Transmission vs. Diffuse Transmission in Circular Dichroism: What to Choose for Probing Solid‐State Samples?

Recent advances in equipment enabled the collection of solid‐state electronic circular dichroism (ECD) spectra using the commercially available integrating sphere attachment for a regular ECD spectrometer. This accessory was designed to reduce negative factors occurring in solid‐state ECD measurements, and is, thereby, very useful for recording diffuse transmittance CD (DTCD) spectra using the pellet technique. In the present article, the operating principle of the integrating sphere and utility of the DTCD method in recording solid‐state ECD spectra is demonstrated. Based on illustrative examples, i.e., 10‐camphorsulfonic acid ammonium, cholest‐4‐en‐3‐one, (3R,4R,5S)‐oseltamivir, and (S)‐linezolid, ECD solid‐state measurements were performed by means of both transmission and diffusion methods and later compared. Selection of these compounds as models for comparative studies was made in view of their different chromophoric systems and the profound importance in the pharmaceutical industry. During the course of this work the benefits and limitations of the use of integrating sphere are presented. The final conclusion is that more relevant solid‐state spectra can be obtained by means of the DTCD method. Chirality 27:441–448, 2015. © 2015 Wiley Periodicals, Inc.     

Design and synthesis of new substrates of HtrA2 protease

HtrA2 belongs to the HtrA (high temperature requirement A) family of ATP-independent serine proteases. The primary function of HtrA2 includes maintaining the mitochondria homeostasis, cell death (by apoptosis, necrosis, or anoikis), and contribution to the cell signaling. Several recent reports have shown involvement of HtrA2 in development of cancer and neurodegenerative disorders. Here, we describe the profiling of HtrA2 protease substrate specificity via the combinatorial chemistry approach that led to the selection of novel intramolecularly quenched substrates. For all synthesized compounds, the highest HtrA2-mediated hydrolysis efficiency and selectivity among tested HtrA family members was observed for ABZ-Ile-Met-Thr-Abu-Tyr-Met-Phe-Tyr(3-NO₂)-NH₂, which displayed a specificity constant k_cat/K_M value of 14,535 M⁻¹ s⁻¹.     

The Cell Division Protein FtsZ from Streptococcus pneumoniae Exhibits a GTPase Activity Delay

The cell division protein FtsZ assembles in vitro by a mechanism of cooperative association dependent on GTP, monovalent cations, and Mg²⁺. We have analyzed the GTPase activity and assembly dynamics of Streptococcus pneumoniae FtsZ (SpnFtsZ). SpnFtsZ assembled in an apparently cooperative process, with a higher critical concentration than values reported for other FtsZ proteins. It sedimented in the presence of GTP as a high molecular mass polymer with a well defined size and tended to form double-stranded filaments in electron microscope preparations. GTPase activity depended on K⁺ and Mg²⁺ and was inhibited by Na⁺. GTP hydrolysis exhibited a delay that included a lag phase followed by a GTP hydrolysis activation step, until reaction reached the GTPase rate. The lag phase was not found in polymer assembly, suggesting a transition from an initial non-GTP-hydrolyzing polymer that switches to a GTP-hydrolyzing polymer, supporting models that explain FtsZ polymer cooperativity.     

Sources,metabolism, and regulation of circulating sphingosine-1-phosphate

Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid that acts either as an intracellular messenger or as a ligand for its membrane receptors. S1P is a normal constituent of blood, where it is found both in plasma and blood cells. Compared with other cell types, sphingolipid metabolism in erythrocytes and platelets has unique features that allow the erythrocytes and platelets to accumulate S1P. In plasma, S1P is bound mainly to HDLs and albumin. Of note, metabolism and biological activity of S1P is to a large extent affected by the type of its carrier. Plasma S1P is characterized by a short half-life, indicating rapid clearance by degradative enzymes and the presence of high-capacity sources involved in maintaining its high concentration. These sources include blood cells, vascular endothelium, and hepatocytes. However, the extent to which each of these contributes to the plasma pool of S1P is a matter of debate. Circulating S1P plays a significant physiological role. It was found to be the key regulator of lymphocyte trafficking, endothelial barrier function, and vascular tone. The purpose of this review is to summarize the present state of knowledge on the metabolism, transport, and origin of plasma S1P, and to discuss the mechanisms regulating its homeostasis in blood.     

Phylogenetic reassessment and biogeography of the Ectateus generic group (Coleoptera: Tenebrionidae: Platynotina)

Owing to the reinterpretation of its morphological synapomorphies, the taxonomic composition of the Ectateus generic group had been ambiguous. The present study scrutinized all existing taxonomic concepts of the group based on a cladistic analysis of the adult morphology of all of the Afrotropical platynotoid Platynotina genera. The phylogenetic relationships were reconstructed using parsimony and Bayesian inference. The results show that all previous taxonomic concepts of the Ectateus generic group concerned paraphyletic entities. The cladistic analysis revealed the following synapomorphies for the taxon: (1) presence of basal indentations of the pronotal disc, (2) ratio of prothorax width to its maximal height > 6.0, and (3) ratio of maximal height of the prothorax to total height < 0.3. Moreover, phylogenetic studies revealed the existence of the Upembarus generic group, a sister‐taxon group to the Ectateus generic group, within the Afrotropical platynotoid Platynotina. Autapomorphic and synapomorphic character mapping show that several taxonomic and nomenclatural changes are needed to consider the particular generic‐level entities traditionally assigned to Afrotropical platynotoid Platynotina as monophyletic lineages. The following taxonomic and nomenclatural adjustments are made in this paper: P teroselinus gen. nov. is erected to accommodate a single species that was previously assigned to Zidalus: Pteroselinus insularis comb. nov. Additionally, the following synonymies are proposed: Anchophthalmops (= Platykochius syn. nov. ), Angolositus (= Aberlencus syn. nov. , = Platymedvedevia syn. nov. ), Glyptopteryx (= Microselinus syn. nov. , = Quadrideres syn. nov. , = Synquadrideres syn. nov. ). In addition, Kochogaster is lowered in rank and is treated as one of the subgenera of Anchophthalmus. Moreover, Pseudoselinus is treated as a subgenus of Upembarus. An identification key to all Afrotropical platynotoid Platynotina genera and subgenera is presented. Zoogeographical analyses revealed the following dispersal barriers for the Ectateus generic group: (1) the Sahara (northern barrier); (2) the dry ecosystems of Botswana, Namibia, and South Africa (southern barrier); and (3) the Congolian rainforests (internal distributional gap). The ancestor of the taxon probably originated in East African ecoregions that predominantly contained wattletrees (acacias) and Commiphora Jacq. Moreover, past climate changes seem to have had a great impact on the observed generic distribution. © 2015 The Linnean Society of London     

Does temperature and oxygen affect duration of intramarsupial development and juvenile growth in the terrestrial isopod Porcellio scaber (Crustacea,Malacostraca)?

According to the temperature-size rule (TSR), ectotherms developing under cold conditions experience slower growth as juveniles but reach a larger size at maturity. Whether temperature alone causes this phenomenon is unknown, but oxygen limitation can play a role in the temperature-size relationship. Oxygen may become limited under warm conditions when the resulting higher metabolism creates a greater demand for oxygen, especially in larger individuals. We examined the independent effects of oxygen concentration (10% and 22% O₂) and temperature (15 °C and 22 °C) on duration of ontogenic development, which takes place within the maternal brood pouch (marsupium), and juvenile growth in the terrestrial isopod common rough woodlouse (Porcellio scaber). Individuals inside the marsupium undergo the change from the aqueous to the gaseous environment. Under hypoxia, woodlice hatched from the marsupium sooner, but their subsequent growth was not affected by the level of oxygen. Marsupial development and juvenile growth were almost three times slower at low temperature, and marsupial development was longer in larger females but only in the cold treatment. These results show that temperature and oxygen are important ecological factors affecting developmental time and that the strength of the effect likely depends on the availability of oxygen in the environment.     

Refined model of optical activity of cis-dihydrodiol metabolites: role of 1,3-diene conformation in the electronic circular dichroism spectra

The effect of the secondary structural feature, that is, nonplanarity of carbon-carbon double bonds, on the rotatory strength of the long-wavelength π-π* electronic transition has been investigated for the series of monocyclic cis-dihydrodiol arene metabolites and model compounds. The contribution from nonplanar C=C bonds to the overall rotatory strength of the π-π* electronic transition is more significant than the contribution from twisted 1,3-diene chromophore. The effect of the substitution pattern in the cyclohexadiene skeleton may be decisive for the sign of the long-wavelength π-π* Cotton effect.     

Growth inhibition and intracellular distribution of Pb ions by the white-rot fungus Abortiporus biennis

When Abortiporus biennis was grown on PbO-amended media, Pb (II) ions accumulated near fungal membrane structures, in the cell wall, and in the cytoplasm of the fungal cells, and their presence caused cell vacuolization. Increased concentration of PbO in the growth media caused a decline in fungal accumulation capacity. Neither PbO solubilization nor an increased level of organic acids underneath the mycelium was found.     

CENP-C is a structural platform for kinetochore assembly

Centromeres provide a region of chromatin upon which kinetochores are assembled in mitosis. Centromeric protein C (CENP-C) is a core component of this centromeric chromatin that, when depleted, prevents the proper formation of both centromeres and kinetochores. CENP-C localizes to centromeres throughout the cell cycle via its C-terminal part, whereas its N-terminal part appears necessary for recruitment of some but not all components of the Mis12 complex of the kinetochore. We now find that all kinetochore proteins belonging to the KMN (KNL1/Spc105, the Mis12 complex, and the Ndc80 complex) network bind to the N-terminal part of Drosophila CENP-C. Moreover, we show that the Mis12 complex component Nnf1 interacts directly with CENP-C in vitro. To test whether CENP-C's N-terminal part was sufficient to recruit KMN proteins, we targeted it to the centrosome by fusing it to a domain of Plk4 kinase. The Mis12 and Ndc80 complexes and Spc105 protein were then all recruited to centrosomes at the expense of centromeres, leading to mitotic abnormalities typical of cells with defective kinetochores. Thus, the N-terminal part of Drosophila CENP-C is sufficient to recruit core kinetochore components and acts as the principal linkage between centromere and kinetochore during mitosis.     

A critical comparative assessment of predictions of protein-binding sites for biologically relevant organic compounds

Protein function annotation and rational drug discovery rely on the knowledge of binding sites for small organic compounds, and yet the quality of existing binding site predictors was never systematically evaluated. We assess predictions of ten representative geometry-, energy-, threading-, and consensus-based methods on a new benchmark data set that considers apo and holo protein structures with multiple binding sites for biologically relevant ligands. Statistical tests show that threading-based Findsite outperforms other predictors when its templates have high similarity with the input protein. However, Findsite is equivalent or inferior to some geometry-, energy-, and consensus-based methods when the similarity is lower. We demonstrate that geometry-, energy-, and consensus-based predictors benefit from the usage of holo structures and that the top four methods, Findsite, Q-SiteFinder, ConCavity, and MetaPocket, perform better for larger binding sites. Predictions from these four methods are complementary, and our simple meta-predictor improves over the best single predictor.