14 results on '"Abian, Olga"'
Search Results
2. Biophysical Screening for Identifying Pharmacological Chaperones and Inhibitors Against Conformational and Infectious Diseases.
- Author
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Velazquez-Campoy A, Sancho J, Abian O, and Vega S
- Subjects
- Animals, Communicable Diseases drug therapy, Communicable Diseases physiopathology, Humans, Ligands, Molecular Chaperones pharmacology, Phenylketonurias drug therapy, Phenylketonurias physiopathology, Protein Conformation, Proteins chemistry, Drug Design, Molecular Targeted Therapy, Proteins metabolism
- Abstract
Experimental and computational screenings are currently widespread tools for identifying either potential ligands for a given target or potential targets for a given chemical compound. In particular, ligand-induced stabilization against thermal denaturation (or thermal shift assay) is an easy and convenient experimental procedure for finding compounds able to control the activity of a protein target (e.g., allosteric or competitive inhibitors interfering with its activity, allosteric activators enhancing its activity, or pharmacological chaperones avoiding aggregation, unfolding or degradation). Despite its simplicity the thermal shift assay does not lack some important drawbacks. Here we describe this methodology, its application to structured and unstructured protein targets, and we discuss successful cases of identification of bioactive compounds for conformational disorders associated to loss of function (phenylketonuria) and infectious diseases (gastric ulcer and hepatitis C).
- Published
- 2016
- Full Text
- View/download PDF
3. Studying the allosteric energy cycle by isothermal titration calorimetry.
- Author
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Martinez-Julvez M, Abian O, Vega S, Medina M, and Velazquez-Campoy A
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- Allosteric Regulation physiology, Protein Binding, Proteins chemistry, Thermodynamics, Calorimetry methods, Proteins metabolism
- Abstract
Isothermal titration calorimetry (ITC) is a powerful biophysical technique which allows a complete thermodynamic characterization of protein interactions with other molecules. The possibility of dissecting the Gibbs energy of interaction into its enthalpic and entropic contributions, as well as the detailed additional information experimentally accessible on the intermolecular interactions (stoichiometry, cooperativity, heat capacity changes, and coupled equilibria), make ITC a suitable technique for studying allosteric interactions in proteins. Two experimental methodologies for the characterization of allosteric heterotropic ligand interactions by ITC are described in this chapter, illustrated with two proteins with markedly different structural and functional features: a photosynthetic electron transfer protein and a drug target viral protease.
- Published
- 2012
- Full Text
- View/download PDF
4. Determination of protein-protein interactions through aldehyde-dextran intermolecular cross-linking.
- Author
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Fuentes M, Segura RL, Abian O, Betancor L, Hidalgo A, Mateo C, Fernandez-Lafuente R, and Guisan JM
- Subjects
- Animals, Electrophoresis, Polyacrylamide Gel, Models, Molecular, Protein Conformation, Rabbits, Aldehydes chemistry, Cross-Linking Reagents chemistry, Dextrans chemistry, Proteins chemistry
- Abstract
A very simple strategy, based on the intermolecular cross-linking of associated proteins by using aldehyde-dextrans, has been proposed to detect protein-protein interactions. Aldehyde-dextran was able to cross-link different enzymes composed of several polypeptide chains (e.g., trypsin and penicillin G acylase), proteolyzated proteins (e.g., extracts from porcine pancreas) and finally, an immunocomplex (horseradish peroxidase/anti-horseradish peroxidase). This cross-linked immunocomplex could be selectively adsorbed on immobilized anti-rabbit IgG. The presence of unspecific covalent attachment between unrelated protein molecules was not detected. Thus, this strategy permits the cross-linking of different protein components and avoids the formation of nonspecific protein-protein associations.
- Published
- 2004
- Full Text
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5. Reversible and strong immobilization of proteins by ionic exchange on supports coated with sulfate-dextran.
- Author
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Fuentes M, Pessela BC, Maquiese JV, Ortiz C, Segura RL, Palomo JM, Abian O, Torres R, Mateo C, Fernández-Lafuente R, and Guisán JM
- Subjects
- Adsorption, Electrophoresis, Polyacrylamide Gel, Ion Exchange Resins, Dextran Sulfate chemistry, Proteins chemistry
- Abstract
New and strong ionic exchange resins have been prepared by the simple and rapid ionic adsorption of anionic polymers (sulfate-dextran) on porous supports activated with the opposite ionic group (DEAE/MANAE). Ionic exchange properties of such composites were strongly dependent on the size of the ionic polymers as well as on the conditions of the ionic coating of the solids with the ionic polymers (optimal conditions were 400 mg of sulfate-dextran 5000 kDa per gram of support). Around 80% of the proteins contained in crude extracts from Escherichia coli and Acetobacter turbidans could be adsorbed on these porous composites even at pH 7. This interaction was stronger than that using conventional carboxymethyl cellulose (CMC) and even others such as supports coated with aspartic-dextran polymer. By means of the sequential use of the new supports and supports coated with polyethyleneimine (PEI), all proteins from crude extracts could be immobilized. In fact, a large percentage (over 50%) could be immobilized on both supports. Finally, some industrially relevant enzymes (beta-galactosidases from Aspergillus oryzae, Kluyveromyces lactis, and Thermussp. strain T2, lipases from Candida antarctica A and B, Candida rugosa, Rhizomucor miehei, and Rhyzopus oryzae and bovine pancreas trypsin and chymotrypsin) have been immobilized on these supports with very high activity recoveries and immobilization rates. After enzyme inactivation, the protein could be fully desorbed from the support, and then the support could be reused for several cycles. Moreover, in some instances the enzyme stability was significantly improved, mainly in the presence of organic solvents, perhaps as a consequence of the highly hydrophilic microenvironment of the support.
- Published
- 2004
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- View/download PDF
6. Epoxy-amino groups: a new tool for improved immobilization of proteins by the epoxy method.
- Author
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Mateo C, Torres R, Fernández-Lorente G, Ortiz C, Fuentes M, Hidalgo A, López-Gallego F, Abian O, Palomo JM, Betancor L, Pessela BC, Guisan JM, and Fernández-Lafuente R
- Subjects
- Adsorption drug effects, Proteins metabolism, Epoxy Compounds pharmacology, Proteins antagonists & inhibitors
- Abstract
The properties of a new commercially available amino-epoxy support (amino-epoxy-Sepabeads) for immobilizing enzymes have been compared to those of conventional epoxy supports. The new support has a layer of epoxy groups over a layer of ethylenediamine that is covalently bound to the support. Thus, this support has a great anionic exchanger power and a high number of epoxy groups. We have found a number of advantages to this new heterofunctional support. Immobilization proceeds at low ionic strength using amino epoxy Sepabeads while requiring high ionic strength using conventional monofunctional epoxy supports. Immobilization is much more rapid using amino-epoxy supports than employing conventional epoxy supports. The possibility of achieving immobilized preparations in which the enzyme orientation may be different to that obtained using the traditional hydrophobic supports (with likely effects in terms of activity or stability). Stability of the immobilized enzyme has been found to be much higher using the new support than in preparations using the conventional ones in many cases. Here we show some examples of these advantages using different enzymes (beta-galactosidases, lipase, glutaryl acylase, invertase, and glucoamylase).
- Published
- 2003
- Full Text
- View/download PDF
7. Targeting intrinsically disordered proteins involved in cancer.
- Author
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Santofimia-Castaño, Patricia, Rizzuti, Bruno, Xia, Yi, Abian, Olga, Peng, Ling, Velázquez-Campoy, Adrián, Neira, José L., and Iovanna, Juan
- Subjects
SMALL molecules ,PROTEINS ,CELLULAR control mechanisms ,MACROMOLECULES ,CANCER - Abstract
Intrinsically disordered proteins (IDPs) do not have a well-defined structure under physiological conditions, but they have key roles in cell signaling and regulation, and they are frequently related to the development of diseases, such as cancer and other malignancies. This has converted IDPs in attractive therapeutic targets; however, targeting IDPs is challenging because of their dynamic nature. In the last years, different experimental and computational approaches, as well as the combination of both, have been explored to identify molecules to target either the hot-spots or the allosteric sites of IDPs. In this review, we summarize recent developments in successful targeting of IDPs, all of which are involved in different cancer types. The strategies used to develop and design (or in one particular example, to repurpose) small molecules targeting IDPs are, in a global sense, similar to those used in well-folded proteins: (1) screening of chemically diverse or target-oriented compound libraries; or (2) study of the interfaces involved in recognition of their natural partners, and design of molecular candidates capable of binding to such binding interface. We describe the outcomes of using these approaches in targeting IDPs involved in cancer, in the view to providing insight, to target IDPs in general. In a broad sense, the designed small molecules seem to target the most hydrophobic regions of the IDPs, hampering macromolecule (DNA or protein)–IDP interactions; furthermore, in most of the molecule–IDP complexes described so far, the protein remains disordered. [ABSTRACT FROM AUTHOR]
- Published
- 2020
- Full Text
- View/download PDF
8. Allosteric Inhibitors of the NS3 Protease from the Hepatitis C Virus.
- Author
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Abian, Olga, Vega, Sonia, Sancho, Javier, and Velazquez-Campoy, Adrian
- Subjects
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PROTEASE inhibitors , *ALLOSTERIC enzymes , *HEPATITIS C virus , *VIRAL proteins , *LIVER cells , *DRUG development , *ENZYME kinetics - Abstract
The nonstructural protein 3 (NS3) from the hepatitis C virus processes the non-structural region of the viral precursor polyprotein in infected hepatic cells. The NS3 protease activity has been considered a target for drug development since its identification two decades ago. Although specific inhibitors have been approved for clinical therapy very recently, resistance-associated mutations have already been reported for those drugs, compromising their long-term efficacy. Therefore, there is an urgent need for new anti-HCV agents with low susceptibility to resistance-associated mutations. Regarding NS3 protease, two strategies have been followed: competitive inhibitors blocking the active site and allosteric inhibitors blocking the binding of the accessory viral protein NS4A. In this work we exploit the intrinsic Zn+2-regulated plasticity of the protease to identify a new type of allosteric inhibitors. In the absence of Zn+2, the NS3 protease adopts a partially-folded inactive conformation. We found ligands binding to the Zn+2-free NS3 protease, trap the inactive protein, and block the viral life cycle. The efficacy of these compounds has been confirmed in replicon cell assays. Importantly, direct calorimetric assays reveal a low impact of known resistance-associated mutations, and enzymatic assays provide a direct evidence of their inhibitory activity. They constitute new low molecular-weight scaffolds for further optimization and provide several advantages: 1) new inhibition mechanism simultaneously blocking substrate and cofactor interactions in a non-competitive fashion, appropriate for combination therapy; 2) low impact of known resistance-associated mutations; 3) inhibition of NS4A binding, thus blocking its several effects on NS3 protease. [ABSTRACT FROM AUTHOR]
- Published
- 2013
- Full Text
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9. Some special features of glyoxyl supports to immobilize proteins
- Author
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Mateo, Cesar, Abian, Olga, Bernedo, Marieta, Cuenca, Emma, Fuentes, Manuel, Fernandez-Lorente, Gloria, Palomo, Jose M., Grazu, Valeria, Pessela, Benevides C.C., Giacomini, Cecilia, Irazoqui, Gabriela, Villarino, Andrea, Ovsejevi, Karen, Batista-Viera, Francisco, Fernandez-Lafuente, Roberto, and Guisán, Jose M.
- Subjects
- *
PROTEINS , *ENZYMES , *ENZYMOLOGY , *HYDROGEN-ion concentration - Abstract
Abstract: This paper described that glyoxyl activated supports are only able to quantitatively immobilize proteins at alkaline pH values and using highly activated supports (otherwise, immobilization is negligible). Furthermore, the immobilization rate depends on the surface density of glyoxyl groups and not on the total concentration of groups in the suspension. Moreover, the temperature presented a dramatic effect on enzyme immobilization rate and different enzymes become immobilized at very different rates. Finally, it was not possible to detect immobilization of mono aminated compounds at neither alkaline nor neutral pH values. In fact, these supports may be used to purify enzymes due to these great differences in immobilization rate (e.g., glutamate racemase). These special features are not shared with other supports usually utilized for covalent immobilization of enzymes (e.g., glutaraldehyde, cyanogen bromide supports) that are able to immobilize proteins by just one very stable bond and suggesting an unique mechanism for glyoxyl agarose supports. In fact, all these results suggested that each individual amino-glyoxyl bond is very weak. Therefore, the first immobilization of proteins on supports activated with glyoxyl groups, in absence of reducing reagents, needs the simultaneous establishment of, at least, two attachments between the protein and the support. This mechanism promotes that proteins become immobilized by the area/s where the highest lysine residues density is located and explain the high enzyme stabilization usually achieved by using this immobilization technique. [Copyright &y& Elsevier]
- Published
- 2005
- Full Text
- View/download PDF
10. Erratum to "Isolated auto-citrullinated regions of PADI4 associate to the intact protein without altering their disordered conformation" [Biophysical Chemistry ...(2024) ...].
- Author
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Neira, José L., Rizzuti, Bruno, Abian, Olga, and Velazquez-Campoy, Adrian
- Subjects
- *
PHYSICAL biochemistry , *PROTEINS - Published
- 2024
- Full Text
- View/download PDF
11. Isolated auto-citrullinated regions of PADI4 associate to the intact protein without altering their disordered conformation.
- Author
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Neira, José L., Rizzuti, Bruno, Abian, Olga, and Velazquez-Campoy, Adrian
- Subjects
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ISOTHERMAL titration calorimetry , *PEPTIDOMIMETICS , *CIRCULAR dichroism , *MOLECULAR docking , *PROTEINS , *ULTRAVIOLET spectroscopy - Abstract
PADI4 is one of the human isoforms of a group of enzymes intervening in the conversion of arginine to citrulline. It is involved in the development of several types of tumors, as well as other immunological illnesses, such as psoriasis, multiple sclerosis, or rheumatoid arthritis. PADI4 auto-citrullinates in several regions of its sequence, namely in correspondence of residues Arg205, Arg212, Arg218, and Arg383. We wanted to study whether the citrullinated moiety affects the conformation of nearby regions and its binding to intact PADI4. We designed two series of synthetic peptides comprising either the wild-type or the relative citrullinated versions of such regions – i.e., a first series of peptides comprising the first three arginines, and a second series comprising Arg383. We studied their conformational properties in isolation by using fluorescence, far-ultraviolet (UV) circular dichroism (CD), and 2D 1H NMR. Furthermore, we characterized the binding of the wild-type and citrullinated peptides in the two series to the intact PADI4, by using isothermal titration calorimetry (ITC), fluorescence, and biolayer interferometry (BLI), as well as by molecular docking simulations. We observed that citrullination did not alter the local conformational propensities of the isolated peptides. Nevertheless, for all the peptides in the two series, citrullination slowed down the kinetic k off rates of the binding reaction to PADI4, probably due to differences in electrostatic effects compared to the presence of arginine. The affinities of PADI4 for unmodified peptides were slightly larger than those of the corresponding citrullinated ones in the two series, but they were all within the same range, indicating that there were no relevant variations in the thermodynamics of binding due to sequence effects. These results highlight details of the self-citrullination of PADI4 and, more generally, of possible auto-catalytic mechanisms taking place in vivo for other citrullinating enzymes or, alternatively, in proteins undergoing citrullination passively. [Display omitted] • Citrullinated regions of PADI4 have the same conformation as wild-type counterparts. • Citrullinated regions bind to PADI4 with different kinetic rates than wild-type. • Citrullinated and wild-type regions bind to PADI4 with nearly the same affinity. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
12. MeCP2-E1 isoform is a dynamically expressed, weakly DNA-bound protein with different protein and DNA interactions compared to MeCP2-E2.
- Author
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Martínez de Paz, Alexia, Khajavi, Leila, Martin, Hélène, Claveria-Gimeno, Rafael, Tom Dieck, Susanne, Cheema, Manjinder S., Sanchez-Mut, Jose V., Moksa, Malgorzata M., Carles, Annaick, Brodie, Nick I., Sheikh, Taimoor I., Freeman, Melissa E., Petrotchenko, Evgeniy V., Borchers, Christoph H., Schuman, Erin M., Zytnicki, Matthias, Velazquez-Campoy, Adrian, Abian, Olga, Hirst, Martin, and Esteller, Manel
- Subjects
PROTEIN-protein interactions ,N-terminal residues ,RETT syndrome ,AMINO acid residues ,PROTEINS ,NEURAL development - Abstract
Background: MeCP2—a chromatin-binding protein associated with Rett syndrome—has two main isoforms, MeCP2-E1 and MeCP2-E2, differing in a few N-terminal amino acid residues. Previous studies have shown brain region-specific expression of these isoforms which, in addition to their different cellular localization and differential expression during brain development, suggest that they may also have non-overlapping molecular mechanisms. However, differential functions of MeCP2-E1 and E2 remain largely unexplored. Results: Here, we show that the N-terminal domains (NTD) of MeCP2-E1 and E2 modulate the ability of the methyl-binding domain (MBD) to interact with DNA as well as influencing the turn-over rates, binding dynamics, response to neuronal depolarization, and circadian oscillations of the two isoforms. Our proteomics data indicate that both isoforms exhibit unique interacting protein partners. Moreover, genome-wide analysis using ChIP-seq provide evidence for a shared as well as a specific regulation of different sets of genes. Conclusions: Our study supports the idea that Rett syndrome might arise from simultaneous impairment of cellular processes involving non-overlapping functions of MECP2 isoforms. For instance, MeCP2-E1 mutations might impact stimuli-dependent chromatin regulation, while MeCP2-E2 mutations could result in aberrant ribosomal expression. Overall, our findings provide insight into the functional complexity of MeCP2 by dissecting differential aspects of its two isoforms. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
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13. Enzyme structure and function protection from gastrointestinal degradation using enteric coatings.
- Author
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Gracia, Ruben, Yus, Cristina, Abian, Olga, Mendoza, Gracia, Irusta, Silvia, Sebastian, Victor, Andreu, Vanesa, and Arruebo, Manuel
- Subjects
- *
SERUM albumin , *EVAPORATION (Chemistry) , *PROTEINS , *PEROXIDASE , *EMULSIONS - Abstract
Abstract Bovine Serum Albumin (BSA) and Horseradish Peroxidase (HRP) have been encapsulated within microparticulated matrices composed of Eudragit RS100 by the water-in-oil-in-water double emulsion solvent evaporation method. Good encapsulation efficiencies were achieved for BSA and HRP, 88.4 and 95.8%, respectively. The stability of the loaded proteins was confirmed by using circular dichroism and fluorescence. The gastroresistance of the protein-loaded microparticles was evaluated under simulated gastric conditions demonstrating the preservation of the structural integrity of the proteins loaded inside the particles. The enzymatic activity of HRP after being released from the enteric microparticles was evaluated by using the peroxidase substrate, revealing that the released enzyme preserved its 100% function. The high drug loadings achieved, reduced cytotoxicity and efficient gastric protection point out towards the potential use of those carriers as oral delivery vectors of therapeutic proteins offering a more controlled targeted release in specific sites of the intestine and an enhanced gastrointestinal absorption. [ABSTRACT FROM AUTHOR]
- Published
- 2018
- Full Text
- View/download PDF
14. Experimental validation of in silico target predictions on synergistic protein targets.
- Author
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Cortes-Ciriano, Isidro, Koutsoukas, Alexios, Abian, Olga, Bender, Andreas, and Velazquez-Campoy, Adrian
- Subjects
PROTEINS ,DRUG development ,PHENOTYPES ,GENETIC testing ,ANTINEOPLASTIC agents - Abstract
A conference paper about an experimental validation of in silico target predictions on synergistic protein targets is presented. It informs that the drug discovery settings, firstly a revival of phenotypic screening strategies. It further informs about the experimental phenotypic screening for cytotoxic compounds with an experimental validation of individual protein targets.
- Published
- 2013
- Full Text
- View/download PDF
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