307,361 research outputs found
EFFECT OF DIVALENT CATIONS ON PORIN INCORPORATION IN PLANAR BLM
No full textBiological membranes contain as a basic structural unit a phospholipid bilayer.
Phospholipid membranes in the form of bilayer films (BLM) are currently used as experimental models for transport phenomena of the biological membranes. In particular, bilayers made of acidic phospholipids, present the advantage to investigate the bioelectrical phenomena across the biological membranes.
This class of phospholipids by having a fixed charge might bind ions that play an important role on many physiological processes. It is known that Ca++ and other alkaline cations change the electrostatic potential of the negatively charged bilayer membranes either absorbing to the phospholipids or accumulating in the aqueous diffuse double layer, and this potential change may induce conductance variation (1).
Cd++ another divalent cation has been shown to exert influence on the permeability of some anaelectrolytes across BLM made of negatively charged phospholipids, but not on the zwitterionic phospholipids (2).
We have studied the effect of divalent cations as Ca++ and Ba++ that exert a cross linking together with a screening effect respectively, on the kinetics of incorporation of the mithocondrial porin in bimolecular lipid membranes made with phosphatydilinositol.
In order to understand how lipids of the bilayer affect protein incorporation at the molecular level we must consider that protein crossing the bilayer has to surmount different regions characterized by different properties as interfacial region, head groups, lipid backbone and hydrocarbon core.
In this study we focused our attention to the role of the interface and head group regions of the BLM in the porin incorporation.
The artificial membranes were made of phosphatydilinositol cromatografically pure in n-decane (1% w/w). The bathing media were KC1 1M and KC1 1M plus CaC12 or BaCl2 10 mM. Porin was added at membrane "black" on the two aqueous bathing solutions. Bilayers conductivity and capacitance measurements were carried out at temperature of 25 ± 2 °C and studied by means of alternate current (f = 1 Hz) recording the voltage after a current to voltage converter in series with the membrane; simultaneously electrical capacitance was measured at 1 KHz before and during porin incorporation (3) .
We noticed that when the membranes are in the presence of Ca++ or Ba++ there was an exceptional stability.
With the different bathing solutions no significant differences were observed in membrane conductance values whereas small capacitance variations were recorded.
Once the membrane has became black porin, at a fixed concentration, was added to the aqueous solutions, and after a lag time due to the diffusion of the protein in the solutions and of the time to overcome the membrane barriers, an increase of the voltage output due to the channels formation was recoded. This phenomenon shows a "S-shaped" kinetics. In contrast to the kinetics of incorporation of the porin in oxidized cholesterol lipid membranes (4), showing an hyperbolic shape. These different kinetics may be explained by the different interface barrier; in fact when the experiment were conducted with porin already present in the bathing solutions before membrane formation, the different kinetics persisted in the phosphatydilinositol and in oxidized cholesterol BLM.
The S shaped kinetics persists also when porin is incorporated in phosphatydilinositol membranes in the presence of Ca++ or Ba++; but the curve in presence of the Ca++ is shifted to the right.
This shift is much more pronounced in the presence of Ba++.
It is known that Ca++ affects the surface potential of BLM containing negatively charged lipids. The protein incorporation into BLM can be regarded as a two steps process: I) the interaction at the surface barrier, and II) the penetration into hydrocarbon region.
In the first step the protein encounter an higher resistance in phosphatydilinositol membrane because it has to overcome the interface region consisting of a diffuse double layer, the membrane surface and the polar head groups.
Ca++, by exerting a cross-linking and screening effect on negatively charged membranes, retards the porin incorporation.
At the moment we are not able to distinguish which of the two effect is prevalent; but results in presence of Ba++, that exerts a screening effect only seem to indicate that the latter effect is prevalent. In order to establish the pure cross-linking effect experiments are in progress.
The influence of the divalent cations on the protein incorporation into BLM, that are sensitive to changes in environmental conditions, make this study appealing in order to provide a better knowledge in the molecular mechanism of the protein incorporation and channel formation.
Work supported by 40% grant from MURST 1991
1) S. G. A. McLaughlin, G. Szabo and G. Eisenman, 1971: J. Gen. physiol., 58, 667.
2) S. Micelli, E. Gallucci and C. Lippe, 1978: Arch. Intern. Physiol. Biochim., 86, 755-759.
3) G. Monticelli, E. Gallucci and S. Micelli, 1990: Proc. X School on Biophysics of Membrane Transport, 1: 328-343, Poland.
4) S. Micelli, G. Monticelli and E. Gallucci, 1990: IBST 1: 221
La canzone tassiana alla Vergine di Loreto. Per una nuova datazione, a partire da uno studio del ms. P1
No full textL’articolo propone una nuova datazione per la canzone tassiana alla Vergine di Loreto. Il componimento, scritto nel 1587, è stato a lungo considerato frutto di un secondo rimaneggiamento. Le motivazioni sono da ricercare nell’VIII stanza dov’è presente il riferimento a un pontefice identificato con Niccolò Sfrondati, Gregorio XIV. Non ci sono, tuttavia, prove materiali di un’iniziale assenza della stanza e la sua introduzione, a seguito dell’elezione avvenuta il 5 dicembre 1590, è in contrasto con i testimoni che la trasmettono.
Lo studio vuole dimostrare come sia difficile postdatare la stampa Berichia del 1589, ma soprattutto analizza il ms. Banco Rari 212 (P1) per il quale vale lo stesso termine ante quem. Fissati gli estremi cronologici si avanza un’ipotesi risolutiva: pensare, sulla base di elementi biografici e intertestuali, che Tasso si stia riferendo a Ugo Boncompagni, Gregorio XIII, permetterebbe di riordinare la storia della canzone compatibilmente con quella dei testimoni
Kinetic parameters of porin incorporation into black lipid membranes of oxidized cholesterol evaluated by means of alternating current
No full textIn this work we report the results of systematical investigation of the time course of porin incorporation into bilayer membranes made of oxidized cholesterol using an ac method (E. Gallucci et al., Biophys. J. 71: 824-831, 1996). This method has the advantages of continuously monitoring resistance and capacitance as a function of time during pore formation until a steady state of the phenomenon is reached. In our case the bilayer membranes are represented, as usual, by an equivalent electrical circuit consisting of one resistance and one capacitance connected in parallel. The total current passing through the membranes is easily monitored, during the insertion process and is interpreted as a probe of the process involved during channel incorporation and assembly. By means of a simple mathematical model, we provide evidence that two concurrent processes are present during the incorporation time course which can be interpreted either as insertion/deinsertion or as positive/negative cooperativity. We observe a phase transition (or similae phenomenon) which seems to take place during the insrtion process, an aspect which has been neglected in most previous investigation of insertion into bilayer membranes. Moreover, the continuous monitoring of capacitance may prove useful in tracking membrane parameters, such as thickness and dielectrical constant, that depend on the lipid and incorporated proteins
Un nuovo sottosistema penale
No full textin BALDI F., GALLUCCI E., GARUTI G., MATTEVI E., PANIZZO F., PICOTTI L., PONGILUPPI C., Le definizioni alternative del processo penale davanti al giudice di pace. Conciliazione, irrilevanza del fatto e condotte riparatori
PORIN INCORPORATION KINETICS IN PHOSPHATIDYLINOSITOL AND OXIDIZED CHOLESTEROL MEMBRANES
No full textConductivity of planar lipid bilayer membranes can be modulated by insertion of protein molecules (1).
Mithocondrial porins form water filled pores in the outer membrane and these channels act selectively on the permeation of hydrophilic solutes.
Aims of this research were: 1) the reconstitution of porin channels in planar lipid bilayer membranes, 2) the analysis and the comparison of porin incorporation kinetics in different lipid membranes: phosphatidylinositol (PI) and oxidized cholesterol, 3) the study of protein concentration effect on the incorporation kinetics, 4) the analysis of the channel reconstitution dependence on the applied voltage.
The artificial membranes were formed, with lipid in n-decane (1% w/w), brushing the solution on a small circular hole between two compartments, 4 ml volume, of a teflon experimental chamber. The two compartments were filled with KCl 1M solution and continuously stirred.
Pt electrodes in bathing solutions were connected to a current to voltage converter and to a sinusoidal voltage source (Vs=VS sin2nt; n=1Hz) so that the voltage at the converter (Vl) resulted directly related to the electrical current crossing the membrane (2).
Experiments were performed at room temperature: 24 ± 1 °C.
Membrane electrical conductances and capacitances were higher in oxidized cholesterol than in PI membranes.
Addition of porin to the solutions bathing a completely black membrane induced an increase of membrane current with time indicating protein fusion and channel formation into the lipid matrix.
In PI membranes the time course of current was S-shaped, irrespective of porin addition before or after membrane formation, but in the first case a delay in channel formation and a slower kinetics were observed (3).
In oxidized cholesterol membranes the current time behaviours were S-shaped when porin was added after membrane formation but resulted of exponential type when porin was present before membrane formation. In this second case faster kinetics were observed and the incorporation started with an higher conductance value.
The time lag between porin addition and the first current variation was shorter in oxidized cholesterol than in PI membranes. For a same porin concentration the incorporation kinetics were faster and higher steady levels were reached in oxidized cholesterol than in PI membranes.
In both types of membranes porin incorporation rate decreased lowering porin concentration; five kinetics of porin incorporation (protein added after membrane formation) in oxidized cholesterol membranes are reported in figure.
Besides the porin concentration also the applied voltage affects the incorporation kinetics and the channel formation.
At a given porin concentration faster kinetics are recorded and higher steady levels are reached increasing the voltage to a critical value after which incorporation is less effective, the kinetics are slower and accompanied by lower steady levels.
The voltage dependence of functional pore reconstitution was more evident in PI than in oxidized cholesterol membranes.
In oxidized cholesterol membrane, at the steady state, nonlinear current-voltage characteristics were obtained. The conductance increases with the voltage to reach a critical value after which inactivation occurs.
At a given applied voltage higher membrane conductances were calculated increasing porin concentration.
Significative differences in the current-voltage relations were obtained depending on the absence or the presence of porin at the membrane formation. In this latter case the membranes had higher conductance values but, at the tested voltages, the range of variability was smaller.
The nature of the lipid matrix can be considered responsible of the different protein incorporation; in particular the surface charge of the lipid bilayer membrane and the interface potential, due to the local accumulation of ions at the membrane-solution interface, play a key role in the incorporation of porin in PI planar bilayer lipid membranes.
These observations suggest the possibility to insert protein channels in lipid layers built on a solid substrate finalized to realize organic semiconductors to be used as biosensor or transducers.
Work supported in part by CNR grant 92.02202.CT14.
1) G. Monticelli - "Pore formation in planar lipid bilayer membranes" Proc. X School on Biophysics of Membrane Transport, Szczyrk (Poland) 1990, 1: 315-327.
2) G. Monticelli, E. Gallucci and S. Micelli - "Experimental data on incorporation of porin molecules in lipid bilayers" Proc. X School on Biophysics of Membrane Transport, Szczyrk (Poland) 1990, 1: 328-343.
3) G. Monticelli, E. Gallucci and S. Micelli - "Pore formation in lipid membranes" Proc. Int. Cong. Memb. and Memb. Processes, Chicago 1990, 1: 175-177
Rehabilitating patients with left spatial neglect by prism exposure during a visuomotor activity
No full textFortis P, Maravita A, Gallucci M, et al. Rehabilitating patients with left spatial neglect by prism exposure during a visuomotor activity. Neuropsychology. 2010;24(6):681-697
Close reading e codifica interpretativa: edizione digitale di libretti d'opera
No full textThis paper reviews the preparatory and operational stages that led to the development of the digital scientific edition of 57 musical dramas staged in the Republic of Lucca between 1636 and 1705 to celebrate an important political event. This case study prompts a reflection on the relationship between digital humanities and opera theater by providing a detailed state of the art: the increased interest in databases and the significant number of amateurs of the genre explain both the detected distance from the textual data and the poor online availability of philologically curated libretti. Instead, the present edition focuses on the content of the operas in terms of textual typologies, use of sources and characters and, in particular, political ideas. In describing the XML-TEI encoding scheme, this paper aims to show interpretive close reading can enhance the analysis of certain concepts, political in this case, through markup. Although acknowledging some limitations, such as the risk of overlapping and the lack of unambiguous IDs, the results highlight a new way of conceiving the commentary of digital editions which provides scholars with the possibility of using digital tools to annotate and visualize exegetical reading.Il contributo ripercorre le fasi riflessive e operative che hanno condotto all’allestimento di un’edizione scientifica digitale di 57 drammi per musica messi in scena nella Repubblica di Lucca tra il 1636 e il 1705 in occasione di un importante momento politico e celebrativo. Il caso specifico permette di riflettere sulla relazione tra digital humanities e teatro d’opera fornendo un dettagliato stato dell’arte: il maggiore interesse per i database e il cospicuo numero di amatori del genere spiegano sia il riscontrato allontanamento dal dato testuale sia la scarsa presenza in rete di libretti curati in termini filologici. La presente edizione, invece, pone al centro dell’attenzione il contenuto delle opere approfondendo la natura dei testi, l’uso delle fonti e dei personaggi, ma soprattutto delle idee politiche. Nel descrivere lo schema di codifica XML-TEI, si avanza la proposta di un close reading interpretativo che valorizzi l’analisi di determinati concetti, politici in questo caso, attraverso la marcatura. I risultati evidenziano un nuovo modo di concepire il commento alle edizioni digitali che, pur riscontrando alcuni limiti come il rischio di overlapping e la mancanza di identificativi univoci, fornisce al critico la possibilità di ricorrere agli strumenti digitali per annotare e visualizzare una lettura esegetica
The role of external applied voltage on kinetic parameters of porin incorporation into black lipid membranes of oxidized cholesterol
No full textThe time course of porin incorporation into black lipid membranes of oxidized cholesterol, as a function of on external applied voltage, was studied by means of an ac method previously reported (E. Gallucci et al., Biophys. J. 71: 824-831, 1996). The kinetic parameters was derived by means of a simple mathematical model before described (D. Meleleo et al., SIF, 1998, Р 53).
At the different voltages Vs examined, the kinetics remained unmodified in its cooperative behavior. The K1 or growth rate parameter describing porin insertion or positive cooperativity seems to be independent of the applied voltage. The K2 or growth rate parameter related to porin deinsertion or negative cooperativity is strongly dependent on applied voltage. In particular K2 decreases with increasing applied voltages (Vs). The v(tm) or value of the maximum depolarization rate, and the Vll(tm) or value of the output voltage when the derivative v(tm) reaches the maximum, both increase with applied voltage (Vs). The kinetic values found indicated that mitochondrial porin is not driven by potential in the membrane, but its insertion depends (for fixed porin concentration, temperature and pH) on the lipid-protein affinity. This result supports previous authors' findings of porin insertion into mitochondrial membranes and asymmetric lipid bilayers (H. Freitag et al., Eur.J. Biochem. 74:483-492, 1993; S.M. Gasser et al.,J. Biol. Chem. 25: 3427- 3430,1983, A. Viese et al., Biophys. J., 70:321-329,1996). On the other hand, as K2 was found to be related with the conductance at the end of porin incorporation, its correlation with Vs reflects the voltage- dependence characteristics of the porin channel
Quality oriented services marketing for eno-gastronomy areas. Findings from a survey of local actors in the Parco Nazionale del Cilento e Vallo di Diano.
No full textThe aim of the study is to investigate the issue of Quality (Gronroos, 2002, Rust, Oliver,
1994) in eno-gastronomy tourism areas from a competitive advantage perspective
(Gallucci, Marino, Mainolfi, 2008). Our research, an exploratory study involves the key
actors (agro-tourism farms, manufacturers/producers of typical products and
institutional stakeholders) in territories characterized by a strong eno-gastronomy
vocation such as the Parco Nazionale del Cilento. We argue that lasting and
sustainable competitive advantage in a territory with a tradition of eno-gastronomy,
depends on the capacity both of local governing boards to manage complex and
intricate functions of integration and coordination, and at the same time, of individual
operators to comply with the standards of quality delineated in a theoretical model
framework. In other words, to package experiential tourism products designed by actors
inside and outside a specific destination, guided by market driven logics. A further aim
is to identify specific actions (best practices) for cooperation at destination level (value
design) and in the value co-creation phase (Pencarelli, Splendiani, 2008). In particular,
from a dual perspective, on the one hand, value created “for” and “with” tourists of
eno-gastronomy areas, beneficiaries and creators of the tourist experience, by
activating a mix of tools to spread tangible and intangible benefits (psychological,
social, emotional) and to minimize costs deriving from client-relation activities, andon the other, “for” the area, i.e. rejecting logics pivoted on mere conservation and protection of resources and based rather, on the modern logics of enhancing destination
heritage (Della Corte, 2009).
KEY WORDS Quality | Destination Management | Tourism Marketing
Evidence for a different assembly of Gramicidin A in oxidized cholesterol black lipid membrane
No full textGramicidin A (GA) is a hydrophobic linear pentadocapeptide that forms cation- selective channels in natural and black lipid membranes (BLM). The channels are believed to consist of single-stranded helices with the NH2 terminals linked together by hydrogen bonds. Linear gramicidins require a lipid bilayer to fold properly. If the phospholipid acyl chains are less than eight carbons long, channel folding is not observed; if the acyl chains are too long, the membrane- spanning channels are destabilized. Besides, embedded proteins might influence the molecular packing lipids (R. E. Koeppe II et al. Biol. Skr. Dan. Selsk, 49-93-98, 1998). In our experiments, we chose oxidized cholesterol black lipid membranes because their thickness (40±10 A) (Ti Tien and A. L. Diana, In Chemistry and Physics of Lipids, 2, 1968) is suited to both gramicidin length (25-30 A) and folding. The time course of gramicidin incorporation into BLM, studied by means of alternating current (E. Gallucci et al., Biophys. J. 71:824- 831, 1986), was sigmoidal independently of the KCI concentration used, indicating a cooperative process of incorporation into BLM. By means of a simple mathematical model (S. Micelli et al, submitted) we estimated the tm value, i.e. the parameter that describes a phase transition process (or similar phenomenon). This phenomenon has been observed by AA as a function of temperature and GA concentration (J. A. Killian and B. De Kruijff, Biophys. 1. 53: 111-117, 1988; Mingtao Ge and J. H. Freed, Biophys. J., 76: 266-280, 1999). On the other hand, the average values of the steady-state conductance against gramicidin concentration, for different KCI concentrations, shows a sigmoidal behavior too. These curves were fitted by means of a four-parameter equation which allowed us to obtain the Hill coefficient, D, or slope factor. The Hill coefficient gives the number of single-stranded helices which form the channel. Our results suggest that the number of channel sub-units depends on the KCI concentration. In our case, at a KCI concentration of 1M, the channel is characterized by two sub-units, in accordance with AA's finding (D. U. Urry, Proc. Natl. Acad. Sci. USA, 86:672, 1971, W. R. Veatch and R. Blond, Biochemisty, 13:5249, 1974), while for KCI concentrations of 0.1 and 0.5 M. the channel is formed by more than two sub-units
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