S3EPY: a Sparky extension for determination of small scalar couplings from spin-state-selective excitation NMR experiments

S3EPY is a Python extension to the program Sparky written to facilitate the assessment of coupling constants from in-phase/antiphase and spin-state-selective excitation (S³E) experiments. It enables the routine use of small scalar couplings by automating the coupling evaluation procedure. S3EPY provides an integrated graphical user interface to programs which outputs graphs and the table of determined couplings.     

The Streamlined Genome of Phytomonas spp. Relative to Human Pathogenic Kinetoplastids Reveals a Parasite Tailored for Plants

Members of the family Trypanosomatidae infect many organisms, including animals, plants and humans. Plant-infecting trypanosomes are grouped under the single genus Phytomonas, failing to reflect the wide biological and pathological diversity of these protists. While some Phytomonas spp. multiply in the latex of plants, or in fruit or seeds without apparent pathogenicity, others colonize the phloem sap and afflict plants of substantial economic value, including the coffee tree, coconut and oil palms. Plant trypanosomes have not been studied extensively at the genome level, a major gap in understanding and controlling pathogenesis. We describe the genome sequences of two plant trypanosomatids, one pathogenic isolate from a Guianan coconut and one non-symptomatic isolate from Euphorbia collected in France. Although these parasites have extremely distinct pathogenic impacts, very few genes are unique to either, with the vast majority of genes shared by both isolates. Significantly, both Phytomonas spp. genomes consist essentially of single copy genes for the bulk of their metabolic enzymes, whereas other trypanosomatids e.g. Leishmania and Trypanosoma possess multiple paralogous genes or families. Indeed, comparison with other trypanosomatid genomes revealed a highly streamlined genome, encoding for a minimized metabolic system while conserving the major pathways, and with retention of a full complement of endomembrane organelles, but with no evidence for functional complexity. Identification of the metabolic genes of Phytomonas provides opportunities for establishing in vitro culturing of these fastidious parasites and new tools for the control of agricultural plant disease.     

Viral SELEX reveals individual and cooperative roles of the C-box and G-box in HIV-2 replication

The 5' UTR of HIV-2 genomic RNA contains signaling motifs that regulate specific steps of the replication cycle. Two motifs of interest are the C-box and the G-box. The C-box is found in the 5' untranslated region upstream of the primer binding site, while the G-box is found downstream from the major splice donor site, encompassing the gag start codon and flanking nucleotides. Together the C-box and the G-box form a long-range base-pairing interaction called the CGI. We and others have previously shown that formation of the CGI affects RNA dimerization in vitro and the positions of the C-box and the G-box are suggestive of potential roles of the CGI in other steps of HIV-2 replication. Therefore, we attempted to elucidate the role of the CGI using a viral SELEX approach. We constructed proviral DNA libraries containing randomized regions of the C-box or G-box paired with wild-type or mutant base-pairing partners. These proviral DNA libraries were transfected into COS-7 cells to produce viral libraries that were then used to infect permissive C8166 cells. The "winner" viruses were sequenced and further characterized. Our results demonstrate that there is strong selective pressure favoring viruses that can form a branched CGI. In addition, we show that the mutation of the C-box alone can enhance RNA encapsidation, and mutation of the G-box can alter the levels of Gag protein isoforms. These results suggest coordinated regulation of RNA translation, dimerization, and encapsidation during HIV-2 replication.     

Endoglin (CD105) expression is regulated by the liver X receptor alpha (NR1H3) in human trophoblast cell line JAR

Human implantation involves invasion of the uterine wall and remodeling of uterine arteries by extravillous cytotrophoblasts. Defects in these early steps of placental development lead to poor placentation and are often associated with preeclampsia, a frequent complication of human pregnancy. One of the complex mechanisms controlling trophoblast invasion involves the activation of the liver X receptor beta (or NR1H2, more commonly known as LXRbeta) by oxysterols known as potent LXR activators. This activation of LXRbeta leads to a decrease of trophoblast invasion. The identification of new target genes of LXR in the placenta could aid in the understanding of their physiological roles in trophoblast invasion. In the present study, we show that the endoglin (ENG) gene is a direct target of the liver X receptor alpha (NR1H3, also known as LXRalpha). ENG, whose gene is highly expressed in syncytiotrophoblasts, is part of the transforming growth factor (TGF) receptor complex that binds several members of the TGFbeta superfamily. In the human placenta, ENG has been shown to be involved in the inhibition of trophoblast invasion. Treatment of human choriocarcinoma JAR cells with T0901317, a synthetic LXR-selective agonist, leads to a significant increase in ENG mRNA and protein levels. Using transfection and electrophoretic mobility shift assays, we demonstrate that LXR (as a heterodimer with the retinoid X receptor) is able to bind the ENG promoter on an LXR response element and mediates the activation of ENG gene expression by LXRalpha in JAR cells. This study suggests a novel mechanism by which LXR may regulate trophoblast invasion in pathological pregnancy such as preeclampsia.     

Influence of zeaxanthin and echinenone binding on the activity of the Orange Carotenoid Protein

In most cyanobacteria high irradiance induces a photoprotective mechanism that downregulates photosynthesis by increasing thermal dissipation of the energy absorbed by the phycobilisome, the water-soluble antenna. The light activation of a soluble carotenoid protein, the Orange-Carotenoid-Protein (OCP), binding hydroxyechinenone, a keto carotenoid, is the key inducer of this mechanism. Light causes structural changes within the carotenoid and the protein, leading to the conversion of a dark orange form into a red active form. Here, we tested whether echinenone or zeaxanthin can replace hydroxyechinenone in a study in which the nature of the carotenoid bound to the OCP was genetically changed. In a mutant lacking hydroxyechinenone and echinenone, the OCP was found to bind zeaxanthin but the stability of the binding appeared to be lower and light was unable to photoconvert the dark form into a red active form. Moreover, in the strains containing zeaxanthin-OCP, blue-green light did not induce the photoprotective mechanism. In contrast, in mutants in which echinenone is bound to the OCP, the protein is photoactivated and photoprotection is induced. Our results strongly suggest that the presence of the carotenoid carbonyl group that distinguishes echinenone and hydroxyechinenone from zeaxanthin is essential for the OCP activity.     

An all-atom model of the chromatin fiber containing linker histones reveals a versatile structure tuned by the nucleosomal repeat length

In the nucleus of eukaryotic cells, histone proteins organize the linear genome into a functional and hierarchical architecture. In this paper, we use the crystal structures of the nucleosome core particle, B-DNA and the globular domain of H5 linker histone to build the first all-atom model of compact chromatin fibers. In this 3D jigsaw puzzle, DNA bending is achieved by solving an inverse kinematics problem. Our model is based on recent electron microscopy measurements of reconstituted fiber dimensions. Strikingly, we find that the chromatin fiber containing linker histones is a polymorphic structure. We show that different fiber conformations are obtained by tuning the linker histone orientation at the nucleosomes entry/exit according to the nucleosomal repeat length. We propose that the observed in vivo quantization of nucleosomal repeat length could reflect nature's ability to use the DNA molecule's helical geometry in order to give chromatin versatile topological and mechanical properties.     

Effect of acyl donor chain length and sugar/acyl donor molar ratio on enzymatic synthesis of fatty acid fructose esters

Lipase-catalyzed synthesis of fatty acid sugar esters through direct esterification was performed in 2-methyl 2-butanol as solvent. Fructose and saturated fatty acids were used as substrates and the reaction was catalyzed by immobilized Candida antarctica lipase. The effect of the initial fructose/acyl donor molar ratio and the carbon-chain length of the acyl donor as well as their reciprocal interactions on the reaction performance were investigated. For this purpose, an experimental design taking into account variations of the molar ratio (from 1:1 to 1:5) and the carbon-chain length of the fatty acid (from C8 to C18) was employed. Statistical analysis of the data indicated that the two factors as well as their interactions had significant effects on the sugar esters synthesis. The obtained results showed that whatever the molar ratio used, the highest concentration (73 g l⁻¹), fructose and fatty acid conversion yields (100% and 80%, respectively) and initial reaction rate (40 g l⁻¹ h⁻¹) were reached when using the C18 fatty acid as acyl donor. Low molar ratios gave the best fatty acid conversion yields and initial reaction rates, whereas the best total sugar ester concentrations and fructose conversion yields were obtained for high molar ratios.     

Enzymatic browning and biochemical alterations in black spots of pineapple [Ananas comosus (L.) Merr.

Penicillium funiculosum Thom. was consistently isolated from pineapple-infected fruitlet (black spots). Polyphenol oxidase, peroxidase, and laccase activities were determined in extracts from contiguous and infected fruitlets. Healthy fruitlets showed a rather high level of polyphenol oxidase (optimum pH 7.0), and this activity was tremendously increased (X 10) in contiguous infected fruitlets. Furthermore, infected fruitlets also exhibited laccase activity (optimum pH 4.0), while peroxidase was rather constant in both fruitlets. Browning reactions were attributed to qualitative and quantitative modifications of the enzymatic equipment (polyphenol oxidase and laccase) (p < 0.0001). In infected fruiltets, sucrose and L-malic acid were present at significantly lower amounts than in healthy ones, likely owing to fungal metabolism (p < 0.0001), whereas cell wall material was three times higher, which could be viewed as a defense mechanism to limit expansion of the mycelium.     

Characterization of scorpion alpha-like toxin group using two new toxins from the scorpion Leiurus quinquestriatus hebraeus.

Two novel toxins, Lqh6 and Lqh7, isolated from the venom of the scorpion Leiurus quinquestriatus hebraeus, have in their sequence a molecular signature (8Q/KPE10) associated with a recently defined group of alpha-toxins that target Na channels, namely the alpha-like toxins [reviewed in Gordon, D., Savarin, P., Gurevitz, M. & Zinn-Justin, S. (1998) J. Toxicol. Toxin Rev. 17, 131-159]. Lqh6 and Lqh7 are highly toxic to insects and mice, and inhibit the binding of alpha-toxins to cockroach neuronal membranes. Although they kill rodents by intracerebroventricular injection, they do not inhibit the binding of antimammal alpha-toxins (e.g. Lqh2) to rat brain synaptosomes, not even at high concentrations. Furthermore, in voltage-clamp experiments, rat brain Na channels IIA (rNav1.2A) expressed in Xenopus oocytes are not affected by Lqh6 nor by Lqh7 below 3 micro m. In contrast, muscular Na channels (rNav1.4 and hNav1.5) expressed in the same cells respond to nanomolar concentrations of Lqh6 and Lqh7 by slowing of Na current inactivation and a leftward shift of the peak conductance-voltage curve. The structural and pharmacological properties of the new toxins are compared to those of other scorpion alpha-toxins in order to re-examine the hallmarks previously set for the alpha-like toxin group.     

Efficiency,thermodynamic and kinetic stability of marketed gadolinium chelates and their possible clinical consequences: a critical review

Gadolinium-based contrast agents are widely used to enhance image contrast in magnetic resonance imaging (MRI) procedures. Over recent years, there has been a renewed interest in the physicochemical properties of gadolinium chelates used as contrast agents for MRI procedures, as it has been suggested that dechelation of these molecules could be involved in the mechanism of a recently described disease, namely nephrogenic systemic fibrosis (NSF). The aim of this paper is to discuss the structure-physicochemical properties relationships of marketed gadolinium chelates in regards to their biological consequences. Marketed gadolinium chelates can be classified according to key molecular design parameters: (a) nature of the chelating moiety: macrocyclic molecules in which Gd3+ is caged in the pre-organized cavity of the ligand, or linear open-chain molecules, (b) ionicity: the ionicity of the complex varies from neutral to tri-anionic agents, and (c) the presence or absence of an aromatic lipophilic residue responsible for protein binding. All these molecular characteristics have a profound impact on the physicochemical characteristics of the pharmaceutical solution such as osmolality, viscosity but also on their efficiency in relaxing water protons (relaxivity) and their biodistribution. These key molecular parameters can also explain why gadolinium chelates differ in terms of their thermodynamic stability constants and kinetic stability, as demonstrated by numerous in vitro and in vivo studies, resulting in various formulations of pharmaceutical solutions of marketed contrast agents. The concept of kinetic and thermodynamic stability is critically discussed as it remains a somewhat controversial topic, especially in predicting the amount of free gadolinium which may result from dechelation of chelates in physiological or pathological situations. A high kinetic stability provided by the macrocyclic structure combined with a high thermodynamic stability (reinforced by ionicity for macrocyclic chelates) will minimize the amount of free gadolinium released in tissue parenchymas.