How Cations Change Peptide Structure

Specific interactions between cations and proteins have a strong impact on peptide and protein structure. We here shed light on the nature of the underlying interactions, especially regarding the effects on the polyamide backbone structure. To do so, we compare the conformational ensembles of model peptides in isolation and in the presence of either Li+ or Na+ cations by state-of-the-art density-functional theory (including van der Waals effects) and gas-phase infrared spectroscopy. These monovalent cations have a drastic effect on the local backbone conformation of turn-forming peptides, by disruption of the H bonding networks and the resulting severe distortion of the backbone conformations. In fact, Li+ and Na+ can even have different conformational effects on the same peptide. We also assess the predictive power of current approximate density functionals for peptide-cation systems and compare to results from established protein force fields as well as to high-level quantum chemistry (CCSD(T)).Comment: 30 pages, 7 figure

Combining ultra-high resolution ion-mobility spectrometry with cryogenic IR spectroscopy for the study of biomolecular ions

Double-resonance spectroscopic schemes in combination with cryogenic ion traps are the go-to techniques when isomer-specific high-resolution spectra are required for analysis of molecular ions. Their limitation lies in the requirement for well-resolved, isomer-specific absorption bands as well as in the potentially time-consuming steps to identify each isomer. We present an alternative approach where isomeric species are readily separated using ion mobility spectrometry (IMS) and selected prior cryogenic spectroscopic analysis. To date, most IMS approaches suffer from relatively low resolution, however, recent technological developments in the field of travelling-wave ion mobility using structures for lossless ion manipulation (SLIM) permit the use of extremely long drift paths, which greatly enhances the resolution. We demonstrate the power of combining this type of ultra-high resolution IMS with cryogenic vibrational spectroscopy by comparing mobility-resolved IR spectra of a disaccharide to those acquired using IR-IR double resonance. This new approach is especially promising for investigation of larger molecules where spectral congestion interferes with double resonance techniques

Gas-phase microsolvation of ubiquitin: investigation of crown ether complexation sites using ion mobility-mass spectrometry.

In this study the gas-phase structure of ubiquitin and its lysine-to-arginine mutants was investigated using ion mobility-mass spectrometry (IM-MS) and electron transfer dissociation-mass spectrometry (ETD-MS). Crown ether molecules were attached to positive charge sites of the proteins and the resulting non-covalent complexes were analysed. Collision induced dissociation (CID) experiments revealed relative energy differences between the wild type and the mutant crown-ether complexes. ETD-MS experiments were performed to identify the crown ether binding sites. Although not all of the binding sites could be revealed, the data confirm that the first crown ether is able to bind to the N-terminus. IM-MS experiments show a more compact structure for specific charge states of wild type ubiquitin when crown ethers are attached. However, data on ubiquitin mutants reveal that only specific lysine residues contribute to the effect of charge microsolvation. A compaction is only observed for one of the investigated mutants, in which the lysine has no proximate interaction partner. On the other hand when the lysine residues are involved in salt bridges, attachment of crown ethers has little effect on the structure

Die Berechnung der induzierten Ladung

One of the main aspects of statistical mechanics is that the properties of a thermodynamics state point do not depend on the choice of the statistical ensemble. It breaks down for small systems e.g. single molecules. Hence, the choice of the statistical ensemble is crucial for the interpretation of single molecule experiments, where the outcome of measurements depends on which variables or control parameters, are held fixed and which ones are allowed to fluctuate. Following this principle, this thesis investigates the thermodynamics of a single polymer pulling experiments within two different statistical ensembles. The scaling of the conjugate chain ensembles, the fixed end-to-end vector (Helmholtz) and the fixed applied force (Gibbs), are studied in depth. This thesis further investigates the ensemble equivalence for different force regimes and polymer-chain contour lengths. Using coarse-grained molecular dynamic simulations, i.e. Langevin dynamics, the simulations were found to complement the theoretical predictions for the scaling of ensemble difference of Gaussian chains in different force-regimes, giving special attention to the zero force regime. After constructing Helmholtz and Gibbs conjugate ensembles for a Gaussian chain, two different data sets of thermodynamic states on the force-extension plane, i.e. force-extension curves, were generated. The ensemble difference is computed for different polymer-chain lengths by using force-extension curves. The scaling of the ensemble difference versus relative polymer-chain length under different force regimes has been derived from the simulation data and compared to theoretical predictions. The results demonstrate that the Gaussian chain in the zero force limit generates nonequivalent ensembles, regardless of its equilibrium bond length and polymer-chain contour length. Moreover, if polymers are charged in confinement, coarse-graining is problematic, owing to dielectric interfaces. Hence, the effect of dielectric interfaces must be taken into account when describing physical systems such as ionic channels or biopolymers inside nanopores. It is shown that the effect of dielectrics is crucial for the dynamics of a biopolymer or an ion inside a nanopore. In the simulations, the feasibility of an efficient and accurate computation of electrostatic interactions in the presence of an arbitrarily shaped dielectric domain is challenging. Several solutions for this problem have been previously proposed in the literature such as a density functional approach, or transforming problem at hand into an algebraic problem ( Induced Charge Computation (ICC) ) and boundary element methods. Even though the essential concept is the same, which is to replace the dielectric interface with a polarization charge density, these approaches have been analyzed and the ICC algorithm has been implemented. A new superior boundary element method has been devised utilizing the force computation via the Particle-Particle Particle-Mesh (P3M) method for periodic geometries (ICCP3M). This method has been compared to the ICC algorithm, the algebraic solutions, and to density functional approaches. Extensive numerical tests against analytically tractable geometries have confirmed the correctness and applicability of developed and implemented algorithms, demonstrating that the ICCP3M is the fastest and the most versatile algorithm. Further optimization issues are also discussed in obtaining accurate induced charge densities. The potential of mean force (PMF) of DNA modelled on a coarsed-grain level inside a nanopore is investigated with and without the inclusion of dielectric effects. Despite the simplicity of the model, the dramatic effect of dielectric inclusions is clearly seen in the observed force profile.Eines der wichtigsten Ergebnisse der statistischen Mechanik ist, dass unterschiedliche statistische Ensembles dieselben thermodynamischen Zustände erzeugen. Dieses Prinzip gilt nicht notwendigerweise für kleine Systeme, wie zum Beispiel einzelne Moleküle oder ein einzelnes Polymer. Deshalb ist die Wahl des statistischen Ensembles von entscheidender Bedeutung für die Interpretation von Einzelmolekülexperimenten ( im Englischen "Single Molecule Experiment" (SME) ), denn das Ergebnis der Messung hängt davon ab, welche Variablen oder Kontrollparameter festgehalten werden und welche fluktuieren können. Ausgehend von diesem Problem haben wir Zugexperimente an einem einzigen Polymer in zwei verschiedenen Ensembles durchgeführt und den thermodynamischen Limes (Anzahl der Polymersegmente wächst gegen unendlich) untersucht. Wir haben zwei konjugierte Ensembles, nämlich das, in dem der End-zu-End Abstand (Helmholtz) festgehalten wurde, mit dem, wo wir die Kraft (Gibbs) festgehalten haben, gründlich und auf verschiedene Arten verglichen. Wir haben den Ensemble-Unterschied als Funktion der Anzahl der Polymersegmente in unterschiedlichen Zugkraftbereichen mittels Molekulardynamik Simulationen untersucht, wobei wir eine Langevin Dynamik benutzt haben. Die untersuchten Messgrössen waren die Bestimmung von sogenannten Kraft-Dehnungskurven, wie sie auch in AFM Experimenten gemessen werden. Diese Kurven wurden für zwei verschieden Gauss Ketten verschiedenster Polymerlänge durchgeführt, einmal mit verschwindender Bondlänge und einmal mit Bondlänge eins. Aufgrund unserer Simulationen konnten wir zeigen, das sowohl Gauss-Ketten mit endlicher, wie auch verschwindender Bondlänge für den Bereich verschwindender Zugkraft einen endlichen Ensembleunterschied besitzen, der nicht von der Kettenlänge abhängt. Dieses Phänomen wurde bereits vor 20 Jahren von R. Neumann beschrieben. Trotz der relativ einfachen Argumente von Neumann gibt es bis heute noch Arbeiten, die diesen Sachverhalt entweder anzweifeln oder verkehrt darstellen. Wir hoffen, durch diesen Teil der Arbeit den Sachverhalt zufriedenstellend aufgeklärt zu haben. Im zweiten Teil der Arbeit behandeln wir geladen Polymere unter einem räumlichen Einschluss. Dies können zum Beispiel Ionen in schmalen Kanälen sein (Ionenkanäle), oder DNA in Nanoporen. In vergröberten Simulationen werden geladene Polymere immer in einem dielektrischen Kontinuum dargestellt. Wasser hat eine relative dielektrische Konstante von 80 bei Raumtemperatur, die dann in dieses Model als Parameter gesteckt wird. Wenn feste Grenzflächen vorhanden sind, haben diese meist niedrige dielektrische Konstanten ($\approx 2$). Diese Grenzflächen haben grosse Auswirkungen auf die elektrostatischen Wechselwirkungen. In den Simulationen ist es wichtig, diese Effekte korrekt *und schnell* zu berechnen. Deshalb haben wir einen effizienten und präzisen Algorithmus entwickelt, der genau dies bewerkstelligt. In der Literatur wurden mehrere Möglichkeiten vorgeschlagen, wie dieses Problem für Simulationen lösbar sein sollte, wie zum Beispiel Dichtefunktionalmethoden, Umwandlung des Problems in ein algebraisches Problem (Induced Charge Computation, ICC) oder die Randelement Methode. Das wesentliche Konzept besteht darin, die Polarisationsladung auf dem dielektrischen Rand so zu bestimmen, dass die dielektrischen Randbedingungen erfüllt werden. Wir haben den ICCP3M Algorithmus entwickelt, dessen Kernstück darin besteht, den P3M Algorithmus zur Bestimmung der induzierten Ladung auf den Randelementen zu benutzen. Durch diesen Trick lässt sich die Ladungsberechnung in CPU Zeit $\mathscr{O}(Nlog N)$, wobei $\mathscr{O}(N)$ die Anzahl der Ladungen im System ist, durchführen. Wir haben den Algorithmus innerhalb des Espresso Programmpakets implementiert und optimiert. Im letzten Teil der Arbeit wurde das Potential der mittleren Kraft einer vergröberten DNA innerhalb einer Nanopore untersucht, wobei wir die Unterschiede zwischen korrekter Behandung der dielektrischen Ränder und der Ignorierung derselben quantifiziert haben. Trotz seiner Einfachheit zeigt unser Modell den dramatischen Einfluss, den die dielektrischen Ränder auf die gemessene efffektive Kraft und das Potential der mittleren Kraft ausüben

Protomers of Benzocaine: Solvent and Permittivity Dependence

The immediate environment of a molecule can have a profound influence on its properties. Benzocaine, the ethyl ester of para-aminobenzoic acid, which finds an application as a local anesthetic (LA), is found to adopt in its protonated form at least two populations of distinct structures in the gas phase and their relative intensities strongly depend on the properties of the solvent used in the electrospray ionization (ESI) process. Here we combine IR-vibrational spectroscopy with ion mobility-mass spectrometry (IM-MS) to yield gas-phase IR spectra of simultaneously m/z and drift-time resolved species of benzocaine. The results allow for an unambiguous identification of two protomeric species - the N- and O-protonated form. Density functional theory (DFT) calculations link these structures to the most stable solution and gas-phase structures, respectively, with the electric properties of the surrounding medium being the main determinant for the preferred protonation site. The fact that the N-protonated form of benzocaine can be found in the gas phase is owed to kinetic trapping of the solution phase structure during transfer into the experimental setup. These observations confirm earlier studies on similar molecules where N- and O-protonation has been suggested

Increased human defensine levels hint at an inflammatory etiology of bisphosphonate-associated osteonecrosis of the jaw: An immunohistological study

<p>Abstract</p> <p>Background</p> <p>Human β-defensins (hBD) are antimicrobial peptides that are an integral part of bone innate immunity. Recently, it could be shown that expression of hBD-1, -2 and -3 were upregulated in cases of osteomyelitis of the jaws. In order to gain insight into the possible impairment of hBD metabolism in bisphosphonate-associated osteonecrosis of the jaws (BONJ), the present exploratory study was designed so as to determine the qualitative and quantitative expression of afore mentioned hBDs in BONJ and infected osteoradionecrosis (ORN), both of which represent inflammatory bone diseases.</p> <p>Methods</p> <p>Bone samples were collected from patients with BONJ (n = 20) and ORN (n = 20). Non-infected healthy bone samples (n = 20) were included as controls. Immunohistological staining in an autostainer was carried out by the (Strept-ABC)-method against hBD-1,-2,-3. Specific positive vs. negative cell reaction of osteocytes (labeling index) near the border of bony resection was determined and counted for quantitative analysis. Number of vital osteocytes vs. empty osteocytes lacunae was compared between groups.</p> <p>Results</p> <p>hBD-1,-2 and -3 could be detected in BONJ as well as ORN and healthy bone samples. Immunoreactivity against hBD-2 and -3 was significantly higher in BONJ than in ORN and healthy jaw bone samples. Number of empty osteocyte lacunae was significantly higher in ORN compared with BONJ (<it>P </it>= 0.001).</p> <p>Conclusion</p> <p>Under the condition of BONJ an increased expression of hBD-1,-2,-3 is detectable, similarly to the recently described upregulation of defensins in chronically infected jaw bones. It remains still unclear how these findings may relate to the pathoetiology of these diseases and whether this is contributing to the development of BONJ and ORN or simply an after effect of the disease.</p

The Anglo-Saxon migration and the formation of the early English gene pool

The history of the British Isles and Ireland is characterized by multiple periods of major cultural change, including the influential transformation after the end of Roman rule, which precipitated shifts in language, settlement patterns and material culture1. The extent to which migration from continental Europe mediated these transitions is a matter of long-standing debate2,3,4. Here we study genome-wide ancient DNA from 460 medieval northwestern Europeans—including 278 individuals from England—alongside archaeological data, to infer contemporary population dynamics. We identify a substantial increase of continental northern European ancestry in early medieval England, which is closely related to the early medieval and present-day inhabitants of Germany and Denmark, implying large-scale substantial migration across the North Sea into Britain during the Early Middle Ages. As a result, the individuals who we analysed from eastern England derived up to 76% of their ancestry from the continental North Sea zone, albeit with substantial regional variation and heterogeneity within sites. We show that women with immigrant ancestry were more often furnished with grave goods than women with local ancestry, whereas men with weapons were as likely not to be of immigrant ancestry. A comparison with present-day Britain indicates that subsequent demographic events reduced the fraction of continental northern European ancestry while introducing further ancestry components into the English gene pool, including substantial southwestern European ancestry most closely related to that seen in Iron Age France5,6

Author Correction: The Anglo-Saxon migration and the formation of the early English gene pool.

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The Anglo-Saxon migration and the formation of the early English gene pool.

The history of the British Isles and Ireland is characterized by multiple periods of major cultural change, including the influential transformation after the end of Roman rule, which precipitated shifts in language, settlement patterns and material culture1. The extent to which migration from continental Europe mediated these transitions is a matter of long-standing debate2-4. Here we study genome-wide ancient DNA from 460 medieval northwestern Europeans-including 278 individuals from England-alongside archaeological data, to infer contemporary population dynamics. We identify a substantial increase of continental northern European ancestry in early medieval England, which is closely related to the early medieval and present-day inhabitants of Germany and Denmark, implying large-scale substantial migration across the North Sea into Britain during the Early Middle Ages. As a result, the individuals who we analysed from eastern England derived up to 76% of their ancestry from the continental North Sea zone, albeit with substantial regional variation and heterogeneity within sites. We show that women with immigrant ancestry were more often furnished with grave goods than women with local ancestry, whereas men with weapons were as likely not to be of immigrant ancestry. A comparison with present-day Britain indicates that subsequent demographic events reduced the fraction of continental northern European ancestry while introducing further ancestry components into the English gene pool, including substantial southwestern European ancestry most closely related to that seen in Iron Age France5,6