89,615 research outputs found
Materials and techniques of Art Nouveau architecture in Italy and Portugal: a first insight for an European route to consistent restoration
No full textThe results of the investigations on building materials and techniques of Casa Major Pessoa, a typical Art
Nouveau construction in Aveiro (Portugal), and two coeval Art Nouveau buildings in Bologna (Italy) are presented
as a methodological contribution to the restoration of this kind of buildings. This is the first step to
ascertain the existence of a common thread between local materials, technologies and architecture in European
countries at the same period. A holistic approach was adopted: materials were investigated along with
architectural, structural and technological features, in order to achieve a first insight into the Art Nouveau
architecture in Europe in particular for its consistent restoration without loss of historical memory
Materiales y tecnologías en la Arquitectura Modernista: casos de estudio de decoración de fachadas en Italia, Portugal y Polonia persiguiendo una restauración racional
No full textThe results of a diagnostic survey on the materials of representative Art Nouveau buildings in Italy, Portugal and Poland are here presented and compared, as a contribution to their understanding and, hence, to support compatible restoration. In particular, the facade decorations were investigated for the appraisal of their materials and technologies, often neglected in current maintenance/restoration works and so cancelled, leading to a severe loss in architectural image. The ongoing diagnostic campaign, in collaboration among different universities, is aimed to set up a database on materials and technologies of Art Nouveau facade decorations at a European scale, as a technical-scientific background for the highlighting of preservation guidelines
The complex process of vitamin A uptake: a first insight by NMR and other biophysical techniques
No full textVitamin A is essential for diverse aspects of life, ranging from embryogenesis to the proper functioning of most adult organs. It circulates in blood bound to serum retinol binding protein (RBP) and is transported into cells by a multitransmembrane receptor termed stimulated by retinoic acid 6 (STRA6)(1). A STRA6-mediated release of retinol from holo-RBP to target organs through a new mechanism has an evolutionary advantage that prevents a possible toxicity deriving from an excessive accumulation of free vitamin A (2). There is also evidence that a specific binding site for the apo-form of the cellular carriers (CRBP) might exist on the cytoplasmic side of the membrane (3).
To gain a first insight into this complex process, we have investigated the effects of biomembrane mimetic systems on CRBP-I and CRBP-II, by means of NMR, Circular Dichroism and Fluorescence measurements.
The interactions of the two homologous proteins with model membranes were studied by recording 15N-HSQC and 15N-TROSY spectra at different molar ratios. Chemical shifts perturbations and line shape analysis provided insights into the interacting residues and proteins conformational dynamics. As the signals were broadened beyond detection at latest steps of the titration, the NMR data have been complemented by CD and Fuorescence measurements.
The results revealed striking differences between the two homologs (4), despite having nearly identical backbone structures (a beta-barrel with two short alpha-helices) and identical retinol-binding motifs. Apo-CRBP-I interacts with model membranes and very likely the helical domain participates in the formation of a protein-membrane “collisional complex” that results in structural changes of the retinol entry site and in a slower protein tumbling. In contrast, NMR data reveal a higher structural flexibility of apo-CRBP-II that allows the existence of more than one conformation. Line shapes analysis performed in the course of the titrations indicated that the protein conformational dispersion is reduced to one preferred state in the presence of phospholipids bilayers. CD spectra showed that the overall structural integrity of the protein is not affected by the presence of the liposomes.
These new evidences complement our earlier results, which had suggested that the two primary cellular retinol carriers exhibit different mechanisms of ligand uptake (5, 6); combined with their distinct tissue distribution this may imply different roles in an intracellular context.
References:
1. Kawaguchi R., Yu J., Honda J., Hu J., Whitelegge J., Ping P., Wiita P., Bok D. and H. Sun Science 315, 820-825 (2007).
2. Kawaguchi R., Yu J., Ter-Stepanian M., Zhong M., Cheng G., Yuan Q., Jin M., Travis G.H., Ong D. and Sun H. ACS Chemical Biology 6, 1041-1051 (2011).
3. Redondo C., Vouropoulou M., Evans J. and Findlay J.B.C. The FASEB J. 22, 1043-1054 (2008).
4. Franzoni L., Baroni F., Cavazzini D., Rossi G.L. and Lücke C., in preparation.
5. Mittag T., Franzoni L., Cavazzini D., Schaffhausen B., Rossi G.L. and Günther U.L. J. Am. Chem. Soc. 128, 9844-9848 (2006).
6. Franzoni L., Cavazzini D., Rossi G.L. and Lücke C. J. Lipid Res. 51, 1332-1343 (2010).
Acknowledgements: the EU-NMR infrastructure HWB●NMR @ Birmingham (UK) is acknowledged for providing access to instrumentation
Biomolecular NMR, a versatile tool for the understanding of protein science: retinoid-binding proteins as an example
No full textProtein science stands at the heart of modern life sciences because it unravels fundamental biological mechanisms and forms the basis for rapid advances in biomedicine and biotechnology. NMR spectroscopy is uniquely suited to study various aspects of protein structure, dynamics, molecular interactions and function, because information for individual residues can be obtained; moreover, kinetic data, low-populated states and the possible formation of intermediates on the reaction pathway can be determined.
The case of retinoid-binding proteins is discussed here, as an example. Vitamin A has diverse biological functions and is essential for human survival. It circulates in blood bound to serum retinol binding protein (RBP) and is transported into cells by a membrane receptor termed STRA6 (1). The cellular trafficking and metabolism of vitamin A are regulated primarily by specific high-affinity carriers called CRBP-I and CRBP-II. They represent an interesting case where structure determination as well as the study of fast dynamics (ps-ns time scale) (2) failed to elucidate the mode of retinol binding and thus to explain their diverse tissue distribution, functional roles and different ligand affinities.
The highly similar structure of the apo and holo forms (a beta-barrel with two short alpha-helices, see the cartoon) exhibits a closed conformation in both proteins, that seemingly offer no access for the ligand. Given the biological relevance of retinoids, the characterization of their protein interactions and targeted release is of special interest. To tackle this challenging subject we have employed a suite of NMR techniques: 15N relaxation dispersion experiments to investigate the proteins dynamics in the slower micros-ms timescale, line-shape analysis of 15N-HSQC spectra recorded during a retinol titration to get insights into the mechanism of ligand binding and H/D exchange experiments to investigate conformational stability. The results allowed to derive a model of retinol uptake, which is different for CRBP-I and CRBP-II (3, 4); moreover, a distinct local flexibility was found to modulate their binding properties (5).
The two proteins deliver retinol to microsomal membrane-bound enzymes, either for esterification with fatty acids (LRAT) (6, 7) or for oxidation to retinaldehyde (RDH) (8). Our current understanding of these processes remains incomplete, but there is evidence that the membrane microenvironment plays a role in the interactions of holo CRBPs with enzymes (8). To address this hypothesis, we have performed a series of NMR experiments with retinol-bound CRBP-I and CRBP-II in the presence of model membranes composed of either anionic or zwitterionic phospholipids, at varying protein:lipid molar ratios and ionic strength.
Both homologues interact with liposomes of anionic phospholipids, but in a significantly different way (9). A conformational rearrangement of the portal region, coupled to a change in protein dynamics, are required for retinol exchange; these processes seem to be triggered by a membrane-collision.
All the differences between CRBP-I and CRBP-II, when dissolved either in buffer or in the presence of biomembrane mimetic systems, may account for their distinct functional roles in the modulation of intracellular retinoid metabolism. Further experiments are in progress to better describe the ongoing processes in a biological context.
(1) Kawaguchi R., Yu J., Honda J., Hu J., Whitelegge J., Ping P., Wiita P., Bok D., Sun H. (2007) Science, 315, 820-825.
(2) Franzoni L., Lücke C., Pérez C., Cavazzini D., Rademacher M., Ludwig C., Spisni A., Rossi G.L., Rüterjans H. (2002) J. Biol. Chem., 277, 21983-21997.
(3) Mittag T., Franzoni L., Cavazzini D., Schaffhausen B., Rossi G.L., Günther U.L. (2006) J. Am. Chem. Soc., 128, 9844-9848.
(4) Franzoni L., Reed M., Cavazzini D., Rossi G.L., Günther U.L., in preparation.
(5) Franzoni L., Cavazzini D., Rossi G.L., Lücke C. (2010) J. Lipid Res., 51, 1332-1343.
(6) Amengual J., Golczak M., Palczewski K., von Lintig J. (2012) J. Biol. Chem., 287, 24216-24227.
(7) Jiang W., Napoli J.L. (2012) Biochim. Biophys. Acta, 1820, 859-869.
(8) Napoli J.L. (2012) Biochim. Biophys. Acta, 1821, 152-167.
(9) Franzoni L., Baroni F., Cavazzini D., Rossi G.L., Lücke C., in preparation
Measurement of total oxyradical scavenging capacity as an useful tool for understanding susceptibility to oxidative stress conditions
No full text
The complex processes of cellular vitamin A uptake and delivery: insights by NMR and other biophysical techniques
No full textVitamin A has diverse biological functions and is essential for human survival. It circulates in blood bound to serum retinol binding protein (RBP) and is transported into cells by a membrane receptor termed STRA6. The cellular trafficking and metabolism of vitamin A are regulated primarily by specific high-affinity carriers called CRBP-I and CRBP-II. Both proteins deliver retinol to membrane-bound enzymes, either for esterification with fatty acids or for oxidation to retinaldehyde. Until now it was not clear whether CRBPs may couple directly to STRA6 and to the enzymes, or the biomembranes modulate ligand uptake and delivery.
To gain insights into these complex processes, we have investigated the interactions of CRBPs (in the apo and holo forms) with biomembrane mimetic systems. NMR experiments were performed at different protein:vesicles molar ratios and by varying the composition of the phospholipid liposomes and the ionic strength. Chemical shifts perturbations and line shape analysis provided information on the interacting residues and proteins conformational dynamics. As the signals were broadened beyond detection at latest steps of the titrations, the NMR data have been complemented by other biophysical measurements.
The results revealed striking differences between CRBP-I and CRBP-II, despite they exhibit the same fold (a beta-barrel with two short alpha-helices) and identical retinol-binding motifs. Moreover, the interactions of the two homologs with the lipid bilayers depend upon the phospholipid composition and ionic strength.
The new evidences complement our previous studies which suggested that the two primary cellular retinol carriers adopt different mechanisms for ligand uptake [1, 2]. These differences may account for their distinct functional roles in the modulation of retinoid metabolism.
[1] T. Mittag, L. Franzoni, D. Cavazzini, B. Schaffhausen, G.L. Rossi, U.L. Günther J. Am. Chem. Soc. 128, 9844-9848 (2006).
[2] L. Franzoni, D. Cavazzini, G.L. Rossi, C. Lücke J. Lipid Res., 51, 1332-1343 (2010)
Sulle tracce di Attis ad Aquileia
No full textThe aim of this article is to highlight the issues concerning the interpretation of documents referring to Attis and his cult in Aquileia. The author focuses on two of the main problems in the study of archaeological documentation: the over-reading of the image and its function. The analysis of the iconographical components of the figure of Attis shows the extreme versatility of the image of the oriental sheperd and highlights how complex it can be to assign a name to that image without any advice provided by the context. Not each oriental sheperd does represent Attis; more over, an image of Attis does not suffice to confirm the practice of his cult in one site
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