Osmosensorische Eigenschaften des Glycinbetain-Transporters BetP aus Corynebacterium glutamicum

Unter hyperosmotischem Stress akkumuliert Corynebacterium glutamicum kompatible Solute durch Synthese und/oder Aufnahme aus dem umgebenden Medium. Die Aufnahme der kompatiblen Solute wird durch die vier sekundären Transportproteine BetP, EctP, ProP und LcoP vermittelt. BetP katalysiert den Transport von Betain im Symport mit zwei Natriumionen und ist auf Aktivitäts- und Expressionsebene reguliert. Um die osmosensorischen und osmoregulatorischen Eigenschaften des Carriers zu untersuchen, wurde BetP gereinigt und in Proteoliposomen aus Escherichia coli Phospholipiden rekonstituiert. Durch spezifische immunologische und proteolytische Analysen konnte der unidirektionale Einbau von BetP in die Liposomenmembran nachgewiesen werden. Die N- und C-terminale Extension ist im Lumen der Proteoliposomen lokalisiert. Untersuchungen zur Ionenselektivität der BetP-Aktivierung in Proteoliposomen ergaben, dass der Transporter spezifisch durch osmotisch induzierte Erhöhungen der internen Kaliumionenkonzentration aktiviert wird. Studien über die osmoregulatorischen und osmosensorischen Eigenschaften von C-terminal verkürzten BetP-Varianten lokalisieren die osmosensorische Domäne von BetP innerhalb der endständigen 25 Aminosäuren der C-terminalen Domäne. Durch ortsspezifische Mutagenese innerhalb dieser putativen Sensordomäne konnten Aminosäuren identifiziert werden, die am Mechanismus der K+-abhängigen BetP-Aktivierung beteiligt sind. Untersuchungen zur Osmoregulation von BetP in Proteoliposomen unterschiedlichen Lipidzusammensetzung ergaben, dass die Osmostress-abhängige Aktivität von BetP durch die Lipidumgebung moduliert werden kann. Die Analysen zur Osmosensorik von BetP in Proteoliposomen wurden durch osmoregulatorische Studien der BetP-Varianten in E. coli- und C. glutamicum-Zellen untermauert

Sperm Lipid Composition in Early Diverged Fish Species: Internal vs. External Mode of Fertilization

The lipid composition of sperm membranes is crucial for fertilization and differs among species. As the evolution of internal fertilization modes in fishes is not understood, a comparative study of the sperm lipid composition in freshwater representatives of externally and internally fertilizing fishes is needed for a better understanding of taxa-specific relationships between the lipid composition of the sperm membrane and the sperm physiology. The lipidomes of spermatozoa from stingray, a representative of cartilaginous fishes possessing internal fertilization, and sterlet, a representative of chondrostean fishes with external fertilization, have been studied by means of nuclear magnetic resonance (NMR), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), electrospray MS, gas chromatography-(GC) MS, and thin-layer chromatography (TLC). NMR experiments revealed higher cholesterol content and the presence of phosphatidylserine in stingray compared to sterlet sperm. Unknown MS signals could be assigned to different glycosphingolipids in sterlet (neutral glycosphingolipid Gal-Cer(d18:1/16:0)) and stingray (acidic glycosphingolipid sulpho-Gal-Cer(d18:1/16:0)). Free fatty acids in sterlet sperm indicate internal energy storage. GC-MS experiments indicated a significant amount of adrenic acid, but only a low amount of docosahexaenoic acid in stingray sperm. In a nutshell, this study provides novel data on sperm lipid composition for freshwater stingray and sterlet possessing different modes of fertilization

Structural modification and biological evaluation of Dmt1-DALDA analogues

The highly charged tetrapeptide Dmt-DALDA (Dmt-D-Arg-Phe-Lys-NH2) has been previously identified as a potent μ-opioid receptor agonist1 and serves as a lead compound for the further development of novel therapeutic (peptidic) opioid analgesics. The present work describes structural modifications of the peptide in order to determine the role of the charges, role of N-methylation, and role of conformation. All prepared compounds have been tested for their in vitro affinity and activity (guinea pig ileum GPI and mouse vas deferens MVD assays), their in vitro permeability (caco-2 test) and in vivo tissue distribution, in- and efflux into and out of mouse brain. These experimental data indicate that : i) side-chain charges are not essential for in vitro activity, ii) the guanidine group of D-Arg2 is important for the blood-brain permeability iii) the conformational constraint of the Phe residue by the benzazepine ring results in highly potent compounds, but is not compatible with the Lys side chain, which can best be removed for high potency. A more detailed discussion of the obtained results will be presented. References: 1. Shimoyama, M.; Szeto, H.H.; Schiller, P.W.; Tagaito, Y.; Tokairin, H.; Eun, C.M., Shimoyama N. Pharmacology 2008, 83, 33-37. 2. Zhao, K.; Luo, G., Zhao, G.-M.; Schiller, P.W.; Szeto, H.H. J. Pharmacol. Exp. Ther. 2003, 304, 425-432

Exploring the Nucleon Structure from First Principles of QCD

Quantum Chromodynamics (QCD) is generally assumed to be the fundamental theory underlying nuclear physics. In recent years there is progress towards investigating the nucleon structure from first principles of QCD. Although this structure is best revealed in Deep Inelastic Scattering, a consistent analysis has to be performed in a fully non-perturbative scheme. The only known method for this purpose are lattice simulations. We first sketch the ideas of Monte Carlo simulations in lattice gauge theory. Then we comment in particular on the issues of chiral symmetry and operator mixing. Finally we present our results for the Bjorken variable of a single quark, and for the second Nachtmann moment of the nucleon structure functions

In vivo antinociception of potent mu opioid agonist tetrapeptide analogues and comparison with a compact opioid agonist - neurokinin 1 receptor antagonist chimera

<p>Abstract</p> <p>Background</p> <p>An important limiting factor in the development of centrally acting pharmaceuticals is the blood-brain barrier (BBB). Transport of therapeutic peptides through this highly protective physiological barrier remains a challenge for peptide drug delivery into the central nervous system (CNS). Because the most common strategy to treat moderate to severe pain consists of the activation of opioid receptors in the brain, the development of active opioid peptide analogues as potential analgesics requires compounds with a high resistance to enzymatic degradation and an ability to cross the BBB.</p> <p>Results</p> <p>Herein we report that tetrapeptide analogues of the type H-Dmt¹-Xxx²-Yyy³-Gly⁴-NH₂are transported into the brain after intravenous and subcutaneous administration and are able to activate the μ- and δ opioid receptors more efficiently and over longer periods of time than morphine. Using the hot water tail flick test as the animal model for antinociception, a comparison in potency is presented between a side chain conformationally constrained analogue containing the benzazepine ring (BVD03, Yyy³: Aba), and a "ring opened" analogue (BVD02, Yyy³: Phe). The results show that in addition to the increased lipophilicity through amide bond N-methylation, the conformational constraint introduced at the level of the Phe³side chain causes a prolonged antinociception. Further replacement of NMe-D-Ala²by D-Arg²in the tetrapeptide sequence led to an improved potency as demonstrated by a higher and maintained antinociception for AN81 (Xxx²: D-Arg) vs. BVD03 (Xxx²: NMe-D-Ala). A daily injection of the studied opioid ligands over a time period of 5 days did however result in a substantial decrease in antinociception on the fifth day of the experiment. The compact opioid agonist - NK1 antagonist hybrid SBCHM01 could not circumvent opioid induced tolerance.</p> <p>Conclusions</p> <p>We demonstrated that the introduction of a conformational constraint has an important impact on opioid receptor activation and subsequent antinociception in vivo. Further amino acid substitution allowed to identify AN81 as an opioid ligand able to access the CNS and induce antinociception at very low doses (0.1 mg/kg) over a time period up to 7 hours. However, tolerance became apparent after repetitive i.v. administration of the investigated tetrapeptides. This side effect was also observed with the dual opioid agonist-NK1 receptor antagonist SBCHM01.</p

Defining a sustainable development target space for 2030 and 2050

With the establishment of the sustainable development goals (SDGs), countries worldwide agreed to a prosperous, socially inclusive, and environmentally sustainable future for all. This ambition, however, exposes a critical gap in science-based insights, namely on how to achieve the 17 SDGs simultaneously. Quantitative goal-seeking scenario studies could help explore the needed systems' transformations. This requires a clear definition of the "target space." The 169 targets and 232 indicators used for monitoring SDG implementation cannot be used for this; they are too many, too broad, unstructured, and sometimes not formulated quantitatively. Here, we propose a streamlined set of science-based indicators and associated target values that are quantifiable and actionable to make scenario analysis meaningful, relevant, and simple enough to be transparent and communicable. The 36 targets are based on the SDGs, existing multilateral agreements, literature, and expert assessment. They include 2050 as a longer-term reference point. This target space can guide researchers in developing new sustainable development pathways

Ligand Recognition of the Major Birch Pollen Allergen Bet v 1 is Isoform Dependent

Each spring millions of patients suffer from allergies when birch pollen is released into the air. In most cases, the major pollen allergen Bet v 1 is the elicitor of the allergy symptoms. Bet v 1 comes in a variety of isoforms that share virtually identical conformations, but their relative concentrations are plant-specific. Glycosylated flavonoids, such as quercetin-3-O-sophoroside, are the physiological ligands of Bet v 1, and here we found that three isoforms differing in their allergenic potential also show an individual, highly specific binding behaviour for the different ligands. This specificity is driven by the sugar moieties of the ligands rather than the flavonols. While the influence of the ligands on the allergenicity of the Bet v 1 isoforms may be limited, the isoform and ligand mixtures add up to a complex and thus individual fingerprint of the pollen. We suggest that this mixture is not only acting as an effective chemical sunscreen for pollen DNA, but may also play an important role in recognition processes during pollination