STUDIO DI COMPOSTI TRIAZINICI MEDIANTE SPETTROSCOPIA NMR

L’analisi conformazionale di un derivato della triazina doppiamente sostituito con gruppi dialchilamminici uguali (chiamato Biboc) è stata affrontata mediante spettri NMR 1H e del 13C. Le quattro catene legate agli azoti amminici, chimicamente uguali, a temperatura ambiente in CDCl3 si differenziano a due a due per i chemical shifts di tutti i nuclei, per la larghezza dei segnali ecc.. Alcuni spettri bidimensionali (COSY, HETCOR e NOESY) hanno permesso di correlare definitivamente tra loro i segnali relativi a ciascun tipo di catena e di mettere a fuoco alcuni interrogativi, fra i quali segnaliamo i tre principali: Quali le condizioni conformazionali che differenziano i due tipi di catene, A e B? Qual è la barriera energetica per l’interconversione della catena A nella B e viceversa? La diversa larghezza di riga nelle catene A e B può essere dovuta alla loro diversa mobilità? La ricerca della conformazione più stabile è stata affidata al calcolo ab initio; i risultati teorici di chemical shift sono stati confrontati con quelli sperimentali. La barriera energetica per l’interconversione A-B (energia di attivazione 12-14 kcal/mole) è stata valutata dalle temperature di coalescenza rilevate su spettri 1H NMR ad alta temperatura. La mobilità dei diversi frammenti molecolari è stata messa in evidenza da misure di tempi di rilassamento T1 del protone in funzione della temperatura. In complesso sono stati studiati due campioni di Biboc in CDCl3 con diversa concentrazione, un campione di Biboc in DMSO-d6 e, per confronto, un campione di un’analoga triazina trisostituita in CDCl3

Computational investigation on the mechanism of oxygen tolerance in the radical SAM enzyme lysine 2,3-aminomutase

The radical S-adenosylmethionine (AdoMet or SAM) enzyme superfamily represents an ensemble of proteins that are able to catalyse biochemical reactions involving organic radical intermediates. These intermediate radicals then undergo a wide range of reactions, many of them difficult to accomplish in the laboratory. The products of such reactions are bioactive compounds of pharmaceutical interest that can be more over used as building blocks for other compounds. Virtually every enzyme belonging to this family is known to be unstable in air due to the requirement for a catalytic, oxygen sensitive [Fe4-S4] cluster that decomposes after oxidative attack by reactive oxygen species (ROS), inactivating the enzyme. The study of these enzymes and their possible application in biotechnology is difficult due to the oxygen sensitivity of the [Fe4-S4] cluster forcing their usage under strictly inert atmosphere. C. subterminale lysine 2,3-aminomutase (CsLAM) is a widely studied radical AdoMet enzyme and a natural oxygen-tolerant variant from B. subtilis (BsLAM) was discovered that catalyses, in presence of air, the interconversion between α- and β-lysine. This project utilised computational methodologies for the assessment of how radical SAM enzymes may manage to shield their FeS cluster from air degradation. Comparison was made of the CsLAM crystal structure with the structural model of oxygen-tolerant BsLAM, here obtained through homology modelling using the 3D structure as its template. Following the validation of the model through PCA, both the CsLAM and the BsLAM enzyme structures were compared and molecular dynamics simulations were used to identify different ways the two enzymes might deal with oxygen. The tunnel searching software CAVER was used to identify those amino acid residues that could obstruct oxygen flow due to their size or their ability to trap oxygen in BsLAM. Quantum mechanics calculations were then performed on the [Fe4-S4] sub-system to retrieve information about the difference in the electrostatics governed by the protein environment. The difference in electrostatics could account for different redox potentials in the two enzymes by making the BsLAM less prone to oxidation by ROS. A selection of amino acid residues were identified as likely to affect the redox potential and mutants of the CsLAM bearing such residues were created. The observation of their dipole moment suggested that the double mutation H131Y/A138S could positively affect the oxygen tolerance in the enzyme

sintesi di derivati idrossi-piridonici a potenziale attivita' antitumorale.

Sono stati sintetizzati e caratterizzati alcuni derivati idrossi-piridonici a potenziale attivita' antitumorale come analoghi strutturali alla ciclopirox olamina, farmaco antifungino con proprietà antitumorali

Deploying Proteins as Electrolyte Additives in Li–S Batteries: The Multifunctional Role of Fibroin in Improving Cell Performance

It is widely accepted that the commercial application of lithium–sulfur batteries is inhibited by their short cycle life, which is primarily caused by a combination of Li dendrite formation and active material loss due to polysulfide shuttling. Unfortunately, while numerous approaches to overcome these problems have been reported, most are unscalable and hence further hinder Li–S battery commercialization. Most approaches suggested also only tackle one of the primary mechanisms of cell degradation and failure. Here, we demonstrate that the use of a simple protein, fibroin, as an electrolyte additive can both prevent Li dendrite formation and minimize active material loss to enable high capacity and long cycle life (up to 500 cycles) in Li–S batteries, without inhibiting the rate performance of the cell. Through a combination of experiments and molecular dynamics (MD) simulations, it is demonstrated that the fibroin plays a dual role, both binding to polysulfides to hinder their transport from the cathode and passivating the Li anode to minimize dendrite nucleation and growth. Most importantly, as fibroin is inexpensive and can be simply introduced to the cell via the electrolyte, this work offers a route toward practical industrial applications of a viable Li–S battery system

Computational investigation on the mechanism of oxygen tolerance in the radical SAM enzyme lysine 2,3-aminomutase

The radical S-adenosylmethionine (AdoMet or SAM) enzyme superfamily represents an ensemble of proteins that are able to catalyse biochemical reactions involving organic radical intermediates. These intermediate radicals then undergo a wide range of reactions, many of them difficult to accomplish in the laboratory. The products of such reactions are bioactive compounds of pharmaceutical interest that can be more over used as building blocks for other compounds. Virtually every enzyme belonging to this family is known to be unstable in air due to the requirement for a catalytic, oxygen sensitive [Fe4-S4] cluster that decomposes after oxidative attack by reactive oxygen species (ROS), inactivating the enzyme. The study of these enzymes and their possible application in biotechnology is difficult due to the oxygen sensitivity of the [Fe4-S4] cluster forcing their usage under strictly inert atmosphere. C. subterminale lysine 2,3-aminomutase (CsLAM) is a widely studied radical AdoMet enzyme and a natural oxygen-tolerant variant from B. subtilis (BsLAM) was discovered that catalyses, in presence of air, the interconversion between α- and β-lysine. This project utilised computational methodologies for the assessment of how radical SAM enzymes may manage to shield their FeS cluster from air degradation. Comparison was made of the CsLAM crystal structure with the structural model of oxygen-tolerant BsLAM, here obtained through homology modelling using the 3D structure as its template. Following the validation of the model through PCA, both the CsLAM and the BsLAM enzyme structures were compared and molecular dynamics simulations were used to identify different ways the two enzymes might deal with oxygen. The tunnel searching software CAVER was used to identify those amino acid residues that could obstruct oxygen flow due to their size or their ability to trap oxygen in BsLAM. Quantum mechanics calculations were then performed on the [Fe4-S4] sub-system to retrieve information about the difference in the electrostatics governed by the protein environment. The difference in electrostatics could account for different redox potentials in the two enzymes by making the BsLAM less prone to oxidation by ROS. A selection of amino acid residues were identified as likely to affect the redox potential and mutants of the CsLAM bearing such residues were created. The observation of their dipole moment suggested that the double mutation H131Y/A138S could positively affect the oxygen tolerance in the enzyme

Tocilizumab for patients with COVID-19 pneumonia. The single-arm TOCIVID-19 prospective trial

BackgroundTocilizumab blocks pro-inflammatory activity of interleukin-6 (IL-6), involved in pathogenesis of pneumonia the most frequent cause of death in COVID-19 patients.MethodsA multicenter, single-arm, hypothesis-driven trial was planned, according to a phase 2 design, to study the effect of tocilizumab on lethality rates at 14 and 30 days (co-primary endpoints, a priori expected rates being 20 and 35%, respectively). A further prospective cohort of patients, consecutively enrolled after the first cohort was accomplished, was used as a secondary validation dataset. The two cohorts were evaluated jointly in an exploratory multivariable logistic regression model to assess prognostic variables on survival.ResultsIn the primary intention-to-treat (ITT) phase 2 population, 180/301 (59.8%) subjects received tocilizumab, and 67 deaths were observed overall. Lethality rates were equal to 18.4% (97.5% CI: 13.6-24.0, P=0.52) and 22.4% (97.5% CI: 17.2-28.3, P<0.001) at 14 and 30 days, respectively. Lethality rates were lower in the validation dataset, that included 920 patients. No signal of specific drug toxicity was reported. In the exploratory multivariable logistic regression analysis, older age and lower PaO2/FiO2 ratio negatively affected survival, while the concurrent use of steroids was associated with greater survival. A statistically significant interaction was found between tocilizumab and respiratory support, suggesting that tocilizumab might be more effective in patients not requiring mechanical respiratory support at baseline.ConclusionsTocilizumab reduced lethality rate at 30 days compared with null hypothesis, without significant toxicity. Possibly, this effect could be limited to patients not requiring mechanical respiratory support at baseline.Registration EudraCT (2020-001110-38); clinicaltrials.gov (NCT04317092)

Correction to: Tocilizumab for patients with COVID-19 pneumonia. The single-arm TOCIVID-19 prospective trial (Journal of Translational Medicine, (2020), 18, 1, (405), 10.1186/s12967-020-02573-9)

Following publication of the original article [1] the authors identified that the collaborators of the TOCIVID-19 investigators, Italy were only available in the supplementary file. The original article has been updated so that the collaborators are correctly acknowledged. For clarity, all collaborators are listed in this correction article