The byssus of the zebra mussel (Dreissena polymorpha): spatial variations in protein composition

The notorious biofouling organism Dreissena polymorpha (the zebra mussel) attaches to a variety of surfaces using a byssus, a series of protein threads that connect the animal to adhesive plaques secreted onto hard substrata. Here, the use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) to characterize the composition of different regions of the byssus is reported. All parts of the byssus show mass peaks corresponding to small proteins in the range of 3.7–7 kDa, with distinctive differences between different regions. Indeed, spectra from thread and plaques are almost completely non-overlapping. In addition, several peaks were identified that are unique to the interfacial region of the plaque, and therefore likely represent specialized adhesive proteins. These results indicate a high level of control over the distribution of proteins, presumably with different functions, in the byssus of this freshwater species.     

Molecular Mechanics Studies of Molybdenum Disulphide Catalysts Parameterisation of Molybdenum and Sulphur

Abstract

A method for the parameterisation of molybdenum disulphide is presented which reproduces the crystal structure accurately. The method involves calculating parameters such that there is no net force contribution from any individual term of the potential on any atom. Ideal bond lengths and bond angles are taken from the X-ray crystal structure; stretching and bending force constants are calculated from a combination of spectroscopic data and quantum mechanics calculations, whereby the energy function with bond length or bond angle is calculated and fitted with an harmonic potential. For the non-bonded Lennard-Jones parameters, the dispersion coefficient C was calculated by an interpolation of existing published parameters using a multiple regression and then the crystal energy was minimised with respect to the van der Waals radius r₀ using a fixed crystal fragment.

These parameters were tested for various models of the hexagonal and rhombohedral forms of MoS₂. RMS fits between structures minimised with molecular mechanics and experimental models ranged from 0.006 Å to 0.012 Å.     

Burkholderia pseudomultivorans sp. nov., a novel Burkholderia cepacia complex species from human respiratory samples and the rhizosphere

Eleven Burkholderia cepacia-like isolates of human clinical and environmental origin were examined by a polyphasic approach including recA and 16S rRNA sequence analysis, multilocus sequence analysis (MLSA), DNA base content determination, fatty acid methyl ester analysis, and biochemical characterization. The results of this study demonstrate that these isolates represent a novel species within the B. cepacia complex (Bcc) for which we propose the name Burkholderia pseudomultivorans. The type strain is strain LMG 26883^T (=CCUG 62895^T). B. pseudomultivorans can be differentiated from other Bcc species by recA gene sequence analysis, MLSA, and several biochemical tests including growth at 42 °C, acidification of sucrose and adonitol, lysine decarboxylase and β-galactosidase activity, and esculin hydrolysis.     

Strain variability in fatty acid composition of Chattonella marina (Raphidophyceae) and its relation to differing ichthyotoxicity toward rainbow trout gill cells

Lipid profiles of three strains (Mexico, Australia, Japan) of Chattonella marina (Subrahmanyan) Hara et Chihara were studied under defined growth (phosphate, light, and growth phase) and harvest (intact and ruptured cells) conditions. Triacylglycerol levels were always <2%, sterols <7%, free fatty acids varied between 2 and 33%, and polar lipids were the most abundant lipid class (>51% of total lipids). The major fatty acids in C. marina were palmitic (16:0), eicosapentaenoic (EPA, 20:5ω3), octadecatetraenoic (18:4ω3), myristic (14:0), and palmitoleic (16:1ω7c) acids. Higher levels of EPA were found in ruptured cells (21.4–29.4%) compared to intact cells (8.5–25.3%). In general, Japanese N‐118 C. marina was the highest producer of EPA (14.3–29.4%), and Mexican CMCV‐1 the lowest producer (7.9–27.1%). Algal cultures, free fatty acids from C. marina, and the two aldehydes 2E,4E‐decadienal and 2E,4E‐heptadienal (suspected fatty acid‐derived products) were tested against the rainbow trout fish gill cell line RTgill‐W1. The configuration of fatty acids plays an important role in ichthyotoxicity. Free fatty acid fractions, obtained by base saponification of total lipids from C. marina showed a potent toxicity toward gill cells (median lethal concentration, LC₅₀ (at 1 h) of 0.44 μg · mL^?1 in light conditions, with a complete loss of viability at >3.2 μg · mL^?1). Live cultures of Mexican C. marina were less toxic than Japanese and Australian strains. This difference could be related to differing EPA content, superoxide anion production, and cell fragility. The aldehydes 2E,4E‐decadienal and 2E,4E‐heptadienal also showed high impact on gill cell viability, with LC₅₀ (at 1 h) of 0.34 and 0.36 μg · mL^?1, respectively. Superoxide anion production was highest in Australian strain CMPL01, followed by Japanese N‐118 and Mexican CMCV‐1 strains. Ruptured cells showed higher production of superoxide anion compared to intact cells (e.g., 19 vs. 9.5 pmol · cell^?1 · hr^?1 for CMPL01, respectively). Our results indicate that C. marina is more ichthyotoxic after cell disruption and when switching from dark to light conditions, possibly associated with a higher production of superoxide anion and EPA, which may be quickly oxidized to produce more toxic derivates, such as aldehydes.     

Genetic and epigenetic integrity assessment of acclimatised papaya plants regenerated directly from shoot-tips following short- and long-term cryopreservation

A vitrification-based cryopreservation protocol was applied to in vitro sourced shoot-tips of four genotypes of Carica papaya; two female (70 and Z6) and two male (B2 and B4). Regeneration of ~58?% (70) and ~59?% (Z6) was recorded for the female genotypes confirming previously published results. Regeneration was at ~77 and ~53?% for the two male genotypes B2 and B4 respectively. Cryo-tube storage and regeneration was tested after 2?C18?months storage in one male (B2) and one female (70) genotype. Regeneration post cryo-storage was similar to 1?h exposure to liquid nitrogen. Individual shoot-tips from the two female and two male genotypes were grown into complete in vitro plants, potted and acclimatised without micropropagation to provide material for randomly amplified DNA fingerprinting (RAF) and amplified DNA methylation polymorphism (AMP) analysis of multiple individuals from in vitro control, plant vitrification solution 2 (PVS2) cryoprotectant control and short (1?h) and long-term cryopreservation treatment plants. No variations were detected for genotype Z6 control and treatment individuals and no RAF variations were detected in any individuals of genotype B2. Small numbers of RAF and AMP variations were detected in some individuals from genotypes B2 (AMP variation only), B4 and 70, but these were also found in controls. Genotype 70 showed the greatest level of variation; genomic DNA variation (RAF) was detected in control and cryopreservation treatment individuals, and the PVS2 control group was the only treatment group without variations for the respective AMP analysis. The variations observed could not be correlated with any phenotypic characteristics 2?months after acclimatisation.     

An optimised 3 M KCl salt-bridge technique used to measure and validate theoretical liquid junction potential values in patch-clamping and electrophysiology

Accurate potential measurements in electrophysiological experiments require correction for liquid junction potentials (LJPs), and, in patch-clamping especially, these can often be ~5–10 mV or more. They can be either calculated, if ion mobilities are known, or measured directly. We describe an optimised system to directly measure LJPs with a patch-clamp amplifier, using as a reference electrode, a freshly-cut 3 M KCl-agar salt-bridge (in polyethylene tubing) with its tip cut off by at least 5 mm during solution changes to eliminate its solution-history-dependent effects. We quantify such history-dependent effects and complement this with a de-novo theoretical analysis of salt diffusion to and from the salt-bridge. Our analysis and experimental results validate the optimised methodology for measuring LJPs, and the use of the Henderson equation for accurately calculating them. The use of this equation is also assessed and generally validated in the light of rigorous Nernst–Planck–Poisson and other numerical simulations and analytical studies of LJPs over recent decades. Digitizing, recording and amplifying the measured potentials increases their accuracy. The measured potentials still need correction for small, well-defined calculable, shifts in LJPs at the 3 M KCl-agar reference. Using this technique, we have measured changes in LJPs for diluted solutions of NaCl, LiCl, KCl, CsCl and NaF, obtaining excellent agreement within ±0.1 mV of predicted values, calculated using ion activities. Our de novo LJP measurements of biionic combinations of the above undiluted salts, and NaI and NaF (with halide anions I^? and F^?), generally also gave excellent agreement with predicted values.     

1-(4-Phenylpiperazin-1-yl)-2-(1H-pyrazol-1-yl)ethanones as novel CCR1 antagonists

A novel series of CCR1 antagonists based on the 1-(4-phenylpiperazin-1-yl)-2-(1H-pyrazol-1-yl)ethanone scaffold was identified by screening a compound library utilizing CCR1-expressing human THP-1 cells. SAR studies led to the discovery of the highly potent and selective CCR1 antagonist 14 (CCR1 binding IC₅₀ = 4 nM using [¹²⁵I]-CCL3 as the chemokine ligand). Compound 14 displayed promising pharmacokinetic and toxicological profiles in preclinical species.     

Azetidines and spiro azetidines as novel P2 units in hepatitis C virus NS3 protease inhibitors

Herein, we report the synthesis and structure–activity relationship studies of new analogs of boceprevir 1 and telaprevir 2. Introduction of azetidine and spiroazetidines as a P2 substituent that replaced the pyrrolidine moiety of 1 and 2 led to the discovery of a potent hepatitis C protease inhibitor 37c (EC₅₀ = 0.8 μM).     

Learning and signal copying facilitate communication among bird species

Signals relevant to different sets of receivers in different contexts create a conflict for signal design. A classic example is vocal alarm signals, often used both during intraspecific and interspecific interactions. How can signals alert individuals from a variety of other species in some contexts, while also maintaining efficient communication among conspecifics? We studied heterospecific responses to avian alarm signals that drive the formation of anti-predator groups but are also used during intraspecific interactions. In three species-rich communities in the western Himalayas, alarm signals vary drastically across species. We show that, independently of differences in their calls, birds respond strongly to the alarm signals of other species with which they co-occur and much more weakly to those of species with which they do not co-occur. These results suggest that previous exposure and learning maintain heterospecific responses in the face of widespread signal divergence. At an area where only two species regularly interact, one species'' calls incorporate the call of the other. We demonstrate experimentally that signal copying allows strong responses even without previous exposure and suggest that such hybrid calls may be especially favoured when pairwise interactions between species are strong.     

HIV-1 envelope determinants for cell tropism and chemokine receptor use

Isolates of human immunodeficiency virus type-1 (HIV-1) display marked differences in their ability to replicate in macrophages and transformed T-cell lines in vitro, a property that has important implications for disease pathogenesis. The restriction in replication between these two CD4-positive cell types is largely at the level of viral entry and is regulated by the viral envelope (env) gene. The envelope protein (Env) is responsible for fusion of the viral and host membranes, and a particular region of Env called the V3-loop has been implicated in regulating viral tropism. However, other regions of Env, such as the V1- and V2-loops, have been shown to modulate the effects of the V3-loop. The discovery that Env initially binds the CD4 molecule on the target cell surface and then makes subsequent interactions with one of several members of the chemokine receptor family has greatly enhanced the molecular understanding of HIV-1 entry. The differential use of chemokine receptors by different viral isolates and their expression in different cell types largely explains viral tropism. The same regions in Env responsible for virus tropism have also been shown to play an important role in mediating chemokine receptor use. The recent crystallization of HIV-1 Env in complex with CD4 illuminates the architecture of the components involved in mediating fusion between the viral and host membranes. The spatial relationship between variable structures of Env previously implicated in tropism and chemokine receptor use and conserved Env structures potentially involved in chemokine receptor binding are discussed.