MS/MS and associated database search algorithms are essential proteomic tools for identifying peptides. Due to their widespread use, it is now time to perform a systematic analysis of the various ...algorithms currently in use. Using blood specimens used in the HUPO Plasma Proteome Project, we have evaluated five search algorithms with respect to their sensitivity and specificity, and have also accurately benchmarked them based on specified false‐positive (FP) rates. Spectrum Mill and SEQUEST performed well in terms of sensitivity, but were inferior to MASCOT, X!Tandem, and Sonar in terms of specificity. Overall, MASCOT, a probabilistic search algorithm, correctly identified most peptides based on a specified FP rate. The rescoring algorithm, PeptideProphet, enhanced the overall performance of the SEQUEST algorithm, as well as provided predictable FP error rates. Ideally, score thresholds should be calculated for each peptide spectrum or minimally, derived from a reversed‐sequence search as demonstrated in this study based on a validated data set. The availability of open‐source search algorithms, such as X!Tandem, makes it feasible to further improve the validation process (manual or automatic) on the basis of “consensus scoring”, i.e., the use of multiple (at least two) search algorithms to reduce the number of FPs.∁
is one of the main causative agents of nosocomial infections and the spread of multidrug-resistant strains is rising. Therefore, novel strategies for therapy are urgently required. The outer membrane ...composition of Gram-negative pathogens and especially of
restricts the efficacy of antibiotic entry into the cell and determines virulence. For efficient outer membrane protein biogenesis, the β-barrel assembly machinery (BAM) complex in the outer membrane and periplasmic chaperones like Skp and SurA are crucial. Previous studies indicated that the importance of individual proteins involved in outer membrane protein biogenesis may vary between different Gram-negative species. In addition, since multidrug-resistant
strains pose a serious global threat, the interference with both virulence and antibiotic resistance by disturbing outer membrane protein biogenesis might be a new strategy to cope with this challenge. Therefore, deletion mutants of the non-essential BAM complex components
and
, of the
homolog
as well as a conditional mutant of
were investigated. The most profound effects for both traits were associated with reduced levels of SurA, characterized by increased membrane permeability, enhanced sensitivity to antibiotic treatment and attenuation of virulence in a
infection model. Strikingly, the depletion of SurA in a multidrug-resistant clinical bloodstream isolate re-sensitized the strain to antibiotic treatment. From our data we conclude that SurA of
serves as a promising target for developing a drug that shows antiinfective activity and re-sensitizes multidrug-resistant strains to antibiotics.
In the course of the preliminary planning of the new Gelnhausen–Fulda railway line which is part of the railway project Hanau–Würzburg/Fulda, two route options with tunnels, bridges, cuts and ...embankments are to be investigated and assessed as efficiently as possible with regard to potential geotechnical risk areas. Due to the geological and hydrogeological conditions within the north‐eastern Hessian Buntsandsteingebirge, several groundwater levels, landslides in tunnel portal areas and karstification structures are to be expected on the two route options, each of which is approx. 45 km long. In order to obtain detailed and, above all, large‐scale knowledge of geotechnical hazards of each route = in the preliminary stage of the exploration programme, geophysical surface investigations are a cost‐ and time‐optimized exploration method in addition to standard drilling. These findings as well as existing subsurface information are combined by means of BIM, so that risk areas can be identified at an early stage of the project and taken into account when selecting the most affordable route.
Im Zuge der Vorplanung der DB‐Neubaustrecke Gelnhausen–Fulda als Teilmaßnahme des Projekts Hanau–Würzburg/Fulda sollen zwei Trassenvarianten mit Tunneln, Brücken, Einschnitten sowie Dämmen untersucht und möglichst effizient hinsichtlich potenzieller geotechnischer Risikobereiche abgeschätzt werden. Aufgrund der geologischen und hydrogeologischen Gegebenheiten innerhalb des nordosthessischen Buntsandsteingebirges ist auf den beiden zu untersuchenden jeweils ca. 45 km langen Trassen mit etlichen Grundwasserstockwerken, Rutschungen in Portalbereichen und Verkarstungsstrukturen zu rechnen. Um bereits in der Vorstufe des Erkundungsprogramms detaillierte und vor allem großräumige Erkenntnisse über die geotechnischen Gefährdungsbilder der einzelnen Trassen zu erhalten, bieten sich zusätzlich zu einem punktuellen Bohrprogramm oberflächengeophysikalische Untersuchungen Erkundungen an. Diese Erkenntnisse sowie geologische und hydrogeoglogische Bestandsunterlagen werden in einem BIM‐Gesamtmodell eingebunden, sodass Risikobereiche möglichst frühzeitig erkannt und bei der Trassenoptimierung berücksichtigt werden können.
The precise spatial localization of single molecules in three dimensions is an important basis for single molecule localization microscopy (SMLM) and tracking. At distances up to a few hundred ...nanometers from the coverslip, evanescent wave coupling into the glass, also known as supercritical angle fluorescence (SAF), can strongly improve the axial precision, thus facilitating almost isotropic localization performance. Specific detection systems, introduced as
(SALM) or
(DONALD), have been developed to exploit SAF in modified two-channel imaging schemes. Recently, our group has shown that off-focus microscopy, i.e., imaging at an intentional slight defocus, can perform equally well, but uses only a single detection arm. Here we compare SALM, off-focus imaging and the most commonly used 3D SMLM techniques, namely cylindrical lens and biplane imaging, regarding 3D localization in close proximity to the coverslip. We show that all methods gain from SAF, which leaves a high detection NA as the only major key requirement to unlock the SAF benefit. We find parameter settings for cylindrical lens and biplane imaging for highest z-precision. Further, we compare the methods in view of robustness to aberrations, fixed dipole emission and double-emitter events. We show that biplane imaging provides the best overall performance and support our findings by DNA-PAINT experiments on DNA-nanoruler samples. Our study sheds light on the effects of SAF for SMLM and is helpful for researchers who plan to employ localization-based 3D nanoscopy close to the coverslip.
Little is known about optimally applying chemotherapeutic agents in a specific temporal sequence to rapidly reduce the tumor load and to improve therapeutic efficacy. The clinical optimization of ...drug efficacy while reducing side effects is still restricted due to an incomplete understanding of the mode of action and related tumor relapse mechanisms on the molecular level. The molecular characterization of transcriptomic drug signatures can help to identify the affected pathways, downstream regulated genes and regulatory interactions related to tumor relapse in response to drug application. We tried to outline the dynamic regulatory reprogramming leading to tumor relapse in relapsed MLL-rearranged pro-B-cell acute lymphoblastic leukemia (B-ALL) cells in response to two first-line treatments: dexamethasone (Dexa) and cytarabine (AraC). We performed an integrative molecular analysis of whole transcriptome profiles of each treatment, specifically considering public knowledge of miRNA regulation via a network-based approach to unravel key driver genes and miRNAs that may control the relapse mechanisms accompanying each treatment. Our results gave hints to the crucial regulatory roles of genes leading to Dexa-resistance and related miRNAs linked to chemosensitivity. These genes and miRNAs should be further investigated in preclinical models to obtain more hints about relapse processes.
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•The surface state of Mn5Ge3 exposed to air and common colvents, is analyzed by XPS.•The surface free energy of Mn5Ge3 is calculated with the OWRK method.•The grafting of Mn5Ge3 films ...by octane- and perfluorodecane-thiol SAMs is proposed.
Hybrid organic/inorganic interfaces could pave the way to chemically designed or new multifunctional electronic devices, in particular in the spintronics field where, for instance, the interfacial spin polarization can be tuned through chemical interactions and surface modifications. We report herein, for the very first time, the assays of self-assembled monolayers (SAMs) formation on Mn5Ge3 surface. Interestingly, Mn5Ge3 is a ferromagnetic metal possessing interesting features for spintronics such as a high Curie temperature, a capability to grow epitaxially on Ge germanium that is a key point for integration in the mainstream Si technology. Mn5Ge3 thin films are synthesized by molecular beam epitaxy under ultra-vacuum condition. We studied the Mn5Ge3 surface prior to deposition of SAMs, meaning the surface exposed to air and different solvents, by XPS and contact angle measurements leading to the value of the surface tension of this surface. Then SAMs of octanethiol and perfluorodecanethiol are formed on Mn5Ge3 surface at room temperature. The best experimental conditions to form the SAMs are found for an immersion time of 36 h and a concentration of 4 mM.
The authors would like to note an error in Figures 1 and 2 of this paper. The graph in Figure 1 incorrectly reflected the overall survival (OS), when it should have displayed the progression-free ...survival (PFS). The caption and median PFS values were correct.
•Severely affected youth with comorbid social phobia (SP) and major depressive disorder (MDD) show reduced reward anticipation and processing compared to typically developing controls.•In the ...clinical group higher reward anticipation correlates with high SP symptoms and lower anticipation with high MDD symptoms.•Results are in line with the theory of heightened vigilance in anxiety and blunted reward processing due to anhedonia in MDD.
Impaired reward processing has been found in individuals with anxiety, but also major depressive disorder (MDD). Here, we studied neural correlates of reward anticipation and processing in a sample of youth with severe social phobia and comorbid depression (SP/MDD) and investigated the specific contribution of SP and MDD symptoms.
15 affected, unmedicated and 25 typically developing (TD) youth completed a monetary gambling task, which included a positive, negative and ambiguous reward condition. Event-related potentials representing cue processing (cue P300), reward anticipation (stimulus preceding negativity, SPN), reward sensitivity (feedback related negativity, FRN) and reward processing (reward P300) were analysed.
Reduced amplitudes of the right hemispheric (r)SPN and reward P300 were observed in SP/MDD compared to TD. Within the SP/MDD group SP symptoms correlated with larger rSPN, and FRN amplitudes. MDD symptoms correlated with smaller rSPN and smaller FRN positive–negative difference wave.
Reward anticipation and feedback processing are reduced in SP/MDD. Higher SP symptoms are associated with stronger neural activation during reward anticipation and reward sensitivity. Depressive symptoms are associated with decreased reward anticipation and sensitivity. Findings are in line with the theory of heightened vigilance in anxiety and blunted reward processing due to anhedonia in MDD.
The study results can inform behavioural interventions for SP and MDD.
Extensive prefractionation is now considered to be a necessary prerequisite for the comprehensive analysis of complex proteomes where the dynamic range of protein abundances can vary from ∼106 for ...cells to ∼1010 for tissues such as blood. Here, we describe a high-resolution 2D protein separation system that uses a continuous free-flow electrophoresis (FFE) device to fractionate complex protein mixtures by solution-phase isoelectric focusing (IEF) into 96 well-defined pools, each separated by ∼0.02−0.10 pH unit depending on the gradient created, followed by rapid (∼6 min per analysis) reversed-phase high-performance liquid chromatography (RP-HPLC) of each FFE pool. Fractionated proteins are readily visualized in a virtual 2D format using software that plots protein loci, pI in the first dimension and relative hydrophobicity (i.e., RP-HPLC retention time) in the second dimension. By coupling a diode-array detector in line with a multiwavelength fluorescence detector, separated proteins can be monitored in the RP-HPLC eluent by both UV absorbance and intrinsic fluorescence simultaneously from a single experiment. Triplicate analyses of standard proteins using a pH 3−10 gradient conducted over a 3-day period revealed a high system reproducibility with a SD of 0.57 (0.05 pH unit) within the FFE pools and 0.003 (0.18 s) for protein retention times in the second-dimension RP-HPLC step. In addition, we demonstrate that the FFE-IEF/RP-HPLC separation strategy can also be applied to complex mixtures of low molecular weight compounds such as peptides. With the facile ability to measure the pH of the isoelectric focused pools, peptide pI values can be estimated and used to qualify peptide identifications made using either MS/MS sequencing approaches or pI discriminated peptide mass fingerprinting. The calculated peak capacity of this 2D liquid-based FFE-IEF/RP-HPLC system is 6720.
Urban-living individuals are exposed to many environmental factors that may combine and interact to influence mental health. While individual factors of an urban environment have been investigated in ...isolation, no attempt has been made to model how complex, real-life exposure to living in the city relates to brain and mental health, and how this is moderated by genetic factors. Using the data of 156,075 participants from the UK Biobank, we carried out sparse canonical correlation analyses to investigate the relationships between urban environments and psychiatric symptoms. We found an environmental profile of social deprivation, air pollution, street network and urban land-use density that was positively correlated with an affective symptom group (r = 0.22, P
< 0.001), mediated by brain volume differences consistent with reward processing, and moderated by genes enriched for stress response, including CRHR1, explaining 2.01% of the variance in brain volume differences. Protective factors such as greenness and generous destination accessibility were negatively correlated with an anxiety symptom group (r = 0.10, P
< 0.001), mediated by brain regions necessary for emotion regulation and moderated by EXD3, explaining 1.65% of the variance. The third urban environmental profile was correlated with an emotional instability symptom group (r = 0.03, P
< 0.001). Our findings suggest that different environmental profiles of urban living may influence specific psychiatric symptom groups through distinct neurobiological pathways.
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