| 1. |
- Dell'Orco, Daniele, et al.
(författare)
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Calcium binding, structural stability and guanylate cyclase activation in GCAP1 variants associated with human cone dystrophy
- 2010
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Ingår i: Cellular and Molecular Life Sciences. - : Springer Science and Business Media LLC. - 1420-9071 .- 1420-682X. ; 67:6, s. 973-984
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Tidskriftsartikel (refereegranskat)abstract
- Guanylate cyclase activating protein 1 (GCAP1) is a neuronal Ca2+ sensor (NCS) that regulates the activation of rod outer segment guanylate cyclases (ROS-GCs) in photoreceptors. In this study, we investigated the Ca2+-induced effects on the conformation and the thermal stability of four GCAP1 variants associated with hereditary human cone dystrophies. Ca2+ binding stabilized the conformation of all the GCAP1 variants independent of myristoylation. The myristoylated wild-type GCAP1 was found to have the highest Ca2+ affinity and thermal stability, whereas all the mutants showed decreased Ca2+ affinity and significantly lower thermal stability in both apo and Ca2+-loaded forms. No apparent cooperativity of Ca2+ binding was detected for any variant. Finally, the nonmyristoylated mutants were still capable of activating ROS-GC1, but the measured cyclase activity was shifted toward high, nonphysiological Ca2+ concentrations. Thus, we conclude that distorted Ca2+-sensor properties could lead to cone dysfunction.
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| 2. |
- Dell'Orco, Daniele, et al.
(författare)
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Delivery success rate of engineered nanoparticles in the presence of the protein corona: a systems-level screening
- 2012
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Ingår i: Nanomedicine: Nanotechnology, Biology and Medicine. - : Elsevier BV. - 1549-9642 .- 1549-9634. ; 8:8, s. 1271-1281
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Tidskriftsartikel (refereegranskat)abstract
- Nanoparticles (NPs) for medical applications are often introduced into the body via intravenous injections, leading to the formation of a protein corona on their surface due to the interaction with blood plasma proteins. Depending on its composition and time evolution, the corona will modify the biological behavior of the particle. For successful delivery and targeting, it is therefore important to assess on a quantitative basis how and to what extent the presence of the corona perturbs the specific interaction of a designed NP with its cellular target. We present a theoretical systems-level analysis, in which peptides have been covalently coupled to the surface of nanoparticles, describing the delivery success rate in varying conditions, with regard to protein composition of the surrounding fluid. Dynamic modeling and parameter sensitivity analysis proved to be useful and computationally affordable tools to aid in the design of NPs with increased success rate probability in a biological context. FROM THE CLINICAL EDITOR: The formation of a protein corona consisting of blood plasma proteins on the surface of intravenously delivered nanoparticles may modify the biological behavior of the particles. This team of investigators present a theoretical systems-level analysis of this important and often neglected phenomenon.
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| 3. |
- Dell'Orco, Daniele, et al.
(författare)
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Dynamics of Conformational Ca2+-Switches in Signaling Networks Detected by a Planar Plasmonic Device
- 2012
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 1520-6882 .- 0003-2700. ; 84:6, s. 2982-2989
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Tidskriftsartikel (refereegranskat)abstract
- Ca2+-sensor proteins regulate a variety of intracellular processes by adopting specific conformations in response to finely tuned changes in Ca2+-concentration. Here we present a surface plasmon resonance (SPR)-based approach, which allows for simultaneous detection of conformational dynamics of four Ca2+-sensor proteins (calmodulin, recoverin, GCAP1, and GCAP2) operating in the vertebrate phototransduction cascade, over variations in Ca2+ concentration in the 0.1-0.6 mu M range. By working at conditions that quantitatively mimic those found in the cell, we show that the method is able to detect subtle differences in the dynamics of each Ca2+-sensor, which appear to be influenced by the presence of free Mg2+ at physiological concentration and by posttranslational modifications such as myristoylation. Comparison between the macroscopic Ca2+-binding constants, directly measured by competition with a chromophoric chelator, and the concerted binding-conformational switch detected by SPR at equilibrium reveals the relative contribution of the conformational change process to the SPR signal. This process appears to be influenced by the presence of other cations that perturb Ca2+-binding and the conformational transition by competing with Ca2+, or by pure electrostatic screening. In conclusion, the approach described here allows a comparative analysis of protein conformational changes occurring under physiologically relevant molecular crowding conditions in ultrathin biosensor layers.
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| 4. |
- Dell'Orco, Daniele, et al.
(författare)
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Mathematical modeling of the protein corona: implications for nanoparticulate delivery systems
- 2014
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Ingår i: Nanomedicine. - 1743-5889. ; 9:6, s. 851-858
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Tidskriftsartikel (refereegranskat)abstract
- This article discusses the role of the protein corona in delivery systems with tagged nanoparticles and how knowledge of the protein corona can help in optimizing delivery. The basic question is whether and how the binding of proteins and other biomolecules at the nanoparticle surface interfere with the interaction between a tag and its receptor. This is an interesting problem in many respects, but most intriguing are the observed differences in delivery efficiency in vivo compared with protein-free in vitro conditions. In order to understand possible situations that the nanoparticle will face in a protein-rich biological environment, we will first describe the formation of a protein corona and thereafter discuss potential perturbations of the delivery systems when moving from in vitro testing to in vivo applications. We emphasize the role of mathematical modeling in optimizing the design of functionalized nanoparticles to achieve high success of delivery.
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| 5. |
- Dell'Orco, Daniele, et al.
(författare)
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Mathematical modeling of the protein corona : Implications for nanoparticulate delivery systems
- 2016. - 2
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Ingår i: Handbook of Immunological Properties of Engineered Nanomaterials: : Haematocompatibility of Engineered Nanomaterials - Haematocompatibility of Engineered Nanomaterials. - : WORLD SCIENTIFIC. - 9789814699167 - 9789814699174 ; 2, s. 53-65
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Bokkapitel (refereegranskat)abstract
- This article discusses the role of the protein corona in delivery systems with tagged nanoparticles and how knowledge of the protein corona can help in optimizing delivery. The basic question is whether and how the binding of proteins and other biomolecules at the nanoparticle surface interfere with the interaction between a tag and its receptor. This is an interesting problem in many respects, but most intriguing are the observed differences in delivery effi ciency in vivo compared with protein-free in vitro conditions. In order to understand possible situations that the nanoparticle will face in a protein-rich biological environment, we will fi rst describe the formation of a protein corona and thereafter discuss potential perturbations of the delivery systems when moving from in vitro testing to in vivo applications. We emphasize the role of mathematical modeling in optimizing the design of functionalized nanoparticles to achieve high success of delivery.
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| 6. |
- Dell'Orco, Daniele, et al.
(författare)
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Modeling the Time Evolution of the Nanoparticle-Protein Corona in a Body Fluid
- 2010
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- Background: Nanoparticles in contact with biological fluids interact with proteins and other biomolecules, thus forming a dynamic corona whose composition varies over time due to continuous protein association and dissociation events. Eventually equilibrium is reached, at which point the continued exchange will not affect the composition of the corona. Results: We developed a simple and effective dynamic model of the nanoparticle protein corona in a body fluid, namely human plasma. The model predicts the time evolution and equilibrium composition of the corona based on affinities, stoichiometries and rate constants. An application to the interaction of human serum albumin, high density lipoprotein (HDL) and fibrinogen with 70 nm N-iso-propylacrylamide/N-tert-butylacrylamide copolymer nanoparticles is presented, including novel experimental data for HDL. Conclusions: The simple model presented here can easily be modified to mimic the interaction of the nanoparticle protein corona with a novel biological fluid or compartment once new data will be available, thus opening novel applications in nanotoxicity and nanomedicine.
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| 7. |
- Dell'Orco, Daniele, et al.
(författare)
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Network-level analysis of light adaptation in rod cells under normal and altered conditions
- 2009
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Ingår i: Molecular BioSystems. - : Royal Society of Chemistry (RSC). - 1742-2051 .- 1742-206X. ; 5:10, s. 1232-1246
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Tidskriftsartikel (refereegranskat)abstract
- Photoreceptor cells finely adjust their sensitivity and electrical response according to changes in light stimuli as a direct consequence of the feedback and regulation mechanisms in the phototransduction cascade. In this study, we employed a systems biology approach to develop a dynamic model of vertebrate rod phototransduction that accounts for the details of the underlying biochemistry. Following a bottom-up strategy, we first reproduced the results of a robust model developed by Hamer et al. (Vis. Neurosci., 2005, 22(4), 417), and then added a number of additional cascade reactions including: (a) explicit reactions to simulate the interaction between the activated effector and the regulator of G-protein signalling (RGS); (b) a reaction for the reformation of the G-protein from separate subunits; (c) a reaction for rhodopsin (R) reconstitution from the association of the opsin apoprotein with the 11-cis-retinal chromophore; (d) reactions for the slow activation of the cascade by opsin. The extended network structure successfully reproduced a number of experimental conditions that were inaccessible to prior models. With a single set of parameters the model was able to predict qualitative and quantitative features of rod photoresponses to light stimuli ranging over five orders of magnitude, in normal and altered conditions, including genetic manipulations of the cascade components. In particular, the model reproduced the salient dynamic features of the rod from Rpe65(-/-) animals, a well established model for Leber congenital amaurosis and vitamin A deficiency. The results of this study suggest that a systems-level approach can help to unravel the adaptation mechanisms in normal and in disease-associated conditions on a molecular basis.
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