Engineered Sortases in Peptide and Protein Chemistry

The transpeptidase sortase A of Staphylococcus aureus (Sa-SrtA) is a valuable tool in protein chemistry. The native enzyme anchors surface proteins containing a highly conserved LPxTG sorting motif to a terminal glycine residue of the peptidoglycan layer in Gram-positive bacteria. This reaction is exploited for sortase-mediated ligation (SML), allowing the site-specific linkage of synthetic peptides and recombinant proteins by a native peptide bond. However, the moderate catalytic efficiency and specificity of Sa-SrtA fueled the development of new biocatalysts for SML, including the screening of sortase A variants form microorganisms other than S. aureus and the directed protein evolution of the Sa-SrtA enzyme itself. Novel display platforms and screening formats were developed to isolate sortases with altered properties from mutant libraries. This yielded sortases with strongly enhanced catalytic activity and enzymes recognizing new sorting motifs as substrates. This minireview focuses on recent advances in the field of directed sortase evolution and applications of these tailor-made enzymes in biochemistry

Limitations of Boulder Detection in Shallow Water Habitats Using High-Resolution Sidescan Sonar Images

Stones and boulders in shallow waters (0–10 m water depth) form complex geo-habitats, serving as a hardground for many benthic species, and are important contributors to coastal biodiversity and high benthic production. This study focuses on limitations in stone and boulder detection using high-resolution sidescan sonar images in shallow water environments of the southwestern Baltic Sea. Observations were carried out using sidescan sonars operating with frequencies from 450 kHz up to 1 MHz to identify individual stones and boulders within different levels of resolution. In addition, sidescan sonar images were generated using varying survey directions for an assessment of range effects. The comparison of images of different resolutions reveals considerable discrepancies in the numbers of detectable stones and boulders, and in their distribution patterns. Results on the detection of individual stones and boulders at approximately 0.04 m/pixel resolution were compared to common discretizations: it was shown that image resolutions of 0.2 m/pixel may underestimate available hard-ground settlement space by up to 42%. If methodological constraints are known and considered, detailed information about individual stones and boulders, and potential settlement space for marine organisms, can be derived

Dynamics of Stone Habitats in Coastal Waters of the Southwestern Baltic Sea (Hohwacht Bay)

Cobbles and boulders on the seafloor are of high ecological value in their function as habitats for a variety of benthic species, contributing to biodiversity and productivity in marine environments. We investigate the origin, physical shape, and structure of habitat-forming cobbles and boulders and reflect on their dynamics in coastal environments of the southwestern Baltic Sea. Stone habitats are not limited to lag deposits and cannot be sufficiently described as static environments, as different dynamic processes lead to changes within the physical habitat structure and create new habitats in spatially disparate areas. Dynamic processes such as (a) ongoing exposure of cobbles and boulders from glacial till, (b) continuous overturning of cobbles, and (c) the migration of cobbles need to be considered. A distinction between allochthonous and autochthonous habitats is suggested. The genesis of sediment types indicates that stone habitats are restricted to their source (glacial till), but hydrodynamic processes induce a redistribution of individual cobbles, leading to the development of new coastal habitats. Thus, coastal stone habitats need to be regarded as dynamic and are changing on a large bandwidth of timescales. In general, wave-induced processes changing the physical structure of these habitats do not occur separately but rather act simultaneously, leading to a dynamic type of habitat

Europa muss Führungsanspruch zeigen: Bilanz der Münchner Sicherheitskonferenz 2020

Nach der Münchner Sicherheitskonferenz (MSC) 2020 liegen die Probleme und Risiken auf dem Tisch. Viel intellektuelle Kraft floss in die Analyse, wie komplex und risikobehaftet die weltweite Sicherheitslage ist. Die Diskussion über kommende Sicherheitsrisiken etwa im Cyberraum, durch die rasante Entwicklung von Dual-Use-Technologien oder auch durch den Klimawandel und seine Folgen spezialisiert sich, zu Recht, immer weiter. Doch beunruhigend ist, wie wenig gleichgerichtete politische Energie zur Bewältigung der Herausforderungen zur Verfügung steht

Improving the proteome coverage of Daphnia magna - implications for future ecotoxicoproteomics studies

Aquatic pollution is an increasing problem and requires extensive research efforts to understand associated consequences and to find suitable solutions. The crustacean Daphnia is a keystone species in lacustrine ecosystems by connecting primary producers with higher trophic levels. Therefore, Daphnia is perfectly suitable to investigate biological effects of freshwater pollution and is frequently used as an important model organism in ecotoxicology. The field of ecotoxicoproteomics has become increasingly prevalent, as proteins are important for an organism's physiology and respond rapidly to changing environmental conditions. However, one obstacle in proteome analysis of Daphnia is highly abundant proteins like vitellogenin, decreasing the analytical depth of proteome analysis. To improve proteome coverage in Daphnia, we established an easy-to-use procedure based on the LC-MS/MS of whole daphnids and the dissected Daphnia gut, which is the main tissue getting in contact with soluble and particulate pollutants, separately. Using a comprehensive spectral library, generated by gas-phase fractionation and a data-independent acquisition method, we identified 4621 and 5233 protein groups at high confidence (false discovery rate < 0.01) in Daphnia and Daphnia gut samples, respectively. By combining both datasets, a proteome coverage of 6027 proteins was achieved, demonstrating the effectiveness of our approach

Gemeinsam stärker: eine Strategie zur Wiederbelebung des transatlantischen Verhältnisses; deutsche Kurzfassung

Die Harvard Kennedy School (HKS) und die Deutsche Gesellschaft für Auswärtige Politik (DGAP) haben im vergangenen Jahr eine Strategiegruppe amerikanischer und europäischer Expertinnen und Experten und ehemaliger Regierungsverantwortlicher gebildet, um die Krise der transatlantischen Beziehungen zu diskutieren und eine Strategie zu ihrer Wiederbelebung und Stärkung zu entwickeln

Die Tagung der Angespannten: nach der Münchner Sicherheitskonferenz

Deutschlands außen- und sicherheitspolitische Entscheiderinnen und Entscheider sind derzeit getrieben von neuen Realitäten, etwa durch technologische Entwicklungen, Risse in den Rüstungskontrollregimen und die Perspektive möglicher Rüstungswettläufe. Der Zerfall der internationalen Ordnung im strategischen Wettbewerb der Großmächte sowie die innere und äußere Schwäche der EU erschüttern den Glauben an die eigene Gestaltungskraft. Bei der MSC deutete sich Bewegung an, doch Entspannung ist nicht geboten

Sortase-Mediated Multi-Fragment Assemblies by Ligation Site Switching

Sortase-mediated ligation (SML) is a powerful tool of protein chemistry allowing the ligation of peptides containing LPxTG sorting motifs and N-terminal glycine nucleophiles. The installation of a sorting motif into the product prohibits the assembly of multiple fragments by SML. Here we report multi-fragment SML based on switchable sortase substrates. Substitution of the Leu residue by disulfide-containing Cys(StBu) results in active sorting motifs, which are inactivatable by reduction. In combination with a photo-protected N-Gly nucleophile, multi-fragment SML is enabled by repetitive cycles of SML and ligation site switching. The feasibility of this approach was demonstrated by a proof-of-concept four-fragment ligation, the assembly of peptide probes for bivalent chromatin binding proteins and oligomerization of peptide antigens. Biochemical and immuno-assays demonstrated functionality of these probes rendering them promising tools for immunology and chromatin biochemistry

Living systematic review and meta-analysis of the prostate MRI diagnostic test with Prostate Imaging Reporting and Data System (PI-RADS) assessment for the detection of prostate cancer:study protocol

INTRODUCTION: The Prostate Imaging Reporting and Data System (PI-RADS) standardises reporting of prostate MRI for the detection of clinically significant prostate cancer. We provide the protocol of a planned living systematic review and meta-analysis for (1) diagnostic accuracy (sensitivity and specificity), (2) cancer detection rates of assessment categories and (3) inter-reader agreement. METHODS AND ANALYSIS: Retrospective and prospective studies reporting on at least one of the outcomes of interest are included. Each step that requires literature evaluation and data extraction is performed by two independent reviewers. Since PI-RADS is intended as a living document itself, a 12-month update cycle of the systematic review and meta-analysis is planned. This protocol is in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses—Protocols statement. The search strategies including databases, study eligibility criteria, index and reference test definitions, outcome definitions and data analysis processes are detailed. A full list of extracted data items is provided. Summary estimates of sensitivity and specificity (for PI-RADS ≥3 and PI-RADS ≥4 considered positive) are derived with bivariate binomial models. Summary estimates of cancer detection rates are calculated with random intercept logistic regression models for single proportions. Summary estimates of inter-reader agreement are derived with random effects models. ETHICS AND DISSEMINATION: No original patient data are collected, ethical review board approval, therefore, is not necessary. Results are published in peer-reviewed, open-access scientific journals. We make the collected data accessible as supplemental material to guarantee transparency of results. PROSPERO REGISTRATION NUMBER: CRD42022343931