Andressa Pinto - Academia.edu (original) (raw)

Papers by Andressa Pinto

Research paper thumbnail of Alteração de atributos químicos no solo de rampas utilizadas no tratamento de águas residuárias

Revista Brasileira de Engenharia Agrícola e Ambiental, 2005

Research paper thumbnail of Probing the Interaction of Brain Fatty Acid Binding Protein (B-FABP) with Model Membranes

PLoS ONE, 2013

Brain fatty acid-binding protein (B-FABP) interacts with biological membranes and delivers polyun... more Brain fatty acid-binding protein (B-FABP) interacts with biological membranes and delivers polyunsaturated fatty acids (FAs) via a collisional mechanism. The binding of FAs in the protein and the interaction with membranes involve a motif called ''portal region'', formed by two small a-helices, A1 and A2, connected by a loop. We used a combination of site-directed mutagenesis and electron spin resonance to probe the changes in the protein and in the membrane model induced by their interaction. Spin labeled B-FABP mutants and lipidic spin probes incorporated into a membrane model confirmed that B-FABP interacts with micelles through the portal region and led to structural changes in the protein as well in the micelles. These changes were greater in the presence of LPG when compared to the LPC models. ESR spectra of B-FABP labeled mutants showed the presence of two groups of residues that responded to the presence of micelles in opposite ways. In the presence of lysophospholipids, group I of residues, whose side chains point outwards from the contact region between the helices, had their mobility decreased in an environment of lower polarity when compared to the same residues in solution. The second group, composed by residues with side chains situated at the interface between the a-helices, experienced an increase in mobility in the presence of the model membranes. These modifications in the ESR spectra of B-FABP mutants are compatible with a less ordered structure of the portal region inner residues (group II) that is likely to facilitate the delivery of FAs to target membranes. On the other hand, residues in group I and micelle components have their mobilities decreased probably as a result of the formation of a collisional complex. Our results bring new insights for the understanding of the gating and delivery mechanisms of FABPs.

Research paper thumbnail of Understanding Chlorocatechol 1,2-Dioxygenase Function: A Promising Player in Bioremediation Processes

Biophysical Journal, 2012

Research paper thumbnail of Structural characterization of a recombinant flagellar calcium-binding protein from Trypanosoma cruzi

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, 2003

Conbercept is a genetically engineered homodimeric protein for the treatment of wet age-related m... more Conbercept is a genetically engineered homodimeric protein for the treatment of wet age-related macular degeneration (wet AMD) that functions by blocking VEGF-family proteins. Its huge, highly variable architecture makes characterization and development of a functional assay difficult. In this study, the primary structure, number of disulfide linkages and glycosylation state of conbercept were characterized by high-performance liquid chromatography, mass spectrometry, and capillary electrophoresis. Molecular modeling was then applied to obtain the spatial structural model of the conbercept-VEGF-A complex, and to study its inter-atomic interactions and dynamic behavior. This work was incorporated into a platform useful for studying the structure of conbercept and its ligand binding functions. Citation: Wu Z, Zhou P, Li X, Wang H, Luo D, et al. (2013) Structural Characterization of a Recombinant Fusion Protein by Instrumental Analysis and Molecular Modeling. PLoS ONE 8(3): e57642.

Research paper thumbnail of EPR Studies of Chlorocatechol 1,2-Dioxygenase: Evidences of Iron Reduction during Catalysis and of the Binding of Amphipatic Molecules

Biophysical Journal, 2005

Chlorocatechol 1,2-dioxygenase from Pseudomonas putida (Pp 1,2-CCD) is a dioxygenase responsible ... more Chlorocatechol 1,2-dioxygenase from Pseudomonas putida (Pp 1,2-CCD) is a dioxygenase responsible for ring cleavage during the degradation of recalcitrant aromatic compounds. We determined the zero-field splitting of the Fe(III) cofactor (jDj ¼ 1.3 6 0.2 cm ÿ1 ) by electron paramagnetic resonance (EPR) experiments that along with other structural data allowed us to infer the Fe(III) coordination environment. The EPR spectrum of the ion shows a significantly decrease of the g ¼ 4.3 resonance upon substrate binding. This result is rationalized in terms of a mechanism previously proposed, where catechol substrate is activated by Fe(III), yielding an exchange-coupled Fe(II)-semiquinone (pair). The Pp 1,2-CCD capacity of binding amphipatic molecules and the effects of such binding on protein activity are also investigated. EPR spectra of spin labels show a protein-bound component, which was characterized by means of spectral simulations. Our results indicate that Pp 1,2-CCD is able to bind amphipatic molecules in a channel with the headgroup pointing outwards into the solvent, whereas the carbon chain is held inside the tunnel. Protein assays show that the enzyme activity is significantly lowered in the presence of stearic-acid molecules. The role of the binding of those molecules as an enzyme activity modulator is discussed.

Research paper thumbnail of Catechol biosensing using a nanostructured layer-by-layer film containing Cl-catechol 1,2-dioxygenase

Biosensors & Bioelectronics, 2006

The detection of aromatic compounds from pesticides and industrial wastewater has become of great... more The detection of aromatic compounds from pesticides and industrial wastewater has become of great interest, since these compounds withstand chemical oxidation and biological degradation, accumulating in the environment. In this work, a highly sensitive biosensor for detecting catechol was obtained with the immobilization of Cl-catechol 1,2-dioxygenase (CCD) in nanostructured films. CCD layers were alternated with poly(amidoamine) generation 4 (PAMAM G4)

Research paper thumbnail of Alteração de atributos químicos no solo de rampas utilizadas no tratamento de águas residuárias

Revista Brasileira de Engenharia Agrícola e Ambiental, 2005

Research paper thumbnail of Probing the Interaction of Brain Fatty Acid Binding Protein (B-FABP) with Model Membranes

PLoS ONE, 2013

Brain fatty acid-binding protein (B-FABP) interacts with biological membranes and delivers polyun... more Brain fatty acid-binding protein (B-FABP) interacts with biological membranes and delivers polyunsaturated fatty acids (FAs) via a collisional mechanism. The binding of FAs in the protein and the interaction with membranes involve a motif called ''portal region'', formed by two small a-helices, A1 and A2, connected by a loop. We used a combination of site-directed mutagenesis and electron spin resonance to probe the changes in the protein and in the membrane model induced by their interaction. Spin labeled B-FABP mutants and lipidic spin probes incorporated into a membrane model confirmed that B-FABP interacts with micelles through the portal region and led to structural changes in the protein as well in the micelles. These changes were greater in the presence of LPG when compared to the LPC models. ESR spectra of B-FABP labeled mutants showed the presence of two groups of residues that responded to the presence of micelles in opposite ways. In the presence of lysophospholipids, group I of residues, whose side chains point outwards from the contact region between the helices, had their mobility decreased in an environment of lower polarity when compared to the same residues in solution. The second group, composed by residues with side chains situated at the interface between the a-helices, experienced an increase in mobility in the presence of the model membranes. These modifications in the ESR spectra of B-FABP mutants are compatible with a less ordered structure of the portal region inner residues (group II) that is likely to facilitate the delivery of FAs to target membranes. On the other hand, residues in group I and micelle components have their mobilities decreased probably as a result of the formation of a collisional complex. Our results bring new insights for the understanding of the gating and delivery mechanisms of FABPs.

Research paper thumbnail of Understanding Chlorocatechol 1,2-Dioxygenase Function: A Promising Player in Bioremediation Processes

Biophysical Journal, 2012

Research paper thumbnail of Structural characterization of a recombinant flagellar calcium-binding protein from Trypanosoma cruzi

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, 2003

Conbercept is a genetically engineered homodimeric protein for the treatment of wet age-related m... more Conbercept is a genetically engineered homodimeric protein for the treatment of wet age-related macular degeneration (wet AMD) that functions by blocking VEGF-family proteins. Its huge, highly variable architecture makes characterization and development of a functional assay difficult. In this study, the primary structure, number of disulfide linkages and glycosylation state of conbercept were characterized by high-performance liquid chromatography, mass spectrometry, and capillary electrophoresis. Molecular modeling was then applied to obtain the spatial structural model of the conbercept-VEGF-A complex, and to study its inter-atomic interactions and dynamic behavior. This work was incorporated into a platform useful for studying the structure of conbercept and its ligand binding functions. Citation: Wu Z, Zhou P, Li X, Wang H, Luo D, et al. (2013) Structural Characterization of a Recombinant Fusion Protein by Instrumental Analysis and Molecular Modeling. PLoS ONE 8(3): e57642.

Research paper thumbnail of EPR Studies of Chlorocatechol 1,2-Dioxygenase: Evidences of Iron Reduction during Catalysis and of the Binding of Amphipatic Molecules

Biophysical Journal, 2005

Chlorocatechol 1,2-dioxygenase from Pseudomonas putida (Pp 1,2-CCD) is a dioxygenase responsible ... more Chlorocatechol 1,2-dioxygenase from Pseudomonas putida (Pp 1,2-CCD) is a dioxygenase responsible for ring cleavage during the degradation of recalcitrant aromatic compounds. We determined the zero-field splitting of the Fe(III) cofactor (jDj ¼ 1.3 6 0.2 cm ÿ1 ) by electron paramagnetic resonance (EPR) experiments that along with other structural data allowed us to infer the Fe(III) coordination environment. The EPR spectrum of the ion shows a significantly decrease of the g ¼ 4.3 resonance upon substrate binding. This result is rationalized in terms of a mechanism previously proposed, where catechol substrate is activated by Fe(III), yielding an exchange-coupled Fe(II)-semiquinone (pair). The Pp 1,2-CCD capacity of binding amphipatic molecules and the effects of such binding on protein activity are also investigated. EPR spectra of spin labels show a protein-bound component, which was characterized by means of spectral simulations. Our results indicate that Pp 1,2-CCD is able to bind amphipatic molecules in a channel with the headgroup pointing outwards into the solvent, whereas the carbon chain is held inside the tunnel. Protein assays show that the enzyme activity is significantly lowered in the presence of stearic-acid molecules. The role of the binding of those molecules as an enzyme activity modulator is discussed.

Research paper thumbnail of Catechol biosensing using a nanostructured layer-by-layer film containing Cl-catechol 1,2-dioxygenase

Biosensors & Bioelectronics, 2006

The detection of aromatic compounds from pesticides and industrial wastewater has become of great... more The detection of aromatic compounds from pesticides and industrial wastewater has become of great interest, since these compounds withstand chemical oxidation and biological degradation, accumulating in the environment. In this work, a highly sensitive biosensor for detecting catechol was obtained with the immobilization of Cl-catechol 1,2-dioxygenase (CCD) in nanostructured films. CCD layers were alternated with poly(amidoamine) generation 4 (PAMAM G4)