A5070781963- Enzyme function and inhibition
- Protein Structure and Dynamics
- Ion channel regulation and function
- Advanced NMR Techniques and Applications
- Viral Infectious Diseases and Gene Expression in Insects
- Trace Elements in Health
- Advanced MRI Techniques and Applications
- X-ray Diffraction in Crystallography
- Chemical Reactions and Mechanisms
- ATP Synthase and ATPases Research
- Crystallization and Solubility Studies
- Amyotrophic Lateral Sclerosis Research
- Mitochondrial Function and Pathology
- Redox biology and oxidative stress
- Amino Acid Enzymes and Metabolism
- Enzyme Structure and Function
- Liver physiology and pathology
- Virus-based gene therapy research
- Photosynthetic Processes and Mechanisms
- Electron Spin Resonance Studies
- Liver Disease and Transplantation
- Cholinesterase and Neurodegenerative Diseases
- Phenothiazines and Benzothiazines Synthesis and Activities
- RNA and protein synthesis mechanisms
- Histone Deacetylase Inhibitors Research
Interuniversity Consortium for Magnetic Resonance
2016-2025
University of Florence
2014-2024
Federico II University Hospital
2019
University of Naples Federico II
2017-2018
Queen Mary University of London
2017
William Harvey Research Institute
2017
Istituto Mediterraneo per i Trapianti e Terapie ad Alta Specializzazione
2017
University of Pavia
2013
Superoxide dismutase-1 (SOD1) mutants, including those with unaltered enzymatic activity, are known to cause amyotrophic lateral sclerosis (ALS). Several destabilizing factors contribute pathogenicity a reduced ability complete the normal maturation process which comprises folding, metal cofactor acquisition, intra-subunit disulphide bond formation and dimerization. Immature SOD1 forms toxic oligomers characteristic large insoluble aggregates within motor system cells. Here we report that...
Abstract In-cell NMR spectroscopy provides precious structural and functional information on biological macromolecules in their native cellular environment at atomic resolution. However, the intrinsic low sensitivity of imposes a big limitation applicability methodology. In this respect, recently developed commercial 1.2 GHz spectrometer is expected to introduce significant benefits. cell samples may suffer from detrimental effects ultrahigh fields, that must be carefully evaluated. Here we...
In-cell NMR spectroscopy is a powerful approach to study protein structure and function in the native cellular environment. It provides precious insights into folding, maturation, interactions, ligand binding of important pharmacological targets directly human cells. However, its widespread application hampered by fact that soluble globular proteins often interact with large components, causing severe line broadening conventional heteronuclear experiments. 19F can overcome this issue, as...
In-cell NMR allows obtaining atomic-level information on biological macromolecules in their physiological environment. Soluble proteins may interact with the cellular environment different ways: either specifically, functional partners, or non-specifically, other components. Such behaviour often causes disappearance of signals. Here we show that by introducing mutations human protein profilin 1, used here as a test case, in-cell signals can be recovered. In cells both specific and...
Abstract Structure‐based drug development is often hampered by the lack of in vivo activity promising compounds screened vitro, due to low membrane permeability or poor intracellular binding selectivity. Herein, we show that ligand screening can be performed living human cells “intracellular protein‐observed” NMR spectroscopy, without requiring enzymatic measurements other cellular assays. Quantitative information obtained fast, inexpensive 1 H experiments, providing dose‐ and time‐dependent...
In-cell NMR can investigate protein conformational changes at atomic resolution, such as those induced by drug binding or chemical modifications, directly in living human cells, and therefore has great potential the context of development it provide an early assessment potency. bioreactors greatly improve cell sample stability over time and, more importantly, allow for recording in-cell data real to monitor evolution intracellular processes, thus providing unique insights into kinetics...
In-cell NMR allows characterizing the folding state of a protein as well posttranslational events at molecular level, in cellular context. Here, initial maturation steps human copper, zinc superoxide dismutase 1 are characterized E. coli cytoplasm by in-cell NMR: from apo protein, which is partially unfolded, to binding causes its final quaternary structure. The selectively binds only one ion, whereas vitro also copper site non-physiological ion. However, no intramolecular disulfide bridge...
Superoxide dismutase 1 (SOD1) is an important metalloprotein for cellular oxidative stress defence, that mutated in familiar variants of Amyotrophic Lateral Sclerosis (fALS). Some mutations destabilize the apo protein, leading to formation misfolded, toxic species. The Copper Chaperone SOD1 (CCS) transiently interacts with and promotes its correct maturation by transferring copper catalyzing disulfide bond formation. By vitro in-cell NMR, we investigated role SOD-like domain CCS (CCS-D2). We...
FlowNMR has the aim of continuously monitoring processes that occur in conditions are not compatible with being carried out within a closed tube. However, it is sample intensive and suitable for samples, such as proteins or living cells, often available limited volumes possibly low concentrations. We here propose dialysis-based modification commercial flowNMR setup allows recycling medium while confining (proteins cells) active volume This approach demonstrated specific cases in-cell NMR...
Candidate drugs rationally designed in vitro often fail due to low efficacy vivo caused by tissue availability or because of unwanted side effects. To overcome the limitations rational drug design, binding candidate their target needs be evaluated cellular context. Here, we applied in-cell NMR investigate a set approved isoform II carbonic anhydrase (CA) living human cells. Some compounds were originally developed toward other targets and later found inhibit CAs. We observed strikingly...
Fluorinated aromatic amino acids (FAAs) are promising tools when studying protein structure and dynamics by NMR spectroscopy. The incorporation FAAs in mammalian expression systems has been introduced only recently. Here, we investigate the effects of proteins expressed human cells, focusing on probability its consequences
Most of our knowledge protein structure and function originates from experiments performed with purified proteins resuspended in dilute, buffered solutions. However, most crowded intracellular environments complex compositions. Significant efforts have been made to develop tools study their native cellular settings. Among these tools, in-cell NMR spectroscopy has the sole technique for characterizing space living cells at atomic resolution physiological temperature. Nevertheless, due...
Systemic lupus erythematosus (SLE) is associated with increased risk of cardiovascular disease (CVD). Among many mechanisms, accelerated atherosclerosis, endothelial dysfunction, and hypercoagulability play a main role. Here, we investigate whether inflammatory, serological clinical markers SLE determine correlate arterial stiffness in patients. Routine blood samples, inflammatory mediators, specific antibodies, 24 h proteinuria were measured 43 patients age sex-matched controls using...
Structure-based drug development suffers from high attrition rates due to the poor activity of lead compounds in cellular and animal models caused by low cell penetrance, off-target binding or changes conformation target protein environment. The latter two effects cause a change apparent affinity compound, which is indirectly assessed assays. To date, direct measurement intracellular remains challenging task. In this work, in-cell NMR spectroscopy was applied measure dissociation constants...
The development of more effective drugs requires knowledge their bioavailability and binding efficacy directly in the native cellular environment. In-cell nuclear magnetic resonance (NMR) spectroscopy is a powerful tool for investigating ligand–target interactions living cells. However, target molecule may be NMR-invisible due to with components, while observing ligand by 1H NMR impractical background. Such limitations can overcome fluorinated ligands 19F in-cell as they bind intracellular...
Background: Thyroid function abnormalities and thyroid autoantibodies have been frequently described in patients with systemic autoimmune diseases as Systemic Sclerosis (SSc). Serum TSH levels are higher SSc more severe skin a worse modified Rodnan score (mRSS). Asymptomatic esophageal involvement due to has never cause of hypothyroidism L-T4 malabsorption HT SSc. Case report: Here we report case 56-year-old female affected by both Hashimoto's thyroiditis (HT) who developed the loss...