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Papers by ana licia veiga

Forum Patrimonio Ambiente Construido E Patrimonio Sustentavel, Feb 9, 2013

Na engenharia e arquitetura, o escaneamento a laser de fachadas revolucionou a forma de se regist... more Na engenharia e arquitetura, o escaneamento a laser de fachadas revolucionou a forma de se registrarem informacoes topograficas e construtivas de detalhe. Scanners a laser (Laser Scanner – LS) podem capturar 12.000 pontos por segundo ou mais, sem a necessidade de refletores. Eles registram nao somente as dimensoes, mas tambem a localizacao espacial, a cor e a textura do objeto alvo. O sistema requer pouco tempo de processamento dos dados a posteriori e uma equipe de execucao minima. Por fim, o escaneamento a laser resulta, ao mesmo tempo, no levantamento fotografico metrico/dimensional e num modelo virtual de altissima precisao. Trata-se, portanto, de uma fotografia tridimensional metrificada. A maquete eletronica, uma vez manipulada nos softwares dedicados, gera inumeros mapas em diversos formatos. Alem do modelo digital, a tecnologia LS pode originar maquetes reais perfeitas, a partir de recursos de Engenharia Reversa (ER). Neste trabalho, objetivamos descrever o funcionamento da tecnologia de escaneamento a laser (LS), assim como provar a eficiencia da aplicacao desta tecnologia no mapeamento urbanistico e arquitetonico de edificacoes patrimoniais e espacos culturais, atraves de um estudo de caso por nos conduzido, juntamente com a empresa MAPTEK, no Espaco Cultural da Avenida Bernardo Monteiro, em Belo Horizonte – MG.

Cadernos Pagu, 2011

Catarina e bolsista da CAPES. Atualmente pesquisa sobre realizadoras de cinema do Brasil e da Arg... more Catarina e bolsista da CAPES. Atualmente pesquisa sobre realizadoras de cinema do Brasil e da Argentina durante as ditaduras militares nos dois países. amveiga@yahoo.com.br 1 Em 2010 as jornadas alcançaram sua terceira edição.

Revista Estudos Feministas, 2014

Bioinformatics (Oxford, England), Jan 28, 2015

The need for more effective and safer pharmaceuticals is a persistent quest. Microbial adaptation... more The need for more effective and safer pharmaceuticals is a persistent quest. Microbial adaptations create the need to permanently develop new antimicrobials, for instance. Similarly, intracellular delivery of drugs is still a challenge and translocation of membranes for drug delivery is an area of intense research. Peptides can be used both as antimicrobial drug leads and drug carrier systems for intracellular delivery. Multifunctional proteins are abundant in viruses but, surprisingly, have never been thoroughly screened for bioactive peptide sequences. Using the AMPA and CellPPD online tools we have evaluated the propensity of viral proteins to comprise antimicrobial (AMP) or cell-penetrating peptides (CPP). Capsid proteins from both enveloped and non-enveloped viruses, and membrane and envelope proteins from enveloped viruses, in a total of 272 proteins from 133 viruses, were screened to detect the presence of potential AMP and CPP sequences. A pool of 2444 and 426 CPP and AMP se...

Journal of Peptide Science, 2015

Despite the intensive study on the mechanism of action of membrane-active molecules such as antim... more Despite the intensive study on the mechanism of action of membrane-active molecules such as antimicrobial and anticancer peptides, most of the biophysical work has been performed using artificial model systems, mainly lipid vesicles. The use of these systems allows full control of the experimental parameters, and to obtain molecular-level detail on the action of peptides, the correlation with biological action is intangible. Recently, several biophysical methodologies have been translated to studies using bacterial and cancer cells. Here, we review biophysical studies on the mechanism of action of antimicrobial and anticancer peptides performed directly on cells. The data in these studies allow to correlate vesicle-based and cell-based studies and fill the vesicle-cell interdisciplinary gap. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2015

Recent patents on anti-infective drug discovery, 2006

The main strategies nowadays to fight AIDS rely on chemical therapy to inhibit the reverse transc... more The main strategies nowadays to fight AIDS rely on chemical therapy to inhibit the reverse transcriptase or protease of HIV. However, a synthetic 36 amino-acids peptide that blocks the entry of the virus in the target cells (enfuvirtide) has recently reached approval for clinical application. This molecule may probably be just the leader of a new generation of drugs that is about to emerge to interrupt the first step in the HIV life cycle, i.e. preventing the virus from actually entering cells. This paper reviews the enfuvirtide path from clinical trials to the attempts to detail its molecular-level mode of action. It is commonly accepted that this peptide would block the fusion between viral and cell plasma membrane through binding to the N-terminal heptad repeat (NHR) region of the viral protein gp41. However, there has been growing evidence that this model of action may be unrealistic, the action of enfuvirtide being more complex and diverse than initially thought. Membrane-assisted local concentration increase and interference with gp120/co-receptor docking may also contribute for the inhibitory action of the peptide. Selected HIV-entry inhibitors on clinical trials are presented to characterize the future drugs in the market in this class.

Significação: Revista de Cultura Audiovisual, 2013

Thrombosis Research, 2004

Envenomation caused by Lonomia obliqua caterpillars is an increasing problem in Southern Brazil. ... more Envenomation caused by Lonomia obliqua caterpillars is an increasing problem in Southern Brazil. The clinical profile is characterized by a profound hemorrhagic disorder. In the present study, we describe the characterization of a fibrin(ogen)olytic factor (lonofibrase) isolated from a venomous secretion of the caterpillars. The crude extract showed a dose-dependent inhibitory effect in the rate of thrombininduced fibrinogen clotting and produced fragmentation of fibrinogen. Isolation of the fibrin(ogen)olytic enzyme was achieved by combining ion exchange chromatography followed by gel filtration in a fast protein liquid chromatography (FPLC) system. A single 35-kDa band was identified and the isolated enzyme named lonofibrase. Lonofibrase rapidly degrades Aa and Bh chains of fibrinogen, also being able to cleave fibrin in a distinct way from that observed with plasmin. The presence of lonofibrase with both fibrinogenolytic and fibrinolytic activities in L. obliqua secretion is coherent with the severe hemorrhagic clinical profile resulting from envenomation caused by these insects.

FEBS Journal, 2013

Cell penetrating peptides (CPPs) can be used as drug delivery systems for different therapeutic m... more Cell penetrating peptides (CPPs) can be used as drug delivery systems for different therapeutic molecules. In this work two novel CPPs, pepR and pepM, designed from two domains of the dengue virus (DENV) capsid protein, were studied for their ability to deliver nucleic acids into cells as non-covalently bound cargo. Translocation studies were performed by confocal microscopy in HepG2, BHK and HEK cell lineages, astrocytes and peripheral blood mononuclear cells. Combined studies in HepG2 cells, astrocytes and BHK cells, at 4 and 37°C or using specific endocytosis inhibitors, revealed that pepR and pepM use distinct internalization routes: pepM translocates lipid membranes directly, while pepR uses an endocytic pathway. To confirm these results, a methodology was developed to monitor the translocation kinetics of both peptides by real-time flow cytometry. Kinetic constants were determined, and the amount of nucleic acids delivered was estimated. Additional studies were performed in order to understand the molecular bases of the peptide-mediated translocation. Peptide-nucleic acid and peptide-lipid membrane interactions were studied quantitatively based on the intrinsic fluorescence of the peptides. pepR and pepM bound ssDNA to the same extent. Partition studies revealed that both peptides bind preferentially to anionic lipid membranes, adopting an a-helical conformation. However, fluorescence quenching studies suggest that pepM is deeply inserted into the lipid bilayer, in contrast with pepR. Moreover, only pepM is able to promote the fusion and aggregation of vesicles composed of zwitterionic lipids. Altogether, the results show that DENV capsid protein derived peptides serve as good templates for novel CPP-based nucleic acid delivery strategies, defining different routes for cell entry.

FEBS Journal, 2007

A small amino acid sequence (LWYIK) inside the HIV-1 gp41 ectodomain membrane proximal region (MP... more A small amino acid sequence (LWYIK) inside the HIV-1 gp41 ectodomain membrane proximal region (MPR) is commonly referred to as a cholesterol-binding domain. To further study this unique and peculiar property we have used fluorescence spectroscopy techniques to unravel the membrane interaction properties of three MPR-derived synthetic peptides: the membrane proximal region peptide-complete (MPRP-C) which corresponds to the complete MPR; the membrane proximal region peptide-short (MPRP-S), which corresponds to the last five MPR amino acid residues (the putative cholesterol-binding domain) and the membrane proximal region peptide-intermediate (MPRP-I), which corresponds to the MPRP-C peptide without the MPRP-S sequence. MPRP-C and MPRP-I membrane interaction is largely independent of the membrane phase. Membrane interaction of MPRP-S occurs for fluid phase membranes but not in gel phase membranes or cholesterol-containing bilayers. The gp41 ectodomain MPR may have a very specific function in viral fusion through the concerted and combined action of cholesterol-binding and non-cholesterol-binding domains (i.e. domains corresponding to MPRP-S and MPRP-I, respectively).

PLoS ONE, 2013

Supercharged proteins are a recently identified class of proteins that have the ability to effici... more Supercharged proteins are a recently identified class of proteins that have the ability to efficiently deliver functional macromolecules into mammalian cells. They were first developed as bioengineering products, but were later found in the human proteome. In this work, we show that this class of proteins with unusually high net positive charge is frequently found among viral structural proteins, more specifically among capsid proteins. In particular, the capsid proteins of viruses from the Flaviviridae family have all a very high net charge to molecular weight ratio (. +1.07/kDa), thus qualifying as supercharged proteins. This ubiquity raises the hypothesis that supercharged viral capsid proteins may have biological roles that arise from an intrinsic ability to penetrate cells. Dengue virus capsid protein was selected for a detailed experimental analysis. We showed that this protein is able to deliver functional nucleic acids into mammalian cells. The same result was obtained with two isolated domains of this protein, one of them being able to translocate lipid bilayers independently of endocytic routes. Nucleic acids such as siRNA and plasmids were delivered fully functional into cells. The results raise the possibility that the ability to penetrate cells is part of the native biological functions of some viral capsid proteins.

Journal of Viral Hepatitis, 2008

We have analysed and identified different membrane-active regions of the Hepatitis C virus (HCV) ... more We have analysed and identified different membrane-active regions of the Hepatitis C virus (HCV) core protein by observing the effect of 18-mer core-derived peptide libraries from two HCV strains on the integrity of different membrane model systems. In addition, we have studied the secondary structure of specific membrane-interacting peptides from the HCV core protein, both in aqueous solution and in the presence of model membrane systems. Our results show that the HCV core protein region comprising the C-terminus of domain 1 and the N-terminus of domain 2 seems to be the most active in membrane interaction, although a role in protein-protein interaction cannot be excluded. Significantly, the secondary structure of nearly all the assayed peptides changes in the presence of model membranes. These sequences most probably play a relevant part in the biological action of HCV in lipid interaction. Furthermore, these membranotropic regions could be envisaged as new possible targets, as inhibition of its interaction with the membrane could potentially lead to new vaccine strategies.

Journal of the American Chemical Society, 2012

Several cationic antimicrobial peptides (AMPs) have recently been shown to display anticancer act... more Several cationic antimicrobial peptides (AMPs) have recently been shown to display anticancer activity via a mechanism that usually entails the disruption of cancer cell membranes. In this work, we designed an 18-residue anticancer peptide, SVS-1, whose mechanism of action is designed to take advantage of the aberrant lipid composition presented on the outer leaflet of cancer cell membranes, which makes the surface of these cells relatively electronegative relative to noncancerous cells. SVS-1 is designed to remain unfolded and inactive in aqueous solution but preferentially fold at the surface of cancer cells, adopting an amphiphilic β-hairpin structure capable of membrane disruption. Membrane-induced folding is driven by electrostatic interaction between the peptide and the negatively charge membrane surface of cancer cells. SVS-1 is active against a variety of cancer cell lines such as A549 (lung carcinoma), KB (epidermal carcinoma), MCF-7 (breast carcinoma) and MDA-MB-436 (breast carcinoma). However, the cytotoxicity towards non-cancerous cells having typical membrane compositions, such as HUVEC and erythrocytes, is low. CD spectroscopy, appropriately designed peptide controls, cell-based studies, liposome leakage assays and electron microscopy support the intended mechanism of action, which leads to preferential killing of cancerous cells.

Journal of Peptide Science, 2013

One of the major challenges in the drug development process is biodistribution across epithelia a... more One of the major challenges in the drug development process is biodistribution across epithelia and intracellular drug targeting. Cellular membrane heterogeneity is one of the major drawbacks in developing efficient and sustainable drug delivery systems, which brings the need to study their interaction with lipids in order to unravel their mechanisms of action and improve their delivery capacities. Cell penetrating peptides (CPPs) are able to translocate almost any cell membrane carrying cargo molecules. However, different CPP use different entry mechanisms, which are often concentration-dependent and cargo-dependent. Being able to quantify the lipid affinity of CPP is of obvious importance and can be achieved by studying the partition extent of CPP into lipid bilayers. The partition constant (K p) reflects the lipid-water partition extent. However, all currently available methodologies are only suitable to determine the partition of single molecules into lipid membranes or entities, being unsuitable to determine the partition of bimolecular or higher order supramolecular complexes. We derived and tested a mathematical model to determine the K p of supramolecular CPP-cargo complexes from fluorescence spectroscopy data, using DNA oligomers as a model cargo. As a proof-of-concept example, the partition extent of two new membrane active peptides derived from dengue virus capsid protein (DENV C protein) with potential CPP properties, in both scenarios (free peptide and complexed with a molecular cargo), were tested. We were able to identify the lipid affinity of these CPP:DNA complexes, thus gaining valuable insights into better CPP formulations.

Journal of Molecular Structure, 2014

h i g h l i g h t s Antibody therapeutic interest has grown in the past decade. Lipid membranes m... more h i g h l i g h t s Antibody therapeutic interest has grown in the past decade. Lipid membranes may play an important role in epitope recognition. Fluorescence spectroscopy can be used to monitor antibody-membrane interactions. Its cost-effectiveness makes it a potential alternative to other techniques.

IUBMB Life, 2014

The current landscapes of novel therapeutic approaches rely mostly on gene-targeted technologies,... more The current landscapes of novel therapeutic approaches rely mostly on gene-targeted technologies, enabling to fight rare genomic diseases, from infections to cancer and hereditary diseases. Although, reaching the action-site for this novel treatments requires to deliver nucleic acids, or other macromolecules into cells, which may pose difficult tasks to pharmaceutical companies. To overcome this technological limitation, a wide variety of vectors have been developed in the past decades and have proven to be successful in delivering various therapeutics. Cell-penetrating peptides (CPP) have been one of the technologies widely studied and have been increasingly used to transport small RNA/DNA, plasmids, antibodies, and nanoparticles into cells. Despite the already proved huge potential that these peptide-based approaches may suggest, few advances have been put to pharmacological or clinical use. This review will describe the origin, development, and usage of CPP to deliver therapeutic agents into cells, with special emphasis on their current application to gene-therapies. Specifically, we will describe the current trials being conducted to treat cancer, gene disorders, and autoimmune diseases using CPP-based therapies.

European Biophysics Journal, 2011

Many cellular phenomena occur on the biomembranes. There are plenty of molecules (natural or xeno... more Many cellular phenomena occur on the biomembranes. There are plenty of molecules (natural or xenobiotics) that interact directly or partially with the cell membrane. Biomolecules, such as several peptides (e.g., antimicrobial peptides) and proteins, exert their effects at the cell membrane level. This feature makes necessary investigating their interactions with lipids to clarify their mechanisms of action and side effects necessary. The determination of molecular lipid/water partition constants (K p) is frequently used to quantify the extension of the interaction. The determination of this parameter has been achieved by using different methodologies, such as UV-Vis absorption spectrophotometry, fluorescence spectroscopy and f-potential measurements. In this work, we derived and tested a mathematical model to determine the K p from f-potential data. The values obtained with this method were compared with those obtained by fluorescence spectroscopy, which is a regular technique used to quantify the interaction of intrinsically fluorescent peptides with selected biomembrane model systems. Two antimicrobial peptides (BP100 and pepR) were evaluated by this new method. The results obtained by this new methodology show that f-potential is a powerful technique to quantify peptide/lipid interactions of a wide variety of charged molecules, overcoming some of the limitations inherent to other techniques, such as the need for fluorescent labeling. Keywords Partition constant Á Zeta-potential Á Fluorescence Á Membrane Á Antimicrobial peptides Membrane-active peptides: 455th WE-Heraeus-Seminar and AMP 2010 Workshop.

Revista Estudos Feministas, 2006

Revista Estudos Feministas, 2009

A historiadora holandesa Kathy Davis compartilha no livro How Feminism Travels across Borders, de... more A historiadora holandesa Kathy Davis compartilha no livro How Feminism Travels across Borders, de maneira autêntica e singular, os resultados de uma ampla pesquisa que buscou abordar a circulação das teorias feministas sobre a saúde da mulher, com a obra que se tornou best seller dentro e fora dos Estados Unidos na década de 1970 e manteve essa posição até o final dos anos 1980, Our Bodies, Ourselves. Davis, que é pesquisadora sênior do Research Institute for History and Culture na Utrecht University, na Holanda, analisa essa obra estadunidense de maneira crítica e com um distanciamento necessário. Sua proposta foi observar o modo como o livro "viajou" por diversos países-foi traduzido para mais de 30 idiomas-e as implicações dessa viagem na maneira de se pensarem o conhecimento feminista e as políticas de saúde em um mundo "globalizado". Ela percebe a produção e a recepção do livro como uma teoria que transita entre as especificidades de um contexto mundial e nos conta que Our Bodies, Ourselves surgiu da compilação das discussões sobre "as mulheres e seus corpos" em encontros pontuais que aconteceram em diversos países, no final dos anos 1960, promovidos pelo grupo Boston Women's Health Book Records-BWHBC. O resultado foi um manual com relatos de experiências pessoais e informações úteis a respeito da saúde das mulheres.

Forum Patrimonio Ambiente Construido E Patrimonio Sustentavel, Feb 9, 2013

Na engenharia e arquitetura, o escaneamento a laser de fachadas revolucionou a forma de se regist... more Na engenharia e arquitetura, o escaneamento a laser de fachadas revolucionou a forma de se registrarem informacoes topograficas e construtivas de detalhe. Scanners a laser (Laser Scanner – LS) podem capturar 12.000 pontos por segundo ou mais, sem a necessidade de refletores. Eles registram nao somente as dimensoes, mas tambem a localizacao espacial, a cor e a textura do objeto alvo. O sistema requer pouco tempo de processamento dos dados a posteriori e uma equipe de execucao minima. Por fim, o escaneamento a laser resulta, ao mesmo tempo, no levantamento fotografico metrico/dimensional e num modelo virtual de altissima precisao. Trata-se, portanto, de uma fotografia tridimensional metrificada. A maquete eletronica, uma vez manipulada nos softwares dedicados, gera inumeros mapas em diversos formatos. Alem do modelo digital, a tecnologia LS pode originar maquetes reais perfeitas, a partir de recursos de Engenharia Reversa (ER). Neste trabalho, objetivamos descrever o funcionamento da tecnologia de escaneamento a laser (LS), assim como provar a eficiencia da aplicacao desta tecnologia no mapeamento urbanistico e arquitetonico de edificacoes patrimoniais e espacos culturais, atraves de um estudo de caso por nos conduzido, juntamente com a empresa MAPTEK, no Espaco Cultural da Avenida Bernardo Monteiro, em Belo Horizonte – MG.

Cadernos Pagu, 2011

Catarina e bolsista da CAPES. Atualmente pesquisa sobre realizadoras de cinema do Brasil e da Arg... more Catarina e bolsista da CAPES. Atualmente pesquisa sobre realizadoras de cinema do Brasil e da Argentina durante as ditaduras militares nos dois países. amveiga@yahoo.com.br 1 Em 2010 as jornadas alcançaram sua terceira edição.

Revista Estudos Feministas, 2014

Bioinformatics (Oxford, England), Jan 28, 2015

The need for more effective and safer pharmaceuticals is a persistent quest. Microbial adaptation... more The need for more effective and safer pharmaceuticals is a persistent quest. Microbial adaptations create the need to permanently develop new antimicrobials, for instance. Similarly, intracellular delivery of drugs is still a challenge and translocation of membranes for drug delivery is an area of intense research. Peptides can be used both as antimicrobial drug leads and drug carrier systems for intracellular delivery. Multifunctional proteins are abundant in viruses but, surprisingly, have never been thoroughly screened for bioactive peptide sequences. Using the AMPA and CellPPD online tools we have evaluated the propensity of viral proteins to comprise antimicrobial (AMP) or cell-penetrating peptides (CPP). Capsid proteins from both enveloped and non-enveloped viruses, and membrane and envelope proteins from enveloped viruses, in a total of 272 proteins from 133 viruses, were screened to detect the presence of potential AMP and CPP sequences. A pool of 2444 and 426 CPP and AMP se...

Journal of Peptide Science, 2015

Despite the intensive study on the mechanism of action of membrane-active molecules such as antim... more Despite the intensive study on the mechanism of action of membrane-active molecules such as antimicrobial and anticancer peptides, most of the biophysical work has been performed using artificial model systems, mainly lipid vesicles. The use of these systems allows full control of the experimental parameters, and to obtain molecular-level detail on the action of peptides, the correlation with biological action is intangible. Recently, several biophysical methodologies have been translated to studies using bacterial and cancer cells. Here, we review biophysical studies on the mechanism of action of antimicrobial and anticancer peptides performed directly on cells. The data in these studies allow to correlate vesicle-based and cell-based studies and fill the vesicle-cell interdisciplinary gap. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.

Biochimica et Biophysica Acta (BBA) - Biomembranes, 2015

Recent patents on anti-infective drug discovery, 2006

The main strategies nowadays to fight AIDS rely on chemical therapy to inhibit the reverse transc... more The main strategies nowadays to fight AIDS rely on chemical therapy to inhibit the reverse transcriptase or protease of HIV. However, a synthetic 36 amino-acids peptide that blocks the entry of the virus in the target cells (enfuvirtide) has recently reached approval for clinical application. This molecule may probably be just the leader of a new generation of drugs that is about to emerge to interrupt the first step in the HIV life cycle, i.e. preventing the virus from actually entering cells. This paper reviews the enfuvirtide path from clinical trials to the attempts to detail its molecular-level mode of action. It is commonly accepted that this peptide would block the fusion between viral and cell plasma membrane through binding to the N-terminal heptad repeat (NHR) region of the viral protein gp41. However, there has been growing evidence that this model of action may be unrealistic, the action of enfuvirtide being more complex and diverse than initially thought. Membrane-assisted local concentration increase and interference with gp120/co-receptor docking may also contribute for the inhibitory action of the peptide. Selected HIV-entry inhibitors on clinical trials are presented to characterize the future drugs in the market in this class.

Significação: Revista de Cultura Audiovisual, 2013

Thrombosis Research, 2004

Envenomation caused by Lonomia obliqua caterpillars is an increasing problem in Southern Brazil. ... more Envenomation caused by Lonomia obliqua caterpillars is an increasing problem in Southern Brazil. The clinical profile is characterized by a profound hemorrhagic disorder. In the present study, we describe the characterization of a fibrin(ogen)olytic factor (lonofibrase) isolated from a venomous secretion of the caterpillars. The crude extract showed a dose-dependent inhibitory effect in the rate of thrombininduced fibrinogen clotting and produced fragmentation of fibrinogen. Isolation of the fibrin(ogen)olytic enzyme was achieved by combining ion exchange chromatography followed by gel filtration in a fast protein liquid chromatography (FPLC) system. A single 35-kDa band was identified and the isolated enzyme named lonofibrase. Lonofibrase rapidly degrades Aa and Bh chains of fibrinogen, also being able to cleave fibrin in a distinct way from that observed with plasmin. The presence of lonofibrase with both fibrinogenolytic and fibrinolytic activities in L. obliqua secretion is coherent with the severe hemorrhagic clinical profile resulting from envenomation caused by these insects.

FEBS Journal, 2013

Cell penetrating peptides (CPPs) can be used as drug delivery systems for different therapeutic m... more Cell penetrating peptides (CPPs) can be used as drug delivery systems for different therapeutic molecules. In this work two novel CPPs, pepR and pepM, designed from two domains of the dengue virus (DENV) capsid protein, were studied for their ability to deliver nucleic acids into cells as non-covalently bound cargo. Translocation studies were performed by confocal microscopy in HepG2, BHK and HEK cell lineages, astrocytes and peripheral blood mononuclear cells. Combined studies in HepG2 cells, astrocytes and BHK cells, at 4 and 37°C or using specific endocytosis inhibitors, revealed that pepR and pepM use distinct internalization routes: pepM translocates lipid membranes directly, while pepR uses an endocytic pathway. To confirm these results, a methodology was developed to monitor the translocation kinetics of both peptides by real-time flow cytometry. Kinetic constants were determined, and the amount of nucleic acids delivered was estimated. Additional studies were performed in order to understand the molecular bases of the peptide-mediated translocation. Peptide-nucleic acid and peptide-lipid membrane interactions were studied quantitatively based on the intrinsic fluorescence of the peptides. pepR and pepM bound ssDNA to the same extent. Partition studies revealed that both peptides bind preferentially to anionic lipid membranes, adopting an a-helical conformation. However, fluorescence quenching studies suggest that pepM is deeply inserted into the lipid bilayer, in contrast with pepR. Moreover, only pepM is able to promote the fusion and aggregation of vesicles composed of zwitterionic lipids. Altogether, the results show that DENV capsid protein derived peptides serve as good templates for novel CPP-based nucleic acid delivery strategies, defining different routes for cell entry.

FEBS Journal, 2007

A small amino acid sequence (LWYIK) inside the HIV-1 gp41 ectodomain membrane proximal region (MP... more A small amino acid sequence (LWYIK) inside the HIV-1 gp41 ectodomain membrane proximal region (MPR) is commonly referred to as a cholesterol-binding domain. To further study this unique and peculiar property we have used fluorescence spectroscopy techniques to unravel the membrane interaction properties of three MPR-derived synthetic peptides: the membrane proximal region peptide-complete (MPRP-C) which corresponds to the complete MPR; the membrane proximal region peptide-short (MPRP-S), which corresponds to the last five MPR amino acid residues (the putative cholesterol-binding domain) and the membrane proximal region peptide-intermediate (MPRP-I), which corresponds to the MPRP-C peptide without the MPRP-S sequence. MPRP-C and MPRP-I membrane interaction is largely independent of the membrane phase. Membrane interaction of MPRP-S occurs for fluid phase membranes but not in gel phase membranes or cholesterol-containing bilayers. The gp41 ectodomain MPR may have a very specific function in viral fusion through the concerted and combined action of cholesterol-binding and non-cholesterol-binding domains (i.e. domains corresponding to MPRP-S and MPRP-I, respectively).

PLoS ONE, 2013

Supercharged proteins are a recently identified class of proteins that have the ability to effici... more Supercharged proteins are a recently identified class of proteins that have the ability to efficiently deliver functional macromolecules into mammalian cells. They were first developed as bioengineering products, but were later found in the human proteome. In this work, we show that this class of proteins with unusually high net positive charge is frequently found among viral structural proteins, more specifically among capsid proteins. In particular, the capsid proteins of viruses from the Flaviviridae family have all a very high net charge to molecular weight ratio (. +1.07/kDa), thus qualifying as supercharged proteins. This ubiquity raises the hypothesis that supercharged viral capsid proteins may have biological roles that arise from an intrinsic ability to penetrate cells. Dengue virus capsid protein was selected for a detailed experimental analysis. We showed that this protein is able to deliver functional nucleic acids into mammalian cells. The same result was obtained with two isolated domains of this protein, one of them being able to translocate lipid bilayers independently of endocytic routes. Nucleic acids such as siRNA and plasmids were delivered fully functional into cells. The results raise the possibility that the ability to penetrate cells is part of the native biological functions of some viral capsid proteins.

Journal of Viral Hepatitis, 2008

We have analysed and identified different membrane-active regions of the Hepatitis C virus (HCV) ... more We have analysed and identified different membrane-active regions of the Hepatitis C virus (HCV) core protein by observing the effect of 18-mer core-derived peptide libraries from two HCV strains on the integrity of different membrane model systems. In addition, we have studied the secondary structure of specific membrane-interacting peptides from the HCV core protein, both in aqueous solution and in the presence of model membrane systems. Our results show that the HCV core protein region comprising the C-terminus of domain 1 and the N-terminus of domain 2 seems to be the most active in membrane interaction, although a role in protein-protein interaction cannot be excluded. Significantly, the secondary structure of nearly all the assayed peptides changes in the presence of model membranes. These sequences most probably play a relevant part in the biological action of HCV in lipid interaction. Furthermore, these membranotropic regions could be envisaged as new possible targets, as inhibition of its interaction with the membrane could potentially lead to new vaccine strategies.

Journal of the American Chemical Society, 2012

Several cationic antimicrobial peptides (AMPs) have recently been shown to display anticancer act... more Several cationic antimicrobial peptides (AMPs) have recently been shown to display anticancer activity via a mechanism that usually entails the disruption of cancer cell membranes. In this work, we designed an 18-residue anticancer peptide, SVS-1, whose mechanism of action is designed to take advantage of the aberrant lipid composition presented on the outer leaflet of cancer cell membranes, which makes the surface of these cells relatively electronegative relative to noncancerous cells. SVS-1 is designed to remain unfolded and inactive in aqueous solution but preferentially fold at the surface of cancer cells, adopting an amphiphilic β-hairpin structure capable of membrane disruption. Membrane-induced folding is driven by electrostatic interaction between the peptide and the negatively charge membrane surface of cancer cells. SVS-1 is active against a variety of cancer cell lines such as A549 (lung carcinoma), KB (epidermal carcinoma), MCF-7 (breast carcinoma) and MDA-MB-436 (breast carcinoma). However, the cytotoxicity towards non-cancerous cells having typical membrane compositions, such as HUVEC and erythrocytes, is low. CD spectroscopy, appropriately designed peptide controls, cell-based studies, liposome leakage assays and electron microscopy support the intended mechanism of action, which leads to preferential killing of cancerous cells.

Journal of Peptide Science, 2013

One of the major challenges in the drug development process is biodistribution across epithelia a... more One of the major challenges in the drug development process is biodistribution across epithelia and intracellular drug targeting. Cellular membrane heterogeneity is one of the major drawbacks in developing efficient and sustainable drug delivery systems, which brings the need to study their interaction with lipids in order to unravel their mechanisms of action and improve their delivery capacities. Cell penetrating peptides (CPPs) are able to translocate almost any cell membrane carrying cargo molecules. However, different CPP use different entry mechanisms, which are often concentration-dependent and cargo-dependent. Being able to quantify the lipid affinity of CPP is of obvious importance and can be achieved by studying the partition extent of CPP into lipid bilayers. The partition constant (K p) reflects the lipid-water partition extent. However, all currently available methodologies are only suitable to determine the partition of single molecules into lipid membranes or entities, being unsuitable to determine the partition of bimolecular or higher order supramolecular complexes. We derived and tested a mathematical model to determine the K p of supramolecular CPP-cargo complexes from fluorescence spectroscopy data, using DNA oligomers as a model cargo. As a proof-of-concept example, the partition extent of two new membrane active peptides derived from dengue virus capsid protein (DENV C protein) with potential CPP properties, in both scenarios (free peptide and complexed with a molecular cargo), were tested. We were able to identify the lipid affinity of these CPP:DNA complexes, thus gaining valuable insights into better CPP formulations.

Journal of Molecular Structure, 2014

h i g h l i g h t s Antibody therapeutic interest has grown in the past decade. Lipid membranes m... more h i g h l i g h t s Antibody therapeutic interest has grown in the past decade. Lipid membranes may play an important role in epitope recognition. Fluorescence spectroscopy can be used to monitor antibody-membrane interactions. Its cost-effectiveness makes it a potential alternative to other techniques.

IUBMB Life, 2014

The current landscapes of novel therapeutic approaches rely mostly on gene-targeted technologies,... more The current landscapes of novel therapeutic approaches rely mostly on gene-targeted technologies, enabling to fight rare genomic diseases, from infections to cancer and hereditary diseases. Although, reaching the action-site for this novel treatments requires to deliver nucleic acids, or other macromolecules into cells, which may pose difficult tasks to pharmaceutical companies. To overcome this technological limitation, a wide variety of vectors have been developed in the past decades and have proven to be successful in delivering various therapeutics. Cell-penetrating peptides (CPP) have been one of the technologies widely studied and have been increasingly used to transport small RNA/DNA, plasmids, antibodies, and nanoparticles into cells. Despite the already proved huge potential that these peptide-based approaches may suggest, few advances have been put to pharmacological or clinical use. This review will describe the origin, development, and usage of CPP to deliver therapeutic agents into cells, with special emphasis on their current application to gene-therapies. Specifically, we will describe the current trials being conducted to treat cancer, gene disorders, and autoimmune diseases using CPP-based therapies.

European Biophysics Journal, 2011

Many cellular phenomena occur on the biomembranes. There are plenty of molecules (natural or xeno... more Many cellular phenomena occur on the biomembranes. There are plenty of molecules (natural or xenobiotics) that interact directly or partially with the cell membrane. Biomolecules, such as several peptides (e.g., antimicrobial peptides) and proteins, exert their effects at the cell membrane level. This feature makes necessary investigating their interactions with lipids to clarify their mechanisms of action and side effects necessary. The determination of molecular lipid/water partition constants (K p) is frequently used to quantify the extension of the interaction. The determination of this parameter has been achieved by using different methodologies, such as UV-Vis absorption spectrophotometry, fluorescence spectroscopy and f-potential measurements. In this work, we derived and tested a mathematical model to determine the K p from f-potential data. The values obtained with this method were compared with those obtained by fluorescence spectroscopy, which is a regular technique used to quantify the interaction of intrinsically fluorescent peptides with selected biomembrane model systems. Two antimicrobial peptides (BP100 and pepR) were evaluated by this new method. The results obtained by this new methodology show that f-potential is a powerful technique to quantify peptide/lipid interactions of a wide variety of charged molecules, overcoming some of the limitations inherent to other techniques, such as the need for fluorescent labeling. Keywords Partition constant Á Zeta-potential Á Fluorescence Á Membrane Á Antimicrobial peptides Membrane-active peptides: 455th WE-Heraeus-Seminar and AMP 2010 Workshop.

Revista Estudos Feministas, 2006

Revista Estudos Feministas, 2009

A historiadora holandesa Kathy Davis compartilha no livro How Feminism Travels across Borders, de... more A historiadora holandesa Kathy Davis compartilha no livro How Feminism Travels across Borders, de maneira autêntica e singular, os resultados de uma ampla pesquisa que buscou abordar a circulação das teorias feministas sobre a saúde da mulher, com a obra que se tornou best seller dentro e fora dos Estados Unidos na década de 1970 e manteve essa posição até o final dos anos 1980, Our Bodies, Ourselves. Davis, que é pesquisadora sênior do Research Institute for History and Culture na Utrecht University, na Holanda, analisa essa obra estadunidense de maneira crítica e com um distanciamento necessário. Sua proposta foi observar o modo como o livro "viajou" por diversos países-foi traduzido para mais de 30 idiomas-e as implicações dessa viagem na maneira de se pensarem o conhecimento feminista e as políticas de saúde em um mundo "globalizado". Ela percebe a produção e a recepção do livro como uma teoria que transita entre as especificidades de um contexto mundial e nos conta que Our Bodies, Ourselves surgiu da compilação das discussões sobre "as mulheres e seus corpos" em encontros pontuais que aconteceram em diversos países, no final dos anos 1960, promovidos pelo grupo Boston Women's Health Book Records-BWHBC. O resultado foi um manual com relatos de experiências pessoais e informações úteis a respeito da saúde das mulheres.