49 research outputs found
Effect of ionic strength on intra-protein electron transfer reactions: The case study of charge recombination within the bacterial reaction center
It is a common believe that intra-protein electron transfer (ET) involving reactants and products that are overall electroneutral are not influenced by the ions of the surrounding solution. The results presented here show an electrostatic coupling between the ionic atmosphere surrounding a membrane protein (the reaction center (RC) from the photosynthetic bacterium Rhodobacter sphaeroides) and two very different intra-protein ET processes taking place within it. Specifically we have studied the effect of salt concentration on: i) the kinetics of the charge recombination between the reduced primary quinone acceptor QA- and the primary photoxidized donor P+; ii) the thermodynamic equilibrium (QA- â QB-) for the ET between QA- and the secondary quinone acceptor QB. A distinctive point of this investigation is that reactants and products are overall electroneutral. The protein electrostatics has been described adopting the lowest level of complexity sufficient to grasp the experimental phenomenology and the impact of salt on the relative free energy level of reactants and products has been evaluated according to suitable thermodynamic cycles. The ionic strength effect was found to be independent on the ion nature for P+ QA- charge recombination where the leading electrostatic term was the dipole moment. In the case of the QA- â QB- equilibrium, the relative stability of QA- and QB- was found to depend on the salt concentration in a fashion that is different for chaotropic and kosmotropic ions. In such a case both dipole moment and quadrupole moments of the RC must be considered
Sensitive biosensors exploiting the minute changes in the capacitance of protein layers associated to the ligand recognition
Soft matter systems interfaced to an electronic device are presently one of the most challenging research activity that has relevance not only for fundamental studies but also for the development of highly performing bio-sensors. Layers of proteins anchored on solid surfaces have small capacitance that undergoes to only minute changes as the ligandâprotein complex is formed. For properly designed systems, the protein layer represents smallest capacitance in a series of capacitors and as such dominates the overall capacitance. When such a protein layer is integrated in a Field Effect Transistor (FET) transduction is remarkably sensitive as the transistor output current is governed by the small changes due to ligand binding. These devices operate in aqueous solutions and are promising as portable sensors for point- of-care applications Two recent achievements will be illustrated: A) the sensitive and quantitative measurement of the weak interactions associated with the binding of neutral enantiomers to Odorant Binding Proteins (OBPs) [1]. immobilized to the gate of a bio-FET. Here the minute change in protein layer capacitance upon binding of S(-)-carvone and R(+)-carvone modulate the response of a water-gated OFET, allowing for chiral differential detection. The FET binding curves modelling provide information on the electrochemical free energies derived from the FET dissociation constants while the electrostatic component is isolated from the threshold voltage shifts. These can be combined with the chemical free energies gathered from the complex formation in solution, overall providing a comprehensive picture of the energy balances for a surface-bound pOBP-carvone complex undergoing chiral interactions. B) Hierarchically organized layers of phospholipids and proteins anchored on the surface of the semiconductor and acting as selective recognition elements independently form the solution ionic strength [2-3]. The charged moieties of the bound proteins along with the counter-ions form a layer that is analogous to an ionic gel. The fixed polyelectrolyte ions generate an electric field that confines the mobile counter-ions in the region of the fixed charges. Eventually a Donnanâs equilibrium is reached and the smallest capacitance in series is associated to the Donnanâs electrical double layer. The molecular recognition process (antigen/antibody in the present case) modify the charge density of the outermost layer and thus its capacitance. This capacitive tuning of the bio-FET response is virtually insensitive to the Debyeâs length value and therefore is compatible with use of the transistor as sensor directly in biological fluids at high ionic strength . [1] M.Y. Mulla, E. Tuccori, M. Magliulo, G. Lattanzi, G. Palazzo, K. Persaud, L Torsi Capacitance-modulated transistor detects odorant binding protein chiral interactions Nat. Commun. 2015, 6, 6010 doi: 10.1038/ncomms7010 [2] M. Magliulo, A. Mallardi, M. Yusuf Mulla, S. Cotrone, B.R. Pistillo, P. Favia, I. Vikholm-Lundin, G. Palazzo, L Torsi Electrolyte-Gated Organic Field-Effect Transistor Sensors Based on Supported Biotinylated Phospholipid Bilayer Adv. Mater. 2013, 25, 2090â2094 DOI: 10.1002/adma.201203587 [3]G. Palazzo, D. De Tullio, M. Magliulo, A. Mallardi, F. Intranovo, M.Y. Mulla, P. Favia, I. Vikholm-Lundin, L. Torsi Detection beyond the Debyeâs length with an electrolyte gated organic field-effect transistor Adv. Mater. 2015, 27, 911-916. DOI: 10.1002/adma.2014035
Soft matter films interfaced to electronic devices: capacitance-modulated field effect transistors integrating protein layers
Soft matter systems interfaced to an electronic device are presently one of the most challenging research activity that has relevance not only for fundamental studies but also for the development of highly performing bio-sensors.
Layers of proteins anchored on solid surfaces have small capacitance that undergoes to only minute changes as the ligandâprotein complex is formed.
For properly designed systems, the protein layer represents smallest capacitance in a series of capacitors and as such dominates the overall capacitance. When such a protein layer is integrated in a Field Effect Transistor (FET) transduction is remarkably sensitive as the transistor output current is governed by the small changes due to ligand binding. These devices operate in aqueous solutions and are promising as portable sensors for point-of-care applications
Two recent achievements will be illustrated:
A) the sensitive and quantitative measurement of the weak interactions associated with the binding of neutral enantiomers to Odorant Binding Proteins (OBPs) [1]. immobilized to the gate of a bio-FET. Here the minute change in protein layer capacitance upon binding of S(-)-carvone and R(+)-carvone modulate the response of a water-gated OFET, allowing for chiral differential detection. The FET binding curves modelling provide information on the electrochemical free energies derived from the FET dissociation constants while the electrostatic component is isolated from the threshold voltage shifts. These can be combined with the chemical free energies gathered from the complex formation in solution, overall providing a comprehensive picture of the energy balances for a surface-bound pOBP-carvone complex undergoing chiral interactions.
B) Hierarchically organized layers of phospholipids and proteins anchored on the surface of the semiconductor and acting as selective recognition elements independently form the solution ionic strength [2-3].
The charged moieties of the bound proteins along with the counter-ions form a layer that is analogous to an ionic gel. The fixed polyelectrolyte ions generate an electric field that confines the mobile counter-ions in the region of the fixed charges. Eventually a Donnanâs equilibrium is reached and the smallest capacitance in series is associated to the Donnanâs electrical double layer. The molecular recognition process (antigen/antibody in the present case) modify the charge density of the outermost layer and thus its capacitance.
This capacitive tuning of the bio-FET response is virtually insensitive to the Debyeâs length value and therefore is compatible with use of the transistor as sensor directly in biological fluids at high ionic strength .
[1] M.Y. Mulla, E. Tuccori, M. Magliulo, G. Lattanzi, G. Palazzo, K. Persaud, L Torsi
Capacitance-modulated transistor detects odorant binding protein chiral interactions
Nat. Commun. 2015, 6, 6010 doi: 10.1038/ncomms7010
[2] M. Magliulo, A. Mallardi, M. Yusuf Mulla, S. Cotrone, B.R. Pistillo, P. Favia, I. Vikholm-Lundin, G. Palazzo, L Torsi
Electrolyte-Gated Organic Field-Effect Transistor Sensors Based on Supported Biotinylated Phospholipid Bilayer
Adv. Mater. 2013, 25, 2090â2094 DOI: 10.1002/adma.201203587
[3] G. Palazzo, D. De Tullio, M. Magliulo, A. Mallardi, F. Intranuovo, M.Y. Mulla, P. Favia, I. Vikholm-Lundin, L. Torsi
Detection beyond the Debyeâs length with an electrolyte gated organic field-effect transistor
Adv. Mater. 2015, 27, 911-916. DOI: 10.1002/adma.201403541
A general approach to the encapsulation of glycoenzymes chains inside calcium alginate gel beads
In this work an enzyme encapsulation general approach, based on the use of calcium alginate hydrogels, is reported. Alginate gels are biodegradable and low cost and have been found to provide a good matrix for the entrapment of sensitive biomolecules. Alginate is an anionic polymer whose gelation occurs by an exchange of sodium ions from the polymer chains with multivalent cations, resulting in the formation of a three dimensional gel network. For gelation alginate is dripped into a calcium chloride solution. The cations diffuse from the continuous phase to the interior of the alginate droplets and form a gelled matrix. By means of this âexternal gelation methodâ beads with a diameter of few millimeters can be obtained (see figure 1).
The entrapment of enzymes in alginate beads suffers some disadvantages, like as low enzyme loading efficiency with reduction of the immobilization yields and reusability, related to the enzyme leakage from the large beads pores (cut off of about 100 kDa).
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Probing light-induced conformational transitions in bacterial photosynthetic reaction centers embedded in trehaloseâwater amorphous matrices
AbstractThe coupling between electron transfer and protein dynamics has been studied in photosynthetic reaction centers (RC) from Rhodobacter sphaeroides by embedding the protein into room temperature solid trehaloseâwater matrices. Electron transfer kinetics from the primary quinone acceptor (QAâ) to the photoxidized donor (P+) were measured as a function of the duration of photoexcitation from 20 ns (laser flash) to more than 1 min. Decreasing the water content of the matrix down to â5Ă103 water molecules per RC causes a reversible four-times acceleration of P+QAâ recombination after the laser pulse. By comparing the broadly distributed kinetics observed under these conditions with the ones measured in glycerolâwater mixtures at cryogenic temperatures, we conclude that RC relaxation from the dark-adapted to the light-adapted state and thermal fluctuations among conformational substates are hindered in the room temperature matrix over the time scale of tens of milliseconds. When the duration of photoexcitation is increased from a few milliseconds to the second time scale, recombination kinetics of P+QAâ slows down progressively and becomes less distributed, indicating that even in the driest matrices, during continuous illumination, the RC is gaining a limited conformational freedom that results in partial stabilization of P+QAâ. This behavior is consistent with a tight structural and dynamical coupling between the protein surface and the trehaloseâwater matrix
Gold nanoparticles obtained by ns-pulsed laser ablation in liquids (ns-PLAL) are arranged in the form of fractal clusters
Gold nanoparticles (AuNPs), synthesized by ns-pulsed laser ablation in liquid (ns-PLAL) in the absence of any capping agents, are potential model systems to study the interactions with biological structures unencumbered by interference from the presence of stabilizers and capping agents. However, several aspects of the physics behind these AuNPs solutions deserve a detailed investigation. The structure in solution of nsPLAL-synthesized AuNPs was investigated in solution by means of small-angle X-ray scattering (SAXS) and dynamic light scattering (DLS). Furthermore, the (dried) NPs have been examined using TEM. The analysis of the SAXS curve shows the presence of a large number of small aggregates with a fractal structure stabilized by
strong long-range repulsive interactions. Fitting of the SAXS curve to a suitable âfractal modelâ allows the estimation of the features of the fractal including the fractal dimension d = 1.9. The latter allows to estimate the fraction of light scattered by fractals of different sizes and thus permits a fair comparison between the DLS and TEM data. Here, a stable abundant population
of fractal clusters is reported reflecting a mechanism where primary AuNPs (size 7.6 nm) are forced to aggregate forming clusters during the collapse of the cavitation bubble. When these clusters are released in the aqueous phase, their large negative charge builds up repulsive interactions that prevent cluster-cluster aggregation imparting colloidal stability
Effect of the Surface Chemical Composition and of Added Metal Cation Concentration on the Stability of Metal Nanoparticles Synthesized by Pulsed Laser Ablation in Water
Metal nanoparticles (NPs) made of gold, silver, and platinum have been synthesized by means of pulsed laser ablation in liquid aqueous solution. Independently from the metal nature, all NPs have an average diameter of 10 ± 5 nm. The ζ-potential values are: â62 ± 7 mV for gold, â44 ± 2 mV for silver and â58 ± 3 for platinum. XPS analysis demonstrates the absence of metal oxides in the case of gold and silver NPs. In the case of platinum NPs, 22% of the particle surface is ascribed to platinum oxidized species. This points to a marginal role of the metal oxides in building the negative charge that stabilizes these colloidal suspensions. The investigation of the colloidal stability of gold NPs in the presence of metal cations shows these NPs can be destabilized by trace amounts of selected metal ions. The case of Ag+ is paradigmatic since it is able to reduce the NP ζ-potential and to induce coagulation at concentrations as low as 3 ÎŒM, while in the case of K+ the critical coagulation concentration is around 8 mM. It is proposed that such a huge difference in destabilization power between monovalent cations can be accounted for by the difference in the reduction potential