Chicken ceruloplasmin. Evidence in support of a trinuclear cluster involving type 2 and 3 copper centers.

Ceruloplasmin was isolated to purity from chicken plasma by a single-step chromatography on amino-ethyl-derivatized Sepharose. Molecular mass, as estimated by nonreducing sodium dodecyl sulfate-electrophoresis, was approximately 140 kDa, slightly higher than that found for ceruloplasmins from other sources. Specific activity as p-phenylenediamine oxidase was five times higher than that reported for mammalian ceruloplasmins. The copper content was estimated to be 5.01 +/- 0.35 atoms per protein molecule, 50% of which was EPR-detectable. The EPR spectrum was completely devoid of any signal typical of the type 2 copper as seen in the other blue multicopper oxidases and in ceruloplasmin from mammalian species. Anaerobic reduction of chicken ceruloplasmin resulted in the disappearance of the 330 nm optical band typical of type 3 copper, which was followed by the appearance of an EPR signal typical of type 2 copper. Subsequently, the type 1 copper and finally the newly formed type 2 copper were reduced. The original optical and EPR spectra were recovered within few minutes upon exposure of reduced ceruloplasmin to air. It is concluded that in oxidized chicken ceruloplasmin type 2 copper interacts with the diamagnetic pair responsible for the 330 nm absorption in such a way as to become EPR-undetectable and that the interaction is relieved by reduction of the pair. Whether this interaction is intrinsically weaker in other blue oxidases and ceruloplasmins studied or is lost with standard preparation procedures remains to be established

Presence of coupled trinuclear copper cluster in mammalian ceruloplasmin is essential for efficient electron transfer to oxygen.

Abstract The reactivity with dioxygen of a mammalian (sheep) ceruloplasmin, anaerobically reduced with ascorbate, was found to depend on the state of the Type 2 and Type 3 copper centers, as monitored by EPR and optical spectroscopy. A complete reoxidation by air after anaerobic reduction with ascorbate was observed with samples (A) purified by the single-step procedure described for chicken ceruloplasmin (Calabrese, L., Carbonaro, M., and Musci, G. (1988) J. Biol. Chem. 263, 6480-6483), while samples prepared by traditional multistep procedure (B) or subjected to freeze-thawing (C) displayed partial and very slow reoxidation, reflecting the functional nonequivalence of blue coppers which is considered a typical property of mammalian ceruloplasmin. The rate of reduction of the 330 nm chromophore was found to increase as a function of the extent and rate of reoxidation of different samples, while the 610 nm band displayed an opposite trend. Samples B and C showed a Type 2 copper signal in the EPR spectrum, while sample A showed practically no Type 2 copper in the oxidized protein, and a transient Type 2-like signal during reduction. The presence of a trinuclear Type 2-Type 3 cluster can therefore be proposed for all ceruloplasmins, and the integrity of the copper-copper coupling is essential for efficient oxidase behavior

La responsabilità civile da illecito trattamento dei dati personali

La tesi affronta il tema della responsabilità civile da illecito trattamento dei dati personali (art. 82 GDPR). Il lavoro si pone l’obbiettivo di dimostrare che, mentre il diritto alla privacy è un diritto essenzialmente personalistico, il diritto alla protezione dei dati è, solo parzialmente, un diritto soggettivo. Per poter predisporre strumenti di tutela più efficaci, è necessario abbandonare la concezione prettamente individualista, che comporta un sistema di responsabilità ad azione del singolo. In particolare, quando i trattamenti involgono attività di profilazione e big data, dovrebbero essere garantite anche finalità di sviluppo etico e sociale. Il diritto alla protezione dei dati dovrebbe, dunque, considerare maggiormente i rischi collettivi e il sistema di accountability dovrebbe essere integrato da obblighi di trasparenza maggiori e da un sistema di controlli più penetranti ed incisivi da parte delle autorità di controllo. La prima parte del lavoro si concentra sull’evoluzione che ha portato all’affermazione del diritto alla protezione dei dati, quale espressione del diritto alla privacy, tenendo in considerazione il contesto tecnologico in cui tale riconoscimento è maturato. L’analisi ripercorre le principali teorie della dottrina e della giurisprudenza statunitensi che hanno contribuito all’affermazione del diritto alla privacy, per dimostrare che esso viene inteso in chiave essenzialmente personalistica. L’evoluzione storica ripercorre le tappe legislative europee e nazionali sulla disciplina dei dati e culmina con un’analisi dei punti di contatto tra diritto alla protezione dei dati, il diritto alla riservatezza, il diritto all’identità personale, il diritto all’autodeterminazione. Nel secondo capitolo, si evidenzia che l’evoluzione esponenziale degli strumenti tecnologici dell’epoca moderna mette in crisi la concezione del diritto alla protezione dei dati in chiave essenzialmente personalistica. In particolare, la creazione di internet, le piattaforme informatiche, i social network, lo sviluppo dell’intelligenza artificiale e gli algoritmi predittivi, rischiano di rendere del tutto inefficiente e inadeguato un sistema di tutela basato su una concezione meramente personalistica del diritto alla protezione dei dati. Si analizzano i rischi collettivi/sociali del trattamento dei dati, illustrando le principali teorie della sorveglianza. Nel terzo capitolo si analizzano le figure soggettive di cui all’art. 82 GDPR e i principi del trattamento. L’analisi evidenzia che il meccanismo di tutela ancora fondato sul consenso determina una forte perdita di effettività del sistema di tutela dei dati. Il GDPR si focalizza maggiormente sulla figura del titolare, su cui gravitano obblighi pregnanti, in particolare quello di accountability. Il titolare è tenuto a svolgere un’attività di ponderazione dei rischi per mettere in atto misure tecniche e organizzative per evitarli o mitigarli. Egli è tenuto a monitorare costantemente i processi della sua attività imprenditoriale che involgono l’utilizzo di dati ed è tenuto a conformare la propria attività nell’osservanza dei principi del GDPR. L’ultima parte del lavoro di ricerca tratta della natura della responsabilità per illecito trattamento dei dati personali. La tesi approfondisce il modello di gestione del rischio disegnato dal GDPR, secondo cui il titolare deve mantenere un approccio proattivo nel prevenire i rischi, dotandosi di una struttura organizzativa che, a tutti i livelli e aree di competenza, adotti le necessarie misure per garantire che i dati personali siano trattati sempre in modo lecito. Da ultimo il lavoro di ricerca approfondisce il problema dei danni risarcibili a seguito di trattamento illecito, con riferimento specifico alla questione dei danni non patrimoniali.This research thesis addresses the issue of civil liability for unlawful processing of personal data (art. 82 GDPR). The research aims to demonstrate that, while the right to privacy is an essentially personal right, the right to data protection is, only partially, a subjective right. In order to provide more effective protection tools, it is necessary to abandon the purely individualistic conception, which involves a system of responsibility based on the action of the individual. In particular, when processing involves profiling and big data, ethical and social development purposes should also be guaranteed. The right to data protection should, therefore, take more account of collective risks and the accountability system should be complemented by greater transparency obligations and a system of more penetrating and incisive controls by supervisory authorities. The first part of the thesis focuses on the evolution that has led to the affirmation of the right to data protection, as an expression of the right to privacy, taking into account the technological context in which this recognition has matured. The analysis retraces the main theories of U.S. doctrine and jurisprudence that have contributed to the affirmation of the right to privacy, to show that it is understood in an essentially personalistic key. The historical evolution traces the stages of European and national legislation on data regulation and culminates with an analysis that highlights how the right to data protection, the right to privacy, the right to personal identity, the right to self-determination are intertwined. In the second chapter, we show that the exponential evolution of technological tools of the modern era undermines the conception of the right to data protection in an essentially personalistic key. In particular, the creation of the internet, computer platforms, social networks, the development of artificial intelligence and predictive algorithms, are likely to make completely inefficient and inadequate a system of protection based on a purely personalistic conception of the right to data protection. The collective/social risks of data processing are analyzed, illustrating the main theories of surveillance. In the third chapter, the subjects referred to in Article 82 GDPR and the principles of processing are analyzed. The analysis shows that the protection mechanism still based on consent determines a strong loss of effectiveness of the data protection system. The GDPR focuses on the figure of the controller, who is burdened by significant obligations, in particular that of accountability. The controller is required to carry out a risk assessment activity in order to implement technical and organizational measures to avoid or mitigate risks. He is required to constantly monitor the processes of his business activity that involve the use of data and is required to comply with the principles of the GDPR. The last part of the research deals with the nature and discipline of liability for unlawful processing of personal data. The thesis delves into the risk management model designed by the GDPR, according to which the controller must adopt a proactive approach in preventing risks through an organizational structure that, at all levels and in all areas of competence, takes the necessary measures to ensure that personal data are always processed lawfully. Lastly, the research explores the issue of the damages resulting from unlawful processing, with specific reference to the issue of immaterial damages. In the European doctrine there is a heated debate around the problem of compensability, both in terms of definition and quantification, of immaterial damages resulting from unlawful processing

Reconstitution of Ceruloplasmin by the Cu(I)-Glutathione Complex: EVIDENCE FOR A ROLE OF Mg2+ AND ATP

The copper-glutathione complex (Cu(I)-GSH) efficiently acted in vitro as the source of Cu(I) in the reconstitution of apoceruloplasmin. Copper was found to reinstate in the various sites in a multistep process, with metal entry into the protein in a first phase, and a second step involving conformational changes of the protein leading to the recovery of the native structural and functional properties. This latter phase was found to be strongly facilitated by Mg2+ or Ca2+ and by ATP. Both Mg2+ and ATP had to be present for optimal reconstitution. These results may shed some light on the mechanisms governing the biosynthesis of ceruloplasmin in vivo. Cu(I)-GSH was the only complex able to reconstitute ceruloplasmin at neutral pH. Glutathione may thus function to shuttle the metal from the membrane copper pump, as the Wilson disease ATPase, and ceruloplasmin in the secretory compartments of the cell. The finding that ceruloplasmin acquires the native conformation after metal entry through a complex pathway triggered by Mg2+ and ATP suggests that they may act as physiological modulators of this process in vivo

Gallbladder Agenesis and Cystic Duct Absence in an Adult Patient Diagnosed by Magnetic Resonance Cholangiography: Report of a Case and Review of the Literature

Gallbladder agenesis (GA) is a rare congenital anomaly of the biliary system often associated with other congenital abnormalities. Patients become symptomatic in 23% of cases. GA is often misinterpreted as other diseases, therefore, leading to unnecessary surgery. We report a case of congenital GA associated to cystic duct absence and a biliary tract abnormality diagnosed by Magnetic Resonance with Cholangiopancreatography

Age-related changes in human ceruloplasmin. Evidence for oxidative modifications.

Human plasma or serum from donors of age comprised between 15 and 95 years was analyzed for paramagnetic and total copper content, as well as for immunoreactive ceruloplasmin content and oxidase activity. All parameters were essentially unaltered, except the paramagnetic copper content, which increased 2-fold upon aging. A dramatic change of the electron paramagnetic resonance spectrum due to ceruloplasmin occurred in individuals over 65 years old and was associated with both an increase of the type 1 copper signal intensity and the appearance of new resonances of a type 2 copper species. Ceruloplasmin was isolated from either young or old donors. Spectroscopic analyses of the isolated proteins confirmed the tendency of type 1 copper to stay reduced in the "young" and oxidized in the "old" protein. The type 2 copper signal observed in most young ceruloplasmin samples was different from the species invariably present in the old protein. The magnetic parameters of the latter species were more consistent with a partially reduced trinuclear copper site. In vitro limited proteolysis resulted in identical fragmentation patterns and kinetics in both proteins. However, changes of the net electric charge were detected in the fragments of the protein isolated from aged individuals, which exhibited a carbonyl content of 0.6 mol of carbonyl/mol of protein. The same pattern of modifications, including a higher carbonyl content (0.65 versus 0.2 mol of carbonyl/mol of protein), could be reproduced by exposure of the young protein to the metal-catalyzed oxidation system iron/ascorbate. These results suggest that during aging ceruloplasmin is subjected to oxidative modifications which are likely to be the source of conformational changes around the copper sites leading to an intramolecular electron rearrangement among the various copper sites

Optimal Operations Management of Hybrid Energy Systems Through Short-Term Atmospheric and Demand Forecasts

Abstract The complexity of systems for energy production through renewable energy sources (RESs) is constantly increasing considering the integration of multiple modules, e.g. different RES sources, energy storage in batteries and connection to national grid for energy trade purpose. This significant complexity could represent a threat but also an opportunity if adequately managed. Aim of this paper is to propose two different approaches to manage the hourly electricity flows between the different components of a hybrid energy system (HES) fueled by PV modules and a wind turbine, equipped with a battery storage system (BES) to satisfy the demand of a user load with the opportunity to sell and purchase the electricity to/from the national grid. The first approach is a heuristic algorithm (HA) which defines robust but constant dispatching criteria of the energy flows between the HES components considering just the current value of energy production and demand with the aim of minimizing the electricity purchased by the grid. On the contrary, the second approach is a mixed integer linear programing (MILP) model which defines the optimal value of the energy flows to maximize the net profit of the HES operations determined by the electricity sales revenues minus the energy purchase costs. The developed MILP leverages the short-term forecast of the atmospheric conditions and user demand as well it considers variable energy sale and purchase pricing in the different daily hours. Both these approaches have been tested and validated through a case study of a residential building in which multiple households live located in the suburban area of Munich (Germany). The obtained results highlight how the MILP outperforms HA considering the net profit achievable weekly due to electricity trade with the grid. In particular, the MILP improve the HA economic performance of the HES operation management of 18% on average over the different months of the year

Healthspan Enhancement by Olive Polyphenols in C. elegans Wild Type and Parkinson’s Models

Parkinson’s disease (PD) is the second most prevalent late-age onset neurodegenerative disorder, affecting 1% of the population after the age of about 60 years old and 4% of those over 80 years old, causing motor impairments and cognitive dysfunction. Increasing evidence indicates that Mediterranean diet (MD) exerts beneficial effects in maintaining health, especially during ageing and by the prevention of neurodegenerative disorders. In this regard, olive oil and its biophenolic constituents like hydroxytyrosol (HT) have received growing attention in the past years. Thus, in the current study we test the health-promoting effects of two hydroxytyrosol preparations, pure HT and Hidrox® (HD), which is hydroxytyrosol in its “natural” environment, in the established invertebrate model organism Caenorhabditis elegans. HD exposure led to much stronger beneficial locomotion effects in wild type worms compared to HT in the same concentration. Consistent to this finding, in OW13 worms, a PD-model characterized by α-synuclein expression in muscles, HD exhibited a significant higher effect on α-synuclein accumulation and swim performance than HT, an effect partly confirmed also in swim assays with the UA44 strain, which features α-synuclein expression in DA-neurons. Interestingly, beneficial effects of HD and HT treatment with similar strength were detected in the lifespan and autofluorescence of wild-type nematodes, in the neuronal health of UA44 worms as well as in the locomotion of rotenone-induced PD-model. Thus, the hypothesis that HD features higher healthspan-promoting abilities than HT was at least partly confirmed. Our study demonstrates that HD polyphenolic extract treatment has the potential to partly prevent or even treat ageing-related neurodegenerative diseases and ageing itself. Future investigations including mammalian models and human clinical trials are needed to uncover the full potential of these olive compounds.Peer Reviewe