Biology subjects

Köhler, J.

Publications and source records attributed to Köhler, J..

5 recordsLinked to original sources

bioRxiv2026

Laudanosine restricts Ebola virus entry by targeting TPC2-dependent endolysosomal trafficking

Late endosome-dependent viruses, including filo- and arenaviruses, rely on host endolysosomal trafficking for productive infection. Here, we used a dual-colour Vesicular stomatitis virus (VSV) based pseudoparticle screen of CytoSorb-derived fractions to identify inhibitors of the Zaire Ebolavirus glycoprotein (GP)-mediated entry. Iterative chromatographic purification and mass spectrometry identified Laudanosine, a degradation product of the clinically used neuromuscular blocker Atracurium, as the antiviral compound. Laudanosine specifically inhibited entry mediated by Ebola, Marburg, Lymphocytic choriomeningitis and Lassa virus glycoproteins without affecting VSV-G-dependent entry. Importantly, Laudanosine inhibited authentic Ebola virus infection without detectable cytotoxicity in cell culture and embryonic zebrafish. Molecular dynamics simulations suggest stable association of Laudanosine with the allosteric inhibitory pocket of the lysosomal two-pore channel (TPC2). Consistently, Laudanosine impairs autophagic flux and disrupts endolysosomal trafficking. Together, our findings identify Laudanosine as a previously unrecognised inhibitor of TPC2-dependent entry of highly lethal viral pathogens.

microbiology↗

bioRxiv2026

Molecular Basis of Angicin Activity

Angicin is a class IId bacteriocin produced by Streptococcus anginosus with activity against Gram-positive pathogens, including Listeria monocytogenes and vancomycin-resistant Enterococcus faecium. While the mannose phosphotransferase system (Man-PTS) has been identified as a receptor in L. monocytogenes, its role in streptococci and the structural determinants of Angicin activity remain unclear. Here, we demonstrate that the Man-PTS is required for Angicin susceptibility in Streptococcus constellatus. A transposon mutant (manM::ISS1) showed complete resistance to Angicin and impaired mannose utilization. Structure-activity relationship analysis of truncated and modified peptides localized antimicrobial activity to the C-terminal region, although none of the variants matched the activity of the full-length peptide. Angicin induced membrane depolarization and pore formation in target bacteria. Residual activity in Man-PTS-impaired L. monocytogenes suggests an additional receptor-independent effect at higher concentrations. In vivo toxicity analysis using zebrafish embryos showed low toxicity at active concentrations. These findings identify the Man-PTS as a receptor for Angicin in streptococci and define structural features associated with its antimicrobial activity.

microbiology↗

bioRxiv2024

The haplotype-resolved Prymnesium parvum (type B) microalga genome reveals the genetic basis of its fish-killing toxins

The catastrophic loss of aquatic life in the Central European Oder River in 2022, caused by a toxic bloom of the haptophyte microalga Prymnesium parvum (in a wide sense, s.l.), underscores the need to improve our understanding of the genomic basis of the toxin. Previous morphological, phylogenetic, and genomic studies have revealed cryptic diversity within P. parvum s.l. and uncovered three clade-specific (types A, B, C) prymnesin toxins. Here, we used state-of-the-art long-read sequencing and assembled the first haplotype-resolved diploid genome of a P. parvum type B, the strain responsible for the Oder disaster. Comparative analyses with type A genomes uncovered a genome-size expansion driven by repetitive elements in type B. We also found conserved chromosomal synteny but divergent evolution in several polyketide synthase (PKS) genes, which are known to underlie toxin production in combination with environmental cues. We identified a specific, approximately 20 kilobase pair comprising deletion in the largest PKS gene of type B that we link to differences of the chemical structure of types A and B prymnesins. Electron-microscopy and flow cytometry confirmed diploidy in the Oder River strain and differences to closely related strains in morphology and ploidy. Our results provide unprecedented resolution of strain diversity in P. parvum and a better understanding of the genomic basis of toxin variability in haptophytes. The reference-quality genome will help to understand changes to microbial diversity in the face of increasing environmental pressures, and provides a basis for strain-level monitoring of invasive Prymnesium in the future.

genomics↗

bioRxiv2019

Incomplete resilience of a shallow lake to a brownification event

Dissolved organic carbon (DOC) concentrations in many freshwater ecosystems of the northern hemisphere have increased in recent decades due to additional terrestrial inputs. This phenomenon, known as brownification, can strongly alter the physical, chemical, and biological traits of aquatic ecosystems. Extreme rainfall can also cause sudden brownification, known as blackwater events in rivers, while longer term effects on lakes are unknown. Here, we investigated the resilience of a small, temperate, shallow lake to a strong natural flooding-induced brownification event in 2011-2012. From initial DOC and total phosphorus (TP) concentrations of ~12 and 0.04 mg L-1, respectively, the lake rapidly reached peak DOC and TP concentrations of 60 and 0.35 mg L-1, respectively. By the following year, water levels had returned close to initial values, yet two additional years of monitoring (until summer 2015) and a more recent sample in spring 2019 showed that the lake did not fully return to its pre-brownification state. Instead, DOC and TP concentrations plateaued at concentrations respectively 1.5-fold and twofold greater than pre-brownification values within less than two years and remained at these concentrations in spring 2019. During this initial recovery period the lake exhibited a decline of phytoplankton and a partial recovery of summer periphyton biomass and production, albeit a full return to pre-brownification values was not recorded in either case. DOC and TP concentrations were positively correlated to phytoplankton biomass and negatively to periphyton. As increases in phytoplankton production outpaced decreasing periphyton production, the net result of this brownification event has been an increase in whole-lake areal summertime primary production. This incomplete resilience to a flooding-induced brownification event implies consequences for several ecological and biogeochemical functions of shallow lakes that warrant further investigation and might contribute to the gradual increase of freshwater DOC concentrations in the northern hemisphere.

ecology↗

bioRxiv2019

Identifying multiple kinetic populations of DNA binding proteins in live cells using single-molecule fluorescence imaging

Understanding how multi-protein complexes function in cells requires detailed quantitative understanding of their association and dissociation kinetics. Analysis of the heterogeneity of binding lifetimes enables interrogation of the various intermediate states formed during the reaction. Single-molecule fluorescence imaging permits the measurement of reaction kinetics inside living organisms with minimal perturbation. However, poor photo-physical properties of fluorescent probes limit the dynamic range and accuracy of measurements of off rates in live cells. Time-lapse single-molecule fluorescence imaging can partially overcome the limits of photobleaching, however, limitations of this technique remain uncharacterized. Here, we present a structured analysis of which timescales are most accessible using the time-lapse imaging approach and explore uncertainties in determining kinetic sub-populations. We demonstrate the effect of shot noise on the precision of the measurements, as well as the resolution and dynamic range limits that are inherent to the method. Our work provides a convenient implementation to determine theoretical errors from measurements and to support interpretation of experimental data.\n\nSTATEMENT OF SIGNIFICANCEMeasuring lifetimes of interactions between DNA-binding proteins and their substrates is important for understanding how they function in cells. In principle, time-lapse imaging of fluorescently-tagged proteins using single-molecule methods can be used to identify multiple sub-populations of DNA-binding proteins and determine binding lifetimes lasting for several tens of minutes. Despite this potential, currently available guidelines for the selection of binding models are unreliable, and the practical implementation of this approach is limited. Here, using experimental and simulated data we identify the minimum size of the dataset required to resolve multiple populations reliably and measure binding lifetimes with desired accuracy. This work serves to provide a guide to data collection, and measurement of DNA-binding lifetimes from single-molecule time-lapse imaging data.

biophysics↗