Institutes
Refine
Year of publication
Document Type
- Doctoral Thesis (311)
- Article (282)
- Preprint (38)
- Contribution to a Periodical (25)
- Book (18)
- Report (2)
Language
- English (521)
- German (154)
- Multiple languages (1)
Has Fulltext
- yes (676)
Is part of the Bibliography
- no (676)
Keywords
- RNA (12)
- SARS-CoV-2 (10)
- NMR spectroscopy (9)
- inflammation (9)
- photochemistry (9)
- NMR (8)
- Biochemistry (7)
- Cell biology (7)
- E2 enzyme (6)
- TRACT (6)
- X-ray crystallography (6)
- oligomerization (6)
- ubiquitination (6)
- Alzheimer’s disease (5)
- COVID19-NMR (4)
- DNA (4)
- DNA-PAINT (4)
- ISGylation (4)
- Mass spectrometry (4)
- fluorescence (4)
- neurodegeneration (4)
- optogenetics (4)
- photolabile protecting groups (4)
- rhodopsin (4)
- structural biology (4)
- Antimicrobial resistance (3)
- Biophysical chemistry (3)
- C. elegans (3)
- Cancer (3)
- Non-structural protein (3)
- PBPK (3)
- RNA structures (3)
- Schmerz (3)
- Solution NMR spectroscopy (3)
- Solution NMR-spectroscopy (3)
- acetylcholine (3)
- autophagy (3)
- drug discovery (3)
- hippocampus (3)
- machine learning (3)
- mass spectrometry (3)
- membrane proteins (3)
- microdialysis (3)
- nuclear receptor (3)
- oligonucleotides (3)
- pain (3)
- polypharmacology (3)
- solid-state NMR (3)
- 5-lipoxygenase (2)
- 5′-UTR (2)
- 6-shogaol (2)
- ABC transporter (2)
- Arzneimittel (2)
- Bacterial biofilms (2)
- Bacterial structural biology (2)
- Baird's rule (2)
- Biotechnology (2)
- Chemical tools (2)
- Chromones (2)
- Covid19-NMR (2)
- Cryoelectron microscopy (2)
- DFT (2)
- EGFR (2)
- EPR (2)
- Endothelial cells (2)
- Enzyme mechanisms (2)
- G-Quadruplex (2)
- G-quadruplexes (2)
- GABARAP (2)
- GPCR (2)
- Host-biofilm interactions (2)
- ISGlyation (2)
- Inflammation (2)
- Innate immune response (2)
- LC3 (2)
- MLL (2)
- Mechanisms of disease (2)
- Medicinal chemistry (2)
- Membrane Proteins (2)
- Membrane proteins (2)
- Metabolism (2)
- NR4A2 (2)
- NanoBRET (2)
- PROTAC (2)
- Permeation and transport (2)
- Persistent wound infections (2)
- Pharmazeutische Technologie (2)
- Pharmazie (2)
- Photochemie (2)
- Photolabile Schutzgruppen (2)
- Precipitation inhibition (2)
- Protein drugability (2)
- RNA modification (2)
- RNA processing (2)
- Rietveld refinement (2)
- SL1 (2)
- SiGe alloys (2)
- Solution-state NMR (2)
- Structural biology (2)
- Supersaturation (2)
- Ubiquitin (2)
- Wirkstofffreisetzung (2)
- Zingiber officinale Roscoe (2)
- accessibility switch (2)
- actin (2)
- adenylyl cyclases (2)
- ankyrin (2)
- aptamers (2)
- bioequivalence (2)
- boron (2)
- boryl anions (2)
- boswellic acids (2)
- click chemistry (2)
- cluster compounds (2)
- computational chemistry (2)
- cyclic nucleotide-gated channels (2)
- deep learning (2)
- dorsal root ganglia (2)
- epigenetics (2)
- erythrocyte (2)
- excited state aromaticity (2)
- farnesoid X receptor (2)
- fluorescent probes (2)
- fluorine (2)
- fragment screening (2)
- germanium (2)
- glucose (2)
- guanylyl cyclases (2)
- inward proton pump (2)
- lactate (2)
- malaria (2)
- microbial rhodopsin (2)
- microscopy (2)
- microtubule-targeting agents (2)
- mitochondrial respiration (2)
- molecular dynamics (2)
- non-alcoholic steatohepatitis (2)
- phosphorylation (2)
- photoisomerization (2)
- photolabile Schutzgruppen (2)
- protein structure (2)
- reactive oxygen species (2)
- rearrangements (2)
- receptor tyrosine kinases (2)
- silicon (2)
- similarity measures (2)
- single-particle tracking (2)
- skin infection model (2)
- spinal cord (2)
- substituent effects (2)
- subvalent compounds (2)
- super-resolution microscopy (2)
- tRNA splicing (2)
- time-resolved spectroscopy (2)
- transcription factor (2)
- tumor microenvironment (2)
- ultrafast spectroscopy (2)
- vascular endothelial cells (2)
- virulence factor (2)
- 1,3-diamine (1)
- 10-Hydroxyaloins A/B (1)
- 140Ce (1)
- 19F (1)
- 19F NMR shifts for fluoroarenes (1)
- 2'-deoxyguanosine riboswitch (1)
- 2-Photonen (1)
- 2D NMR spectroscopy (1)
- 2′-O-methylation (1)
- 3D electron diffraction (1)
- 3D reconstruction and image processing (1)
- 4-fluoroamphetamine (1)
- 5'-UTR (1)
- 5-lipoxygenase; (1)
- 9-HODE (1)
- 9-borafluorene (1)
- ABC Transporter (1)
- ABC proteins, ribosome recycling (1)
- ABC transporters (1)
- ABCE1 (1)
- ADAR (1)
- AF4 (1)
- AF4–MLL (1)
- AMP-activated kinase (AMPK) (1)
- ATG4 (1)
- ATG8 (1)
- ATP binding (1)
- ATPases (1)
- ATTO 390 (1)
- Ab initio calculations (1)
- Absorption modeling (1)
- Acute leukemia, chromosomal translocation, MLL, KMT2A, AF4-MLL, reciprocal fusion protein, MLL-r leukemia, (1)
- Acute myeloid leukemia (AML) (1)
- Adeno-associated virus vector (1)
- Adenosine A1 receptor (1)
- Age (1)
- Airport emissions (1)
- Akute lymphoblastische Leukämie (1)
- Alkylation of RNA (1)
- Aloe (1)
- Aloe lateritia ENGLER (1)
- Alterung (1)
- Alzheimer's disease (1)
- Analytical chemistry (1)
- Angiogenesis (1)
- Anthranoids (1)
- Anthraquinones (1)
- Anthrones (1)
- Antibiotic resistance (1)
- Antigen carrier (1)
- Antigen processing and presentation (1)
- Antimuscarinic and Papaverine-Like Activity (1)
- Antisense agents (1)
- Apoptosis (1)
- Aptamer (1)
- Aptamere (1)
- Arc protein (1)
- Armadillo repeat protein (1)
- Arzneimittelanalogon (1)
- Asphodelaceae (1)
- Assignment (1)
- Atg8 (1)
- Aurora kinase (1)
- Autophagy (1)
- B-cell lymphoma (1)
- BACE1 (1)
- BET proteins (1)
- BH3 mimetics (1)
- BRG1 (1)
- Bacterial Signal Transduction (1)
- Bacterial biofilm (1)
- Binding Kinetics (1)
- Binding kinetic (1)
- Bindungsanalyse (1)
- BioID (1)
- Biochemische Analyse (1)
- Bioenabling formulations (1)
- Bioenergetics (1)
- Bioequivalence (1)
- Biological techniques (1)
- Biologie (1)
- Biomaterials – proteins (1)
- Biophysics (1)
- Biophysik (1)
- Biopysikalische Chemie (1)
- Biowaiver (1)
- Bispecific (1)
- Black Lipid Membrane (1)
- Brain (1)
- Broken symmetry (1)
- Bärnighausen tree (1)
- B−B bonds (1)
- B−H bonds (1)
- CAR-immunotherapy (1)
- CEST (1)
- CNN (1)
- CNP (1)
- COVID-19 (1)
- CPVT (1)
- CRBN (1)
- CRISPR/Cas9 (1)
- CXCL13 (1)
- CXCR5 (1)
- Caenorhabditis elegans (1)
- Calcium (1)
- Calstabin2/FKBP12.6 (1)
- Cancer models (1)
- Capsule Synthesis (1)
- Carbanions (1)
- Cardiovascular disease (1)
- Catalysis (1)
- Cell division (1)
- Cell-free Protein Expression (1)
- Cell-free Protein Synthesis (1)
- Cell-free protein synthesis (1)
- Cellular microbiology (1)
- Cellular neuroscience (1)
- Cellular suicide switch (1)
- Ceramide Synthase (1)
- ChIP-Seq (1)
- Chaperones (1)
- Chemical characterization (1)
- Chemical modification (1)
- Chemie (1)
- Chemistry (1)
- Chikungunya (1)
- Chloroxospezies (1)
- Chrimson (1)
- Clinical relevant dissolution specification (1)
- Colitis assoziate colorectal cancer (1)
- Colorectal cancer (1)
- Complex I (1)
- Complex II (1)
- Computational biology and bioinformatics (1)
- Controlled freezing/thawing (1)
- Covid19-nmr (1)
- Cryo-concentration (1)
- Cryo-electron Tomography (1)
- Crystal and Molecular Structure (1)
- Curcuminoid concentrations (1)
- Cy5.5 (1)
- Cyclization (1)
- CysLTR1 (1)
- Cysteine‐Rich Domain (CRD) (1)
- DDR (1)
- DEAD-Box Helicase (1)
- DFT+MBD calculations (1)
- DNA G-quadruplex (1)
- DNA conjugate (1)
- DNA damage response (1)
- DNA methylation (1)
- DNA pulldown (1)
- DNA recognition (1)
- DNA-protein-interactions (1)
- DNP (1)
- DNP in solids (1)
- Daclatasvir (1)
- Degrader design (1)
- Designed ankyrin repeat protein (1)
- Desinfektionsmittel (1)
- DiI (1)
- Diffuse large B-cell lymphoma (1)
- Dipol (1)
- Directional freezing (1)
- Diseases (1)
- Dissolution (1)
- Dissolution nanoparticles (1)
- Disulfide bond (1)
- Disulphide Bridges (1)
- Drug Delivery (1)
- Drug delivery systems (1)
- Drug discovery (1)
- Drug screening (1)
- Drug substance bottles (1)
- Dynamics (1)
- E3 Ligase (1)
- EPR spectroscopy (1)
- ERK3 (1)
- ERQC (1)
- Efflux system (1)
- Electron Paramagnetic Resonance (1)
- Electron microscopy (1)
- Electrospinning (1)
- Elektrochemie (1)
- Enantioselektive Katalyse (1)
- Endothelial cell (1)
- Energie (1)
- Enthalpy (1)
- Entwicklungsbiologie (1)
- FAIRE-Seq (1)
- FBDD (1)
- FBS (1)
- FPR (1)
- Faltungsdynamik (1)
- Fatty acid synthases (1)
- Ferroptosis (1)
- Festkörperstruktur (1)
- Flash photolysis (1)
- Flavin homeostasis (1)
- Fluorene derivatives (1)
- Food effect (1)
- Fortpflanzung (1)
- Fragment (1)
- Freezing and thawing (1)
- Functional aspects of cell biology (1)
- Fusion proteins (1)
- G protein-coupled receptors (1)
- G-protein-coupled receptors (1)
- G-quadruplex binders (1)
- G2A receptor (1)
- GC-MS (1)
- GIIb/IIIa-receptor (1)
- Gastrointestinaltrakt (1)
- GdIr2Si2 (1)
- Gene therapy (1)
- Genetic engineering (1)
- Genetics (1)
- Genexpression (1)
- Genome engineering (1)
- Genregulation (1)
- Geschichte der Pharmazie (1)
- Glucose (1)
- Glutathione (1)
- Guanylatzyklasen (1)
- Gβγ subunit (1)
- H/ACA-RNP (1)
- HCV (1)
- HIV protease inhibitors (1)
- Haloferax volcanii (1)
- HeLa cells (1)
- Helicase (1)
- Hepatitis B virus (1)
- Herpes Simplex Virus Type 1 (HSV-1) (1)
- Heterocycles (1)
- High Performance Liquid (1)
- Histidine Kinases (1)
- Homeoviscous adaptation (1)
- Homonataloins A and B (1)
- Hydrodynamik (1)
- IAP (1)
- INS (1)
- ITGA9 (1)
- Image processing (1)
- Immune evasion (1)
- Immunochemistry (1)
- Immunology (1)
- Immunseren (1)
- In situ hybridization (1)
- In vitro wound models (1)
- Infectious diseases (1)
- Inflammatory Bowel Disease (1)
- Influenza (1)
- Innovation (1)
- Instrumentation & Devices (1)
- Ion transport (1)
- Ischämie (1)
- Isolation, separation and purification (1)
- Isothermal Titration Calorimetry (1)
- Jet engine oil (1)
- Juckreiz (1)
- Jurkat (1)
- K48-linked (1)
- K63-linked (1)
- KCNT1 (1)
- KMT2A (1)
- KR2 (1)
- Kaliumkanal (1)
- Kaliumkanal-Aktivatoren (1)
- Kd determination (1)
- Ketonkörper (1)
- Kinase (1)
- Kinase Inhibitors (1)
- Kinase inhibitors (1)
- Kinetik (1)
- Knochenmarksmikromilieu (1)
- Knotless phytochrome (1)
- LASS (1)
- LC-MS-based lipid mediator analysis (1)
- LC3/GABARAP (1)
- LILBID-MS (1)
- LIMK1 (1)
- LIMK2 (1)
- LINC00607 (1)
- LINE-1 (1)
- LIR (1)
- LIR motif (1)
- LIR motifs (1)
- Lehramt (1)
- Leukaemia (1)
- Leukemia (1)
- Leukocytes (1)
- Lewis acid (1)
- Ligands (1)
- Lipid Environment (1)
- Lipid packing (1)
- Lipidinteraktion (1)
- Lipoprotein Structure (1)
- Liposomes (1)
- M1 Ubiquitination (1)
- MACS (1)
- MAPK6 (1)
- MET (1)
- MET receptor (1)
- MFS-type tripartite system (1)
- MHC I (1)
- MLH1 (1)
- MLL-r acute leukemia (1)
- MODY (1)
- Macrodomain (1)
- Makrophagen (1)
- Massenspektrometrie (1)
- Mechanistic (1)
- Medication Appropriatness Index (1)
- Membrane Protein (1)
- Membrane fluidity (1)
- Membrane homeostasis (1)
- Membranes (1)
- Mesoporous silica (1)
- Meta effect (1)
- Metabolic Regulation (1)
- Metabolismus (1)
- Mice (1)
- Microbial Rhodopsins (1)
- Microbiology (1)
- Microdialysis (1)
- Mitochondria (1)
- Mitochondrial dysfunction (1)
- Mitosis (1)
- Modification (1)
- Molecular Interactions (1)
- Molecular Neuroscience (1)
- Molecular biology (1)
- Molecular glue (1)
- Monolage (1)
- Multi-domain proteins (1)
- Multidrug Transporters (1)
- Multimedikation / Polypharmazie (1)
- Multimorbidität (1)
- Multiphoton microscopy (1)
- Mycobacterium tuberculosis (1)
- N-glycoproteome (1)
- NADPH oxidase (1)
- NASH (1)
- NMR Solution Structure (1)
- NMR shift calculation (1)
- NMR solution structure (1)
- NMRI (1)
- NOESY (1)
- NPR2 (1)
- NR4A3 (1)
- NS5A (1)
- Nanocarriers (1)
- Nanodiscs (1)
- Nanotechnologie (1)
- Natural product (1)
- Natural products (1)
- Neuropathic pain (1)
- New E3-ligase ligands (1)
- Nox (1)
- Nox inhibition (1)
- Nucleic acid-binding domain (1)
- Nucleolar protein 14 (1)
- Nukleinsäuren (1)
- Nukleäre Rezeptoren (1)
- OTULIN (1)
- Oligonukleotid (1)
- Oncogenes (1)
- Oncoprotein activation (1)
- Ophthalmologie (1)
- Optogenetics (1)
- Organische Synthese (1)
- Organokatalyse (1)
- Orphan nuclear receptor (1)
- Outer Membrane Protein RcsF (1)
- Overhauser effect (1)
- Oxanthrone Glucosyls (1)
- Oxidative phosphorylation (1)
- PAC-1 (1)
- PBPK modelling (1)
- PBPK/PD (1)
- PC12APPsw (1)
- PDF-Global-Fit (1)
- PELDOR (1)
- PELDOR / DEER (1)
- PELDOR/DEER spectroscopy (1)
- PIM kinases (1)
- PLGA nanoparticles (1)
- PRIMUM-Studie (1)
- PXRD (1)
- Pain behavior (1)
- Paramyxoviruses (1)
- Parkinson’s disease (1)
- Partial agonist (1)
- Patch Clamp (1)
- Patch-clamp (1)
- Peptide nucleic acids (1)
- Pharmacology (1)
- Pharmazeutische Chemie (1)
- Phenalen, Heteroatom-Dotiert (1)
- Phosphoproteome (1)
- Photocages (1)
- Photochemistry (1)
- Photoisomerization (1)
- Photolabile protecting groups (1)
- Photoschalter (1)
- Photosynthesis (1)
- Physical chemistry (1)
- Physiologically Based Pharmacokinetic Modelling (1)
- Phytochrome (1)
- Pigment Green 7 (1)
- Pneumoviruses (1)
- Polyketide synthases (1)
- Polyketides (1)
- Polypeptid transport-associated domains (1)
- Post-Hartree-Fock methods (1)
- Potassium channels (1)
- Pre‐Ligand Assembly Domain (PLAD) (1)
- Process characterization (1)
- Process time (1)
- Proteasome inhibitor (1)
- Protein Kinase (1)
- Protein biosynthesis (1)
- Protein engineering (1)
- Protein folding (1)
- Protein synthesis (1)
- Proteins (1)
- Proteomics (1)
- Proteostasis (1)
- Pruritus (1)
- Pseudocontact shift (1)
- Pseudouridine (1)
- Pseudouridylation (1)
- Pulverdiffraktometrie (1)
- Puromycin (1)
- QbD (1)
- Quality control (1)
- Quantenpunkte (1)
- Quinonoids (1)
- R values (1)
- RMN (1)
- RNA PT (1)
- RNA cleavage (1)
- RNA degradation (1)
- RNA dynamics (1)
- RNA modifications (1)
- RNA synthesis (1)
- RNA therapy (1)
- RNA, biochemistry, mass spectrometry (1)
- RNA-Seq (1)
- RNA-binding proteins (1)
- RND (1)
- RXR (1)
- RXR-Selektivität (1)
- RXR-Subtypen (1)
- Rab27a (1)
- Radiotherapy (1)
- Regulation der Genexpression (1)
- Regulation of lipid metabolism (1)
- Release testing (1)
- Repair (1)
- Repression <Genetik> (1)
- Retinoid X Rezeptoren Subtypen (1)
- Retrotransposition (1)
- Rh(II) catalysis (1)
- Rhabdomyosarcoma (1)
- Rhamnus purshianus D. C. (1)
- RhlB (1)
- Rhodopsine (1)
- Ribonucleoproteins (1)
- Ribosome Recycling (1)
- Ribosome biogenesis (1)
- RyR2 (1)
- Ryanodin (1)
- Röntgenstrukturanalyse (1)
- S107 (1)
- SAR (1)
- SEDDS (1)
- SERCA (1)
- SHAPE analysis (1)
- SL5a (1)
- SL5b (1)
- SL5b + c (1)
- SL5c (1)
- SMEDDS (1)
- SNEDDS (1)
- SNP (1)
- SOFI (1)
- SPM (1)
- STED microscopy (1)
- STM (1)
- Saccharomyces cerevisiae (1)
- Schleimpilze (1)
- Screening (1)
- Selectivity (1)
- Semi-empiricalcalculations (1)
- Sexualität (1)
- Sila-Substitution (1)
- Silacyclobutenes (1)
- Silafullerane (1)
- Silane (1)
- Silole (1)
- Silylation (1)
- Single-Molecule Localization Microscopy (SMLM) (1)
- Slack (1)
- Smac mimetic (1)
- Small molecules (1)
- Sodium ion pump (1)
- Solid Oral Dosage Forms (1)
- Solid dispersion (1)
- Solubility (1)
- Spektroskopie (1)
- Sphingolipids (1)
- Sphingosinkinase 2 (1)
- Spinach (1)
- Spinlabel (1)
- Stem-cell research (1)
- Stereochemistry (1)
- Stress response (1)
- Stroke (1)
- Structural Biology (1)
- Structure refinement (1)
- Subtomogram averaging (1)
- T-cell (1)
- TBK1 (1)
- TERRA RNA (1)
- TICT (1)
- TSEN (1)
- Target validation (1)
- Taspase1 (1)
- Temperature mapping (1)
- Tetrelbindung (1)
- TgF344-AD (1)
- Theoretical chemistry (1)
- Thyroidhormone (1)
- Tiermodell (1)
- Time-resolved spectroscopy (1)
- Tissue distribution (1)
- TmRh2Si2 (1)
- Transcription factors (1)
- Transcriptional regulation (1)
- Transient absorption (1)
- Transport Drugs (1)
- Transporters (1)
- Transposon systems (1)
- Tumour-suppressor proteins (1)
- Two-component System (1)
- UBA5 (1)
- UBL (1)
- UFIM (1)
- UFM1 (1)
- ULK4 (1)
- UV/Vis spectroscopy (1)
- Ubiquitin chains (1)
- Ubiquitinome (1)
- Ultrafine particles (UFPs) (1)
- Ultrakurzzeit-Spektroskopie (1)
- Unc-51-like kinase (1)
- VHL (1)
- Viologene (1)
- Vioprolide A (1)
- Viral entry (1)
- Virologie (1)
- Virus-host Interaction (1)
- Wasserlöslichkeit (1)
- Wound dressings (1)
- Wound infection (1)
- X-ray structure analysis (1)
- Xenocoumacin (1)
- ZIKV (1)
- Zolpidem (1)
- Zolpidem; Food-effect, PBBM, PBPK (1)
- Zweiphotonen-Anregung (1)
- Zyklisierung (1)
- absolute quantification of RNA modifications (1)
- ab initio calculations (1)
- accessory proteins (1)
- actin cytoskeleton dynamics (1)
- acute leukemia (1)
- acyclovir (1)
- adaptor mediated recruitment (1)
- additive manufacturing (1)
- adenosine–arginine conjugate (1)
- aging (1)
- alternative matrices (1)
- ambiguous structures (1)
- amorphous solid dispersions (1)
- amyloid beta (1)
- amyloid precursor protein (1)
- amyloidogenesis (1)
- analgesia (1)
- analytical chemistry (1)
- angiogenesis-related cell functions (1)
- anilinopyrimidine (1)
- antibiotic resistance (1)
- antigen presentation (1)
- antigen processing (1)
- antisense technology (1)
- antiviral response (1)
- antiviral signaling (1)
- apoptosis (1)
- aptamer (1)
- arachidonic acid (AA) (1)
- arrhythmia model (1)
- astrogliosis (1)
- atropisomers (1)
- attention module (1)
- atypical MAPK kinase (1)
- automated assignment (1)
- axon bifurcation (1)
- axon branching (1)
- azo compounds (1)
- baicalein (1)
- barstar (1)
- benchmark standards (1)
- benzazepinone (1)
- benzothiazoles (1)
- big data (1)
- bilin-binding photoreceptors (1)
- bio-enabling formulations (1)
- bioavailability (1)
- bioconjugation (1)
- biomarker (1)
- biophysics (1)
- bioprinting (1)
- biorelevant dissolution (1)
- bio‐enabling formulations (1)
- bisubstrate inhibitors (1)
- body-on-a-chip (1)
- bond activation (1)
- boron heterocycles (1)
- breast milk (1)
- brightness (1)
- buchu (1)
- c-Myc (1)
- cGKI (1)
- cGMP signalling (1)
- cMYC (1)
- caged compounds (1)
- calsequestrin (1)
- cancer (1)
- cancer associated fibroblasts (1)
- canine cancer (1)
- capture (1)
- carbazole ligands (1)
- carbon direct detection (1)
- catalytic mechanism (1)
- cell cycle (1)
- cell migration (1)
- cell stress (1)
- cell-free expression (1)
- cell-free protein synthesis (1)
- cellular uptake and localization (1)
- ceramide synthase (1)
- cerium (1)
- cerumen (1)
- channelrhodopsin (1)
- charge transfer (1)
- chemical shift assignment (1)
- chemiluminescence (1)
- chemistry education (1)
- chemoenzymatic synthesis (1)
- chemogenomics (1)
- cholesterol (1)
- chronobiology (1)
- circular dichroism (1)
- classical Hodgkin lymphoma (1)
- clustering (1)
- co-crystal structure (1)
- coarse-grained simulations (1)
- cofilin (1)
- colon (1)
- combinatorial approaches (1)
- commercialization (1)
- comparative oncology (1)
- complex I (1)
- complex IV (1)
- computer-aided diagnosis (1)
- conductivity (1)
- confocal Raman microscopy (1)
- confocal microscopy (1)
- connection to nature (1)
- continuous rotation (1)
- copper phthalocyanine (1)
- correlated electrons (1)
- coumarin (1)
- coumarin 6 (1)
- covalent inhibitors (1)
- creams (1)
- cross-correlation functions (1)
- cross-section (1)
- cryo-electron microscopy (1)
- crystal structure (1)
- curcumin (1)
- cyanines (1)
- cycloaddition (1)
- cysteine reactivity (1)
- cysteine-redox regulation (1)
- cytokine (1)
- data curation (1)
- data projection (1)
- data science (1)
- de novo design (1)
- dendritic spines (1)
- denoising (1)
- de novo design (1)
- diacylglycerol kinase (1)
- dialysis (1)
- diazo compounds (1)
- diboranes (1)
- differential scanning fluorimetry (1)
- diffusion states (1)
- digestion artifact (1)
- dissolution (1)
- dopamine D2S/D3 receptor (1)
- drug abstinence (1)
- drug development (1)
- drug metabolism and distribution (1)
- drug release (1)
- drug resistance (1)
- drug targets (1)
- drug therapy (1)
- drug-likeness (1)
- dynamic nuclear polarization (1)
- dynamics (1)
- efflux pump (1)
- eicosanoid (1)
- electron beam induced deposition (1)
- electron crystallography (1)
- electron transfer system (1)
- electronic nose (1)
- elegans (1)
- enamides (1)
- end maturation (1)
- endolysosomal system (1)
- endonuclease (1)
- endoplasmic reticulum (1)
- endothelial cells (1)
- energy barriers (1)
- enolase-1 (1)
- environmental education (1)
- enzyme catalysis (1)
- enzyme kinetics (1)
- epitranscriptome (1)
- epitranscriptomics (1)
- ethnobotany (1)
- etravirine (1)
- evolution (1)
- exosomes (1)
- extracellular matrix (1)
- extract (1)
- famotidine (1)
- fatty acid (1)
- fatty acid/biosynthesis (1)
- filopodia (1)
- flavolignan (1)
- fluidity (1)
- fluorescent labeling (1)
- fluorophore (1)
- flux growth (1)
- folding landscapes (1)
- food effect (1)
- fragment-based design (1)
- fragment-based drug design (1)
- fragment-based screening (1)
- frankincense (1)
- free mRNA (1)
- fulgide (1)
- gallein (1)
- gapmers (1)
- gene expression (1)
- genetic code expansion (1)
- glucose transport (1)
- glutathione (1)
- glutathionylation (1)
- glycerophospholipid (1)
- glycolysis (1)
- green gap (1)
- guanidine riboswitch (1)
- guanidine-II riboswitch (1)
- hTERT (1)
- hairpin polyamides (1)
- heart-on-a-chip (1)
- hepatische Ketogenese (1)
- hepatocyte nuclear factor 4α (1)
- heteronuclear detection (1)
- hierarchical clustering (1)
- high content imaging (1)
- high dimensional complex data (1)
- high-throughput sequencing (1)
- histamine (1)
- histone deacetylase (1)
- histone modifications (1)
- homogeneous catalysis (1)
- human milk (1)
- human neurons (1)
- human pain models (1)
- human pharmacology (1)
- hxt0 strain (1)
- hydromethylthionine (1)
- hypoxia (1)
- iPSc (1)
- immune system (1)
- in silico tools (1)
- in vitro activities (1)
- in vivo neuroimaging (1)
- in-vitro–in-vivo correlation (1)
- infant health (1)
- information literacy (1)
- interaction partners (1)
- interest in animals (1)
- interface engineering (1)
- intracellular internalization (1)
- intrinsically disordered region (1)
- inflammatory pain (1)
- ion beam induced deposition (1)
- ionization (1)
- isopropanol (1)
- isothermal titration calorimetry (ITC) (1)
- isotope labeling (1)
- itch (1)
- kidney-on-a-chip (1)
- kinase (1)
- kinase inhibitors (1)
- kinetics (1)
- labelfree quantitative mass spectrometry (1)
- lattice-energy minimization (1)
- leucocytes (1)
- leukocyte adhesion (1)
- leukotriene (1)
- leukotriene A4 hydrolase (1)
- lichtaktivierbare Werkzeuge (1)
- ligand design (1)
- ligases (1)
- light control (1)
- light-induced polymerization (1)
- lipid (1)
- lipid based formulation (1)
- lipid metabolism (1)
- lipid suspension (1)
- lipidic cubic phase (1)
- lipidomics (1)
- liposomes (1)
- lipoxin (1)
- lipoxygenase (1)
- lipoxygenase inhibitor (1)
- liquid chromatography–mass spectrometry (1)
- liver fibrosis (1)
- liver-on-a-chip (1)
- long non-coding RNA (1)
- longitudinale Medikationsveränderung (1)
- lung cancer (1)
- lung tumor heterogeneity (1)
- lung-on-a-chip (1)
- lysosome (1)
- lysosomes (1)
- mRNA surveillance (1)
- macromolecular machines (1)
- macrophage (1)
- magnetic exchange coupling constants (1)
- magnetic resonance imaging (1)
- magnetism (1)
- main group elements (1)
- maternal diet (1)
- mature milk (1)
- meat adulteration (1)
- medicinal chemistry (1)
- medicinal plant (1)
- memantine (1)
- membrane anchor; substrate-binding protein dependent secondary transport; TRAP-associated extracytoplasmic immunogenic (TAXI); tripartite ATP-independent periplasmic (TRAP) (1)
- membrane dynamics (1)
- membrane fluidity (1)
- membrane mimics (1)
- membrane organization (1)
- membrane protein complexes (1)
- membrane receptors (1)
- membranes/fluidity (1)
- memory consolidation (1)
- metabolic sydrome (1)
- metabolomics (1)
- metallo β lactamases (1)
- metastasis (1)
- mice (1)
- micro-physiological systems (1)
- microRNA (1)
- microsomal prostaglandin E2 (1)
- mitochondria (1)
- mixed valence compounds (1)
- modeling & simulation (1)
- modeling and simulation (1)
- molecular docking (1)
- molecular electronics (1)
- molecular machines (1)
- molecular switch (1)
- molekulare Werkzeuge (1)
- multi-objective optimization (1)
- multi-target drug (1)
- multi-task learning (1)
- multicolor imaging (1)
- multidrug resistance protein 4 (1)
- multiple sclerosis (1)
- multiresistant bacteria (1)
- multispectral flow cytometry (1)
- multitarget drugs (1)
- multitasking protein (1)
- n_TOF (1)
- nanodiscs (1)
- nanomedicine (1)
- natural compound synthesis (1)
- nerve injury (1)
- neural network (1)
- neurodegenerative disorders (1)
- neuromuscular system (1)
- neuropathic pain (1)
- neuropathy (1)
- neurotropic virus (1)
- neutron (1)
- nitrogen direct detection (1)
- nitroindoles (1)
- nitrosylation (1)
- nociception (1)
- non-alcoholic fatty liver disease (1)
- non-canonical amino acid (1)
- non-coding RNAs (1)
- non-invasive imaging (1)
- non-natural nucleotide (1)
- nonstructural proteins (1)
- nuclear magnetic resonance (NMR) (1)
- nuclear magnetic resonance spectroscopy (1)
- nuclear receptor related-1 (1)
- nuclear receptor-related 1 (1)
- nucleobases (1)
- nucleophilic boron (1)
- nucleophilic substitution (1)
- nucleoside analysis (1)
- nucleosynthesis (1)
- oligonucleotide (1)
- oncogene promoters (1)
- one-dimensional convolutional neural network (1)
- one-pot reaction (1)
- optogenetic (1)
- organs-on-a-chip (1)
- osteopontin (1)
- overtaking innovation (1)
- oxidative folding (1)
- p38 (1)
- pSer domain (1)
- pair distribution function analysis (1)
- pair distribution function refinement (1)
- pair distribution functions (1)
- pancreatic cancer (1)
- peptide antibiotics (1)
- peptide loading complex (1)
- peptide transport (1)
- performance tests (1)
- peripheral injury (1)
- peroxisome prolifer ators-activated receptor-γ (1)
- phage lysis proteins (1)
- pharmacokinetics (1)
- pharmacokinetics/ pharmacodynamics (PK/PD), (1)
- pharmacological activity (1)
- pharmacophore model (1)
- phase separation (1)
- phenotypic screening (1)
- phospholipids/phosphatidylglycerol (1)
- photo-tethering (1)
- photoacid (1)
- photobleaching (1)
- photocage (1)
- photocages (1)
- photocaging (1)
- photocycle (1)
- photoschalbare (1)
- photospaltbare Linker (1)
- photoswitch (1)
- photoswitches (1)
- phototherapeutic window (1)
- phytochemical composition (1)
- phytochromes (1)
- platelets (1)
- polymers (1)
- polyubiquitin (1)
- potassium (1)
- potassium transporter (1)
- powder data (1)
- precipitation inhibitor (1)
- precursor synthesis (1)
- privileged structures (1)
- problem-based learning (1)
- proline isomerization (1)
- prostaglandin (1)
- prostaglandin E (1)
- prostaglandins (1)
- protein X-ray crystallography (1)
- protein capture (1)
- protein compartmentalization (1)
- protein engineering (1)
- protein folding (1)
- protein kinase inhibitor (1)
- protein motif (1)
- protein oxidation (1)
- protein tyrosine phosphatase (1)
- protein-protein interactions (1)
- proteins (1)
- proteostasis stress (1)
- proton transfer (1)
- pseudokinase (1)
- pseudouridylation (1)
- purification of protein complexes (1)
- quantenpunkte (1)
- quantum yield (1)
- quercetin (1)
- quinacridone (1)
- radical reactions (1)
- random forest regressor (1)
- real-time NMR spectroscopy (1)
- receptor condensates (1)
- receptor cross-interaction (1)
- receptor dynamics (1)
- red light (1)
- redox chemistry (1)
- regulatory science (1)
- release testing (1)
- resmetirom (1)
- resolution of inflammation (1)
- resolvin (1)
- respiratory chain (1)
- resveratrol (1)
- retinal (1)
- retinoid X receptor (1)
- rhabdomyosarcoma (1)
- rhodamine 123 (1)
- ribosomal exit tunnel (1)
- ribosome-associated quality control (1)
- riboswitch (1)
- ribozymes (1)
- rivastigmine (1)
- s-process (1)
- screening system (1)
- selbst-anordnende (1)
- self-assembled monolayers (1)
- self-initiated photografting and photopolymerization (1)
- semisolids (1)
- siRNAs (1)
- signal transduction (1)
- silibinin (1)
- silybin (1)
- single particle cryo-EM (1)
- single-molecule FRET (1)
- single-molecule imaging (1)
- single-molecule localization microscopy (1)
- single-particle cryoEM (1)
- single-trajectory analysis (1)
- site-directed spin labeling (1)
- site-specific labeling (1)
- skin delivery (1)
- skin-on-a-chip (1)
- smFRET (1)
- small protein (1)
- small-molecule inhibitor (1)
- smooth muscle cells (1)
- sodium alkoxide (1)
- solid-phase synthesis (1)
- solid-state NMR spectroscopy (1)
- soluble epoxide hydrolase (1)
- solution-state NMR (1)
- solvate (1)
- somatosensory system (1)
- soraphen (1)
- spiro compounds (1)
- splice-modulating oligonucleotides (1)
- stability of fluorescent label (1)
- stearic acid (1)
- stereoselective synthesis (1)
- streptozotocin (1)
- stress dependent RNA modification dynamics (1)
- structural proteins (1)
- structure determination (1)
- structure determination from powder data (1)
- structure-activity relationships (1)
- sugar transport assays (1)
- sugar transport inhibitors (1)
- sulfamide (1)
- super-SMEDDS (1)
- super-SNEDDS (1)
- supercomplex (1)
- supersaturating drug delivery systems (1)
- supersaturation (1)
- surface modification (1)
- surface-initiated polymerization (1)
- surfactant (1)
- swelling (1)
- synaptic transmission (1)
- synaptic vesicle recycling (1)
- synthase-1 (1)
- t(4;11) leukemia (1)
- tRNA (1)
- target variability (1)
- temperature jump (1)
- tetrapyrrole-binding photoreceptors (1)
- therapeutic equivalence (1)
- therapy (1)
- thermal stability (1)
- thiol inhibitors (1)
- thyroid hormone receptor (1)
- tissue distribution (1)
- tissue-plasma-ratio (1)
- toll-like receptor (1)
- topical formulations (1)
- total scattering technique (1)
- total scattering techniques (1)
- trafficking (1)
- transcription (1)
- transient complex (1)
- tumor (1)
- type 2 diabetes (1)
- ubiquitin-conjugating enzymes (1)
- umpolung (1)
- validation (1)
- vascular biology (1)
- vascular biology/endothelial cells (1)
- venetoclax (1)
- viral immune escape (1)
- virtual labeling (1)
- virus (1)
- vitamin B12 (1)
- voltage-gated proton channels (1)
- voltage-sensing domains (1)
- voltage-sensing phosphatases (1)
- weakly supervised learning (1)
- x-ray crystallography (1)
- yeast fatty acid synthase (1)
- zinc finger (1)
- zolpidem (1)
- zoo education (1)
- µ-protein (1)
Institute
- Biochemie, Chemie und Pharmazie (676)
- Präsidium (43)
- Medizin (35)
- Buchmann Institut für Molekulare Lebenswissenschaften (BMLS) (31)
- Zentrum für Biomolekulare Magnetische Resonanz (BMRZ) (30)
- Biowissenschaften (23)
- MPI für Biophysik (15)
- Physik (12)
- Zentrum für Arzneimittelforschung, Entwicklung und Sicherheit (ZAFES) (5)
- Geowissenschaften / Geographie (4)
In vitro release testing as an alternative to establishing bioequivalence of drug products in vivo
(2020)
Generische Arzneimittel werden als Arzneimittel definiert, die im Vergleich zu einem Referenzarzneimittel hinsichtlich der meisten pharmazeutischen Aspekte identisch sind.
Um die therapeutische Äquivalenz zum Referenzprodukt sicherzustellen, sind Bioverfügbarkeitsstudien erforderlich. Für Arzneimittel, die als feste, perorale, schnell freisetzende Darreichungsformen formuliert sind, kann auf den Nachweis der Bioäquivalenz in vivo zugunsten eines vergleichenden Freisetzungstests in vitro im Rahmen eines sogenannten Biopharmaceutics Classification System (BCS) basierten Biowaivers verzichtet werden.
Der BCS-basierte Biowaiver ist ein vielversprechendes Instrument, welches Kosteneinsparungen sowie eine Verringerung des regulatorischen Aufwands im Zuge der behördlichen Zulassung von Generika ermöglicht und dazu beitragen kann, die Zugänglichkeit unentbehrlicher Arzneimittel zu verbessern. Dabei gibt es jedoch auch Hürden, welche die weitläufige Anwendung des Verfahrens verhindern: Unklare Löslichkeits- und Permeabilitätsklassifizierungen von Wirkstoffen, Arzneimittel, welche die in vitro Freisetzungskriterien nicht erfüllen, sowie Zweifel an der Eignung der regulatorischen Spezifikationen, Freisetzungsunterschiede in vitro erfassen zu können, die für das Verhalten in vivo relevant sind.
In der vorliegenden Dissertation werden diese Probleme thematisiert, indem eine umfassende Bewertung der Anwendbarkeit und Einschränkungen des BCS-basierten Biowaivers in seinem aktuell regulatorisch vorgeschriebenen Ablauf vorgenommen wird. Mögliche Anpassungen des Verfahrens wurden auf der Grundlage experimenteller in vitro Daten untersucht, bewertet und mithilfe von in silico Simulationsmodellen auf die Situation in vivo extrapoliert.
Throughout their life cells of eukaryotic organisms can be confronted with a variety of proteotoxic stresses and in order to survive, corresponding resistance mechanisms had to evolve. Proteotoxic stresses can cause misfolding of proteins and accumulation of toxic protein aggregates. Failure to remove aggregates of misfolded proteins compromises cellular function and can ultimately cause cell death and disease. To deal with this challenge, cells utilize a complex network of protein quality control pathways, including chaperones, the ubiquitin-proteasome system and the autophagy system.
Another mechanism to cope with proteotoxic stresses is the stalling of translation initiation in order to save valuable resources and prevent faulty translation. Upon stress, intrinsically disordered RNA-binding proteins such as TIA-1 or G3BP1/2 are recruited to stalled preinitiation complexes and a network of multivalent interactions between RNAs and proteins is formed. These mRNP networks can merge with each other and phase separate into membraneless liquid-like structures called stress granules (SGs). Once stress is released, SGs are quickly resolved and translation continues. Yet, chronic stress or mutations of SG-associated proteins can cause persistent SGs, which can sequester misfolded proteins and have been linked to neurodegenerative diseases such as amyotrophic lateral sclerosis or frontotemporal dementia.
In mammalian cells, three isoforms of the small ubiquitin-related modifier (SUMO), SUMO1, SUMO2 and SUMO3 are covalently attached to lysine residues of target proteins. SUMO conjugation is catalyzed via an enzymatic cascade of an heteromeric E1 activating enzyme, the E2 conjugating enzyme Ubc9 and in some cases one of a limited number of E3 SUMO ligases. SUMOylation is a dynamic modification and can be reversed by SUMO isopeptidases, the best characterized of which belong to the SENP family. Cellular stresses such as heat or oxidative stress strongly induce SUMOylation resulting in increased numbers of poly-SUMOylation (formation of SUMO2/3 chains) on nuclear proteins.
The SUMO-targeted ubiquitin ligase (STUbL) RNF4 harbors four SUMO interaction motifs in its N-terminal domain. This feature allows RNF4 to specifically bind poly-SUMOylated proteins and catalyze their proteolytic or non-proteolytic ubiquitylation.
A variety of substrate proteins have been shown to undergo SUMO-primed ubiquitylation by RNF4 in response to stress or DNA damage. RNF4-mediated ubiquitylation is often a signal for proteolytic degradation of these substrates.
In this work we aimed by identify novel RNF4 targets, in heat-stressed cells in order to gain a wider understanding of the nuclear proteotoxic stress response. Analysis by mass spectrometry revealed that a large fraction of RNF4-interacting proteins in heatstressed cells are nuclear RNA-binding proteins, many of which shuttle outside the nucleus and associate with SGs upon stress. We validated, that nuclear RNA-binding proteins, such as TDP-43 and hnRNP M are indeed heat-induced targets of SUMOprimed ubiquitylation by RNF4.
These initial results led us to further investigate the links between the SUMO/RNF4-mediated, nuclear protein quality control and the dynamics of cytosolic heat- or arsenite-induced SGs. SUMO2/3 and RNF4 are mainly nuclear proteins and we confirmed that they do not associate with SGs. Yet, we could demonstrate that depletion of SUMO2/3, the E3 SUMO ligase PML or RNF4 as well as chemical inhibition of SUMOylation strongly delayed SG clearance upon stress release, indicating that a functional STUbL pathway is essential for the timely clearance of SGs.
Next, we investigated how stress-induced poly-SUMOylation is regulated. Our data shows that SENP levels and activities are reduced in response to heat and arsenite stress, which allows the buildup of poly-SUMO chains on nuclear proteins. Limitation of poly-SUMOylation by overexpression of the SUMO chain-specific isopeptidases SENP6 and SENP7 induced SG formation. In contrast, poly-SUMO-priming by chemical depletion of SENP6 with the drug hinokiflavone drastically limited SG formation upon stress treatment. These results indicate a clear role of chain-specific SENPs in the regulation of stress-induced poly-SUMOylation and SG dynamics.
Last, we investigated whether the STUbL pathway could affect the phase separation of FUSP525 (an ALS-linked mutant of the SG-associated protein FUS) and observed that perturbations of the STUbL pathway lead to an increased phase separation of FUSP525L.
Thus, our work connects the SUMO/RNF4 protein quality control mechanism to the dynamics of SGs supporting the hypothesis that release of proteotoxic stress in the nucleus facilitates the clearance of cytosolic SGs. Thereby, we discovered a previously unknown link between the nuclear and cytosolic axis of proteotoxic stress response.
In the past decade, tissue-resident innate lymphoid cells (ILC) have become a central field of immunological research. ILC are a family of innate immune cells comprising cytotoxic Natural Killer (NK) cells and the non-cytotoxic helper like ILC1, ILC2 and ILC3. They mirror the functions and phenotypes of T cells, but do not require rearranged antigen-specific receptors for their rapid response to signals from injured or infected tissue. As potent cytokine producers being enriched in mucosal tissue, ILC play an essential role in tissue maintenance and regulating immunity to chronic inflammation and infection (Vivier et al., 2018). Although heterogeneity and plasticity of ILC complicates their classification, the pathophysiology of a broad variety of autoimmune and chronic inflammatory diseases have been associated with dysregulations in ILC subset distribution and functions (Dzopalic et al., 2019). This highlights their importance in human health and disease and accounts for the need for markers unambiguously describing the different ILC subtypes. This work introduces NKp65, a C-type lectin-like receptor (CTLR) encoded in the natural killer gene complex by the KLRF2 gene, as an exclusive marker for human ILC3. NKp65 expression especially discerns ILC3-like NK cell precursor from mature NK cells which express the NKp65-relative NKp80. Moreover, flow cytometric analysis of NKp65 expression aids in the demarcation of natural cytotoxicity receptor (NCR) expressing ILC3, from the closely related but functionally distinct RORt+ LTi cells and NCR- ILC3. This work further provides insights into NK cell development by in vitro differentiation studies in which NKp65 expressing cells are generated in presence of OP9 feeder cells and cytokines to support development. In such cultures, NKp65 expressing in vitro ILC (ivILC) acquire NKp80 expression in a Notch-dependent manner indicating their differentiation into mature NK cells. Acquisition of NK cell phenotypic markers is accompanied by NKp65 downregulation which leads to the mutually exclusive expression of NKp80 on NK cells and NKp65 on ILC3-like cells. Further insights are provided into the functional consequences of NKp65 engagement by its cognate high affinity ligand ‘keratinocyte-associated C-type lectin’ (KACL) which is selectively expressed on human keratinocytes (Bauer et al., 2015; Spreu et al., 2010). Expressed on ivILC, NKp65 mediates killing of KACL expressing target cells, suggesting that NKp65-KACL interaction promotes cellular cytotoxicity. In this context, the observed metalloproteinase dependent shedding of NKp65 might play a role in the termination of the cellular interaction. The findings on the regulation of NKp65 expression demonstrate the presence of a functional STAT5 response element in the KLRF2 promoter endowing a transcriptional control of NKp65 expression by IL-7 signaling. This provides an interesting link between the dependency of ILC3 on IL-7 signaling for their maintenance and the specific expression of NKp65 on these cells.
In summary, this study provides new insights into the physiologic expression of the CTLR NKp65 on human ILC3. The dependency of NKp65 surface expression on sustained STAT5 signaling provided by IL-7 underlines the connection of NKp65 expression and an ILC3 phenotype which might contribute to promote future research in discerning the interspersed pathways of ILC3 and NK cell development. The tissue and cell specific expression of NKp65 on ILC3 and its ligand KACL on keratinocytes of the human skin further suggests an important role of this genetically coupled receptor-ligand pair in tissue specific immunosurveillance.
[Nachruf] Hugo Fasold
(2018)
Den photoaktivierbaren Schutzgruppen PPGs wurde als wichtige Werkzeuge, um z. B. biologische Prozesse zu untersuchen, in den letzten Jahren eine beachtliche Aufmerksamkeit zuteil. Der Einsatz von PPGs weist gegenüber chemischen Schutzgruppen wertvolle Vorteile auf, was deren Einsatz für biologische Konzepte attraktiv macht. Insbesondere, da keine weiteren Reagenzien außer Licht als Mittel für die Photolyse benötigt werden. Darüber hinaus ist es möglich, durch Einsatz moderner Lasertechnik, eine homogene Bestrahlung des Reaktionsvolumens mit einer hohen Lichtdosis auf einer, im Vergleich zu klassischen Lichtquellen, kürzeren Zeitskala zu gewährleisten.
Die Diversität der einsetzbaren photosensitiven Schutzgruppen, kommt einerseits der Vielfalt der anwachsenden biochemischen Fragestellungen insofern zugute, als dass die ausgewählten PPGs auf verschiedenste Anforderungen der zu untersuchenden Systeme zugeschnitten werden können. Anderseits kann die, durch einige Problemstellungen, erforderte chromatische Orthogonalität der eingesetzten Schutzgruppen, gewährleistet
werden, deren Umsetzung sich in den letzten Jahren in zahlreichen Studien als erfolgsversprechend erwies. Beide Aspekte sind unter anderem Gegenstand der vorliegenden Arbeit.
Zum einen sollte das Photocaged Puromycin als photolabiles Antibiotikum Derivat, mithilfe der Cumarinylmethyl-Schutzgruppe (DEACM-Puromycin) optimiert werden und mit dem vorherigen Nitrobenzyl-geschützten NVOC-Puromycin mittels spektroskopischer und biochemischer Methoden verglichen werden. Zum anderen sollte eine neue Strategie etabliert werden, mit deren Hilfe das photolytisch entschützte Puromycin erneut deaktiviert werden kann.
DEACM-Puromycin konnte mithilfe eines fünfstufigen Syntheseweges hergestellt werden. Ausgehend von 7-Amino-4-methylcumarin, dessen allylische Methylgruppe durch die Riley-Reaktion mit Selendioxid Se2O zum entsprechenden DEACM-Aldehyd oxidiert wurde und anschließende Reduzierung in Anwesenheit von NaBH4 zum Cumarinalkohol DEACM-OH. Eine nicht toxische Variante, bei welcher Selendioxid umgangen wurde, zeichnete sich ebenso als zielführend aus. DEACM-OH und Puromycin konnten im Anschluss über ein Carbonat-Intermediat miteinander als Carbamat verknüpft und mithilfe von HPLC aufgetrennt werden.
Die Vorrangigkeit des neuen photolabilen Puromycin Derivates (DEACM-Puromycin), wurde zuerst mithilfe der Laser-NMR-Spektroskopie sowie HPLC Verfahren erfasst. Spektroskopische Analysen im Rahmen einer Kollaboration mit dem AK von Professor Wachtveitl bestätigten, dass DEACM-Puromycin für biologische Anwendungen geeignetere photolytische Eigenschaften, wie z. B. einen höheren Extinktionskoeffizienten, eine bathochrome Verschiebung des Absorptionsmaximums, sowie eine höhere Quantenausbeute und Uncaging Effizienz der Photolyse, aufwies. Basierend auf einer vergleichbaren HOMO-LUMO Differenz beider Verbindungen (DEACM-OH und DEACM-Puromycin), konnte die Spaltung der Schutzgruppe mithilfe der Differenz der Fluoreszenzslebensdauern mit einer Rate von 0,71*108 s-1 charakterisiert werden. Dies war im Vergleich zum vorherigen NVOC-Puromycin um eine Größenordnung höher. Weitere durchgeführte spektroskopische Methoden wurden mittels quantenchemischer Rechnungen unterstützt, um wichtige Erkenntnisse der kinetischen und dynamischen Abläufe der Photolyse des geschützten Puromycins anzueignen, z. B.:
• Der zur Photolyse von DEACM-Puromycin konkurrierende Fluoreszenzprozess, kann durch protische Medien unterdrückt werden. Dies und die somit ermöglichten Wasserstoffbrückenbindungen, welche die entstehenden ionischen Intermediate während der Photolyse stabilisieren, könnten sich für die Erhöhung der Quantenausbeute der photolytischen Abspaltung von DEACM-Puromycin zu Nutze gemacht werden.
• Anhand von Experimenten auf der ultrakurzen Zeitskala wurde die Population eines angeregten S1-ICT-Zustandes detektiert. Bei diesem findet, in Anwesenheit von polarem Lösungsmittel, einen Ladungstransfer der Diethylaminoreste auf den Cumarinring statt.
• Polare Lösungsmittel bewirken ebenfalls den Übergang zu einem TICT Zustand, welcher als nichtstrahlende Relaxation die Fluoreszenz des DEACM-Puromycins reduziert. Die Auswahl von Substituenten sowie polaren und protischen Lösungsmitteln zur Begünstigung der ICT- sowie TICT- Zustände, könnte zukünftig zur Optimierung der Photolyse Effizienz herangezogen werden.
Die gelungene Optimierung des geschützten Puromycins als ein photosensitives Antibiotikum, durch die DEACM-Schutzgruppe, konnte mittels eines XTT-Zellviabilität-Experimentes mit sf9-Insektenzellen nachgewiesen werden. Zusätzlich konnte die lichtkontrollierte Puromycylierung zur Visualisierung neu synthetisierter Proteine als weitere Funktion des optimierten photoaktivierbaren Puromycins in Zusammenarbeit mit dem AK. Schumann (MPI für
Hirnforschung, Frankfurt am Main) nachgeprüft werden. Obwohl beide photocaged Verbindungen, DEACM- sowie das vorherige NVOC-Puromycin, eine vergleichbare Zellpermeabilität zu den präparierten Neurozellen aufwiesen, zeigte DEACM-Puromycin unter gleichen Bedingungen nach der Belichtung ein signifikant intensiveres Puromycylierungsignal als NVOC-Puromycin. Unter der Annahme, dass sich mehr NVOC- als DEACM-Puromycin in
den Zellen befand, bestätigt diese Beobachtung die Vorrangigkeit des DEACM-Derivates, aufgrund seiner bereits optimierten photochemischen Eigenschaften. Durch den Einsatz eines Zwei-Photonen-Lasers, konnte die Eignung von DEACM-Puromycin für die raumselektive Steuerung der Detektion von neu exprimierten Proteinen, mit größerer Auflösung auf subzellularem Level, nachgewiesen werden.
...
This PhD thesis is dedicated to the extension of the portfolio of nuclear magnetic resonance (NMR) methods to characterize ribonucleic acids (RNAs). Only within the last few decades it has been realized that the cellular role of RNA goes well beyond the central dogma of molecular biology. In fact, RNA takes part in numerous cellular processes, executes numerous functions and acts either as a single player or in larger complexes, mostly RNA-protein complexes (RNPs) such as the ribosome or the spliceosome. This versatility in RNA function is coupled to a structural variety and the ability to adopt multiple long-lived and intricate conformations. Due to this high molecular complexity special demands are placed on the methods that are required for RNA structural characterization. With the ability to capture dynamics at atomic resolution and to measure under close to native conditions, NMR spectroscopy is undoubtedly a prime method for this purpose.
A general introduction to the current state of research, selected achievements as well as challenges in the field of NMR spectroscopy on RNA is given in Chapter I. This thesis is further composed of three independent chapters covering the three separate projects, which form the main body of work within the course of this thesis.
The imino group found in two of the four RNA nucleobases is generally considered to be the most powerful reporter group in the process of the NMR spectroscopic characterization of RNA. Its resonance assignment provides key information for a rapid determination of the RNA’s secondary structure. This is possible, since the imino proton can only be detected, if it is protected from rapid solvent exchange through hydrogen bonding interactions or, in rare cases, steric shielding. Consequently, information on flexible regions of RNA that are not protected against solvent exchange cannot be derived using this NMR spy. It is a key finding of the thesis that nucleobase interactions can also be mapped through the amino groups, as they similarly take part in base pairing or RNA-ligand interactions. Notably, solvent exchange of the amino protons is always slower compared to the imino proton. Thus, 1H,15N resonances of the amino group can be detected even for dynamic regions of RNA. Moreover, focusing on characterizing amino groups of RNA nucleobases increases the number of available reporters as amino groups are present in three out of four RNA nucleobases.
However, there is a reason that up to work conducted in this thesis, amino groups have not been used for monitoring RNA nucleobases: the rate of the C-NH2 bond rotation is most often close to the chemical shift differences of the two non-identical amino proton resonances, in particular for guanosines and adenosines, amino resonances regularly remain elusive in NMR spectra. Therefore, we developed experiments that excite double quantum (DQ) coherences of the two amino group protons and that further utilize 13C-detection. Results on these experiments are discussed in Chapter II and show that the rotational exchange can be avoided by evolving double quantum instead of single quantum (SQ) coherence in the indirect dimension of a 13C-detected C(N)H-HDQC experiment. The new experiment enables the detection of a full set of sharp amino resonances. The advantages of this experiment are immediately apparent when comparing the number of observable imino resonances in a classic 1H,15N-HSQC spectrum with the number of amino resonances obtained in the 13C-detected C(N)H-HDQC spectrum of the same RNA.
Furthermore, based on the newly available resonance assignment of amino groups, we developed a 13C-detected “amino”-NOESY experiment to obtain precious additional structural restraints. The 13C-detected “amino”-NOESY experiment enables the observation of NOE contacts that are not accessible using other 1H-detected NOESY experiment. Among these new NOE contacts are valuable, inter-residual correlations, which are otherwise scarce in RNA due to the proton deficiency of its nucleobases. We showed that the newly obtained NOE contacts are especially important in the structure determination of RNAs with only few NOE restraints. Under such circumstances, the inclusion of the newly obtained amino NOE contacts lead to a significant improvement in the root-mean-square deviation (RMSD) of the three-dimensional structure of the 34 nts GTP class II aptamer. Together the novel 13C-detected NMR experiments developed within this PhD project provide a valuable alternative for the imino-based characterization of nucleobase interactions.
...
Die Akademische Feier der Vereinigung von Freunden und Förderern der Goethe-Universität schafft es wie keine andere Veranstaltung, jedes Jahr aufs Neue in nur knapp zwei Stunden den Wissenschaftskosmos dieser Universität abzubilden: In kurzweiligen, auch für Laien verständlichen Vorstellungen skizzierten die Laudatoren, worüber die 13 Preisträgerinnen und Preisträger in ihren ausgezeichneten Arbeiten geforscht hatten.