buffers at pH 2.5 results in the formation of long (microm), straight fibrils while, at pH 3.6, short (<500 nm) fibrils form. At higher ionic strengths (0.2-0.4 M) at pH 1.5-3.6, the fibrils have a distinct curved and nodular morphology. To determine the conformational properties of beta(2)m within in vitro fibrils of different morphologies, limited proteolysis of each fibril type using pepsin was performed and the resulting peptide fragments identified by tandem mass spectrometry. For comparison, the proteolytic degradation patterns of monomeric beta(2)m and seven synthetic peptides spanning the entire sequence of the intact protein were similarly analysed. The results show that fibrils with different morphologies result in distinct digestion patterns. While the curved, worm-like fibrils are relatively weakly protected from proteolysis, the long, straight fibrils formed at pH 2.5 at low ionic strength show only a single cut-site at Val9, demonstrating that substantial refolding of the initially acid-denatured and unprotected state of beta(2)m occurs during assembly. The data demonstrate that the organisation of the polypeptide chain in fibrils with different morphological features differs considerably, despite the fact that the fibrils possess a common cross-beta architecture.     

Phenolic compounds,essential oil composition,and antioxidant activity of Angelica purpurascens (Avé-Lall.) Gill

In this study, methanol extracts (MEs) and essential oil (EO) of Angelica purpurascens (Avé-Lall.) Gill obtained from different parts (root, stem, leaf, and seed) were evaluated in terms of antioxidant activity, total phenolics, compositions of phenolic compound, and essential oil with the methods of 2,2-azino-bis(3ethylbenzo-thiazoline-6-sulfonic acid (ABTS•⁺), 2,2-diphenyl-1-picrylhydrazil (DPPH•) radical scavenging activities, and ferric reducing/antioxidant power (FRAP), the Folin–Ciocalteu, liquid chromatography−tandem mass spectrometry (LC−MS/MS), and gas chromatography-mass spectrometry (GC−MS), respectively. The root extract of A. purpurascens exhibited the highest ABTS•⁺, DPPH•, and FRAP activities (IC₅₀: 0.05 ± 0.0001 mg/mL, IC₅₀: 0.06 ± 0.002 mg/mL, 821.04 ± 15.96 µM TEAC (Trolox equivalent antioxidant capacity), respectively). Moreover, EO of A. purpurascens root displayed DPPH• scavenging activity (IC₅₀: 2.95 ± 0.084 mg/mL). The root extract had the highest total phenolic content (438.75 ± 16.39 GAE (gallic acid equivalent), µg/mL)). Twenty compounds were identified by LC−MS/MS. The most abundant phenolics were ferulic acid (244.39 ± 15.64 μg/g extract), benzoic acid (138.18 ± 8.84 μg/g extract), oleuropein (78.04 ± 4.99 μg/g extract), and rutin (31.21 ± 2.00 μg/g extract) in seed, stem, root, and leaf extracts, respectively. According to the GC−MS analysis, the major components were determined as α-bisabolol (22.93%), cubebol (14.39%), α-pinene (11.63%), and α-limonene (9.41%) among 29 compounds. Consequently, the MEs and EO of A. purpurascens can be used as a natural antioxidant source.     

Electrochemical reactivity at redox-molecule-based nanoelectrode ensembles

A model describing electrochemical reactivity at nanoelectrode ensembles consisting of redox-molecule-based active sites immobilized on otherwise passivated electrode surfaces is presented. A mathematical treatment in terms of hemispherical diffusion of redox-active solutes to a layer of independent molecule-based nanoelectrode sites is shown to be equivalent to one in terms of a bimolecular diffusion-limited reaction between a layer of immobilized redox molecules and a reservoir of redox-active solutes. This equivalence derives from the fact that in both cases the mass-transfer problem is essentially that of hemispherical diffusion. The model is further developed to consider rate limitation by both the bimolecular redox reaction between the active-site molecule and redox molecules in solution and the heterogeneous redox reaction between the electrode and the active-site molecule. Analytical expressions are derived for the current–voltage relation corresponding to catalyzed electron transfer at an ensemble of redox-molecule-based nanoelectrode sites, and the expressions are used to interpret preliminary data for ultrasensitive electrochemical detection in flow streams via an electrochemical amplification process that is thought to involve redox mediation by individual analyte molecules adsorbed onto monolayer-coated electrodes.