A5091654640- DNA Repair Mechanisms
- Bacterial Genetics and Biotechnology
- Advanced Fluorescence Microscopy Techniques
- Advanced biosensing and bioanalysis techniques
- CRISPR and Genetic Engineering
- DNA and Nucleic Acid Chemistry
- Click Chemistry and Applications
- Photosynthetic Processes and Mechanisms
- Genomics and Chromatin Dynamics
- Antibiotic Resistance in Bacteria
- Photodynamic Therapy Research Studies
- PARP inhibition in cancer therapy
- Enzyme Structure and Function
- Advanced Electron Microscopy Techniques and Applications
- Escherichia coli research studies
- Photochromic and Fluorescence Chemistry
- Advanced Biosensing Techniques and Applications
- Plant Genetic and Mutation Studies
- Virus-based gene therapy research
- Nanoplatforms for cancer theranostics
- Bacterial biofilms and quorum sensing
- Molecular Biology Techniques and Applications
- Hepatitis B Virus Studies
- Lanthanide and Transition Metal Complexes
- Biotin and Related Studies
St Vincents Institute of Medical Research
2024
The Francis Crick Institute
2021-2024
Illawarra Health and Medical Research Institute
2019-2023
University of Wollongong
2018-2023
University of Groningen
2018-2021
Saarland University
2016-2021
Helmholtz Centre for Infection Research
2019
German Center for Infection Research
2019
Helmholtz Institute for Pharmaceutical Research Saarland
2019
Abstract The activation of eukaryotic origins replication occurs in temporally separated steps to ensure that chromosomes are copied only once per cell cycle. First, the MCM helicase is loaded onto duplex DNA as an inactive double hexamer. Activation after recruitment a set firing factors assemble two Cdc45–MCM–GINS (CMG) holo-helicases. CMG formation leads underwinding on path establishment fork, but whether becomes melted at this stage unknown 1 . Here we use cryo-electron microscopy image...
To prevent detrimental chromosome re-replication, DNA loading of a double hexamer the minichromosome maintenance (MCM) replicative helicase is temporally separated from unwinding. Upon S-phase transition in yeast, unwinding achieved two steps: limited opening helix and topological separation strands. First, Cdc45, GINS Polε engage MCM to assemble CMGE with partially hexamers that nucleate melting. In second step, triggered by Mcm10, CMGEs separate completely, eject lagging-strand template...
In Escherichia coli, damage to the chromosomal DNA induces SOS response, setting in motion a series of different repair and tolerance pathways. polymerase IV (pol IV) is one three specialised polymerases called into action during response help cells tolerate certain types damage. The canonical view field that pol primarily acts at replisomes have stalled on damaged template. However, results several studies indicate also other substrates, including single-stranded gaps left behind...
Biofilm formation is a key mechanism of antimicrobial resistance. We have recently reported two classes orally bioavailable C-glycosidic inhibitors the Pseudomonas aeruginosa lectin LecB with antibiofilm activity. They proved efficient in target binding, were metabolically stable, nontoxic, selective, and potent inhibiting bacterial biofilm. Here, we designed synthesized six new carboxamides 24 sulfonamides for detailed structure-activity relationship clinically representative variants....
In bacteria, genetic recombination is a major mechanism for DNA repair. The RecF, RecO and RecR proteins are proposed to initiate by loading the RecA recombinase onto DNA. However, biophysical mechanisms underlying this process remain poorly understood. Here, we used genetics single-molecule fluorescence microscopy investigate whether RecF function together, or separately, in live Escherichia coli cells. We identified conditions which functions genetically separable. Single-molecule imaging...
Abstract The bacterial RecF, RecO, and RecR proteins are an epistasis group involved in loading RecA protein into post-replication gaps. However, the targeting mechanism that brings these to appropriate gaps is unclear. Here, we propose may involve a direct interaction between RecF DnaN. In vivo, commonly found at replication fork. Over-expression of but not RecO or ATPase mutant, extremely toxic cells. We provide evidence molecular basis toxicity lies replisome destabilization....
While buffer cocktails remain the most commonly used method for photostabilization and photoswitching of fluorescent markers, intramolecular triplet-state quenchers emerge as an alternative strategy to impart fluorophores with 'self-healing' or even functional properties such photoswitching. In this contribution, we evaluated combinations both approaches show that inter- quenching processes compete each other. We find although rate is additive, photostability limited by faster pathway. Often...
Abstract When replication forks encounter template DNA lesions, the lesion is simply skipped in some cases. The resulting lesion-containing gap must be converted to duplex permit repair. Some filling occurs via switching, a process that generates recombination-like branched intermediates. Escherichia coli Uup and RadD proteins function different pathways UvrA-like ABC family ATPase. RecQ-like SF2 Loss of both functions uncovers frequent RecA-independent deletion events plasmid-based assay....
Abstract The combination of the two complementary imaging modalities 19 F magnetic resonance (MRI) and fluorescence (FLI) possesses high potential for biological medical applications. Herein we report first design, synthesis, dual detection validation, cytotoxic testing four promising BODIPY dyes MRI–fluorescence detection. Using straightforward Steglich reactions, small fluorinated alcohols were easily covalently tethered to a dye in yields, leaving its properties unaffected. synthesized...
Several functions have been proposed for the Escherichia coli DNA polymerase IV (pol IV). Although much research has focused on a potential role pol in assisting III replisomes bypass of lesions, is rarely found at replication fork vivo. Pol expressed increased levels E. cells exposed to exogenous damaging agents, including many commonly used antibiotics. Here we present live-cell single-molecule microscopy measurements indicating that double-strand breaks induced by antibiotics strongly...
Under many conditions the killing of bacterial cells by antibiotics is potentiated DNA damage induced reactive oxygen species (ROS) 1–3 . A primary cause ROS-induced cell death accumulation double-strand breaks (DSBs) 1,4–6 polymerase IV (pol IV), an error-prone produced at elevated levels in experiencing damage, has been implicated both ROS-dependent and DSBR 7–15 Here, we show using single-molecule fluorescence microscopy that DSBs promote pol activity two ways. First, exposure to...
pol VICE391 (RumAʹ2B) is a low-fidelity polymerase that promotes considerably higher levels of spontaneous "SOS-induced" mutagenesis than the related E. coli V (UmuDʹ2C). The molecular basis for enhanced was previously unknown. Using single molecule fluorescence microscopy to visualize enzymes, we discovered elevated are likely due, in part, prolonged binding RumB genomic DNA leading increased synthesis compared UmuC. We have generated steric gate variant (pol VICE391_Y13A) readily...
Genetically encodable fluorescent proteins have revolutionized biological imaging in vivo and vitro. Despite their importance, photophysical properties, i. e., brightness, count-rate photostability, are relatively poor compared to synthetic organic fluorophores or quantum dots. Intramolecular photostabilizers were recently rediscovered as an effective approach improve properties of fluorophores. Here, direct conjugation triplet-state quenchers redox-active substances creates high local...
Abstract Genetically encodable fluorescent proteins have revolutionized biological imaging in vivo and vitro . Since there are no other natural tags with comparable features, the impact of for research cannot be overemphasized. Despite their importance, photophysical properties, i.e., brightness, count-rate photostability, relatively poor compared to synthetic organic fluorophores or quantum dots. Intramolecular photostabilizers were recently rediscovered as an effective approach improve...
Abstract While buffer cocktails remain the gold-standard for photostabilization and photoswitching of fluorescent markers, intramolecular triplet-state quenchers emerge as an alternative strategy to impart fluorophores with ‘self-healing’ or even functional properties such photoswitching. In this contribution, we evaluated various combinations both approaches show that inter- quenching processes compete each other rather than being additive synergistic. Often dominate photophysical situation...
Abstract DNA polymerase IV (pol IV) is expressed at increased levels in Escherichia coli cells suffering high of damage. In a recent single-molecule imaging study, we demonstrated that elevating the pol concentration not sufficient to provide access binding sites on nucleoid, suggesting other factors may recruit its substrates once becomes damaged. Here extend this work, investigating proteins UmuD and RecA as potential modulators activity. promotes long-lived association with whereas...