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  • ItemCapítulo de livro
    Application of Rare Earth Cation-Exchanged Nanozeolite as a Support for the Immobilization of Fungal Lipase and their Use in Biodiesel Production
    (2021-01-01) de Paula Guarnieri, Guilherme [UNESP]; de Vasconcellos, Adriano [UNESP]; de Moraes, Fábio Rogério [UNESP]; Nery, José Geraldo [UNESP]; Universidade Estadual Paulista (UNESP)
    For the last 50 years, zeolite ions exchanged with lanthanum (La3+) and cerium (Ce3+) rare earth cations have mostly been used as fluid catalytic cracking catalysts (FCCs) in the petrochemical industries for the production of fuels. However, environmental concerns and restrictive legislation have led researchers to develop clean technologies for energy production of biofuels such as biodiesel from renewable sources. In this context, a new class of biocatalysts composed of nanozeolite ions exchanged with rare earth cations (La3+, Ce3+, Gd3+, Dy3+) and fungal lipases such as Thermomyces lanuginosus lipase (TLL) and Rhizomucor miehei lipase (RML) has been applied as heterogeneous catalysts for the transesterification of non-edible microalgae oils to fatty acid ethyl esters (FAEEs). Experimental results indicate that TLL immobilized on rare earth nanozeolitic supports showed more activity in comparison with RML. Considering these facts, this chapter looks at such parameters as amount of immobilized enzyme, enzymatic activity, and FAEE yield, along with bioinformatics approaches and the different behaviors of both nanozeolite–enzymes complexes (TLL and RML).
  • ItemArtigo
    Brown Spider Venom Phospholipase-D Activity upon Different Lipid Substrates
    (2023-02-01) Chaves-Moreira, Daniele; Gremski, Luiza Helena; de Moraes, Fábio Rogério [UNESP]; Vuitika, Larissa; Wille, Ana Carolina Martins; Hernández González, Jorge Enrique [UNESP]; Chaim, Olga Meiri; Senff-Ribeiro, Andrea; Arni, Raghuvir Krishnaswamy [UNESP]; Veiga, Silvio Sanches; Universidade Federal do Paraná (UFPR); Universidade Estadual Paulista (UNESP); Universidade Estadual de Ponta Grossa (UEPG); University of California San Diego
    Brown spider envenomation results in dermonecrosis, characterized by an intense inflammatory reaction. The principal toxins of brown spider venoms are phospholipase-D isoforms, which interact with different cellular membrane components, degrade phospholipids, and generate bioactive mediators leading to harmful effects. The Loxosceles intermedia phospholipase D, LiRecDT1, possesses a loop that modulates the accessibility to the active site and plays a crucial role in substrate. In vitro and in silico analyses were performed to determine aspects of this enzyme’s substrate preference. Sphingomyelin d18:1/6:0 was the preferred substrate of LiRecDT1 compared to other Sphingomyelins. Lysophosphatidylcholine 16:0/0:0 was preferred among other lysophosphatidylcholines, but much less than Sphingomyelin d18:1/6:0. In contrast, phosphatidylcholine d18:1/16:0 was not cleaved. Thus, the number of carbon atoms in the substrate plays a vital role in determining the optimal activity of this phospholipase-D. The presence of an amide group at C2 plays a key role in recognition and activity. In silico analyses indicated that a subsite containing the aromatic residues Y228 and W230 appears essential for choline recognition by cation-π interactions. These findings may help to explain why different cells, with different phospholipid fatty acid compositions exhibit distinct susceptibilities to brown spider venoms.
  • ItemArtigo
    Exploring the Folding Mechanism of Dimeric Superoxide Dismutase
    (2023-02-16) Mouro, Paulo R. [UNESP]; Sanches, Murilo N. [UNESP]; Leite, Vitor B. P. [UNESP]; Chahine, Jorge [UNESP]; Universidade Estadual Paulista (UNESP)
    The Cu/Zn Human Superoxide Dismutase (SOD1) is a dimeric metalloenzyme whose genetic mutations are directly related to amyotrophic lateral sclerosis (ALS), so understanding its folding mechanism is of fundamental importance. Currently, the SOD1 dimer formation is studied via molecular dynamics simulations using a simplified structure-based model and an all-atom model. Results from the simplified model reveal a mechanism dependent on distances between monomers, which are limited by constraints to mimic concentration dependence. The stability of intermediates (during the int state) is significantly affected by this distance, as well as by the presence of two folded monomers prior to dimer formation. The kinetics of interface formation are also highly dependent on the separation distance. The folding temperature of the dimer is about 4.2% higher than that of the monomer, a value not too different from experimental data. All-atom simulations on the apo dimer give binding free energy between monomers similar to experimental values. An intermediate state is evident for the apo form at a separation distance between monomers slightly larger than the native distance which has little formed interface between monomers. We have shown that this intermediate is stabilized by non-native intra- and intercontacts.
  • ItemArtigo
    Immunoprophylactic properties of the Corynebacterium pseudotuberculosis-derived MBP:PLD:CP40 fusion protein
    (2022-12-01) Barral, Thiago Doria; Kalil, Mauricio Alcantara; Mariutti, Ricardo Barros [UNESP]; Arni, Raghuvir Krishnaswamy [UNESP]; Gismene, Carolina [UNESP]; Sousa, Fernanda Severo; Collares, Tiago; Seixas, Fabiana Kommling; Borsuk, Sibele; Estrela-Lima, Alessandra; Azevedo, Vasco; Meyer, Roberto; Portela, Ricardo Wagner; Universidade Federal da Bahia (UFBA); Universidade Estadual Paulista (UNESP); Universidade Federal de Pelotas; Universidade Federal de Minas Gerais (UFMG)
    Abstract: Caseous lymphadenitis (CLA) is a disease that affects small ruminants, and the best way to prevent its spread on a herd is through immunoprophylaxis. Thus, we aimed to evaluate the MBP:PLD:CP40 fusion protein as a new CLA immunogen. The fusion protein was constructed by combining Corynebacterium pseudotuberculosis PLD and CP40 proteins with maltose-binding protein (MBP) as an intrinsic adjuvant. The antigenicity, allergenic potential, prediction of B epitopes, binding to MHC receptors, and docking on the Toll-Like 2 receptor were evaluated in silico. MBP:PLD:CP40 was expressed and purified. 40 BALB/c were divided into four groups (G1 – control, G2 – Saponin, G3 – MBP:PLD:CP40, and G4 – rPLD + rCP40). Total IgG, IgG1, and IgG2a were quantified, and the expressions of cytokines after splenocyte in vitro stimulation were assessed. Mice were challenged 42 days after the first immunization. The in silico analysis showed that MBP:PLD:CP40 has immunogenic potential, does not have allergic properties, and can dock on the TRL2 receptor. MBP:PLD:CP40 stimulated the production of IgG1 antibodies in a fivefold proportion to IgG2a, and TNF and IL-17 were significantly expressed in response to the antigenic stimuli. When rPLD and rCP40 were used together for immunization, they could induce IFN-γ and IL-12, but with no detectable antibody production. The G3 and G4 groups presented a survival of 57.14% and 42.86%, respectively, while the G1 and G2 mice were all dead 15 days after the challenge. MBP:PLD:CP40 partially protected the mice against C. pseudotuberculosis infection and can be considered a potential new CLA immunogen. Key points: • The fusion protein induced more IgG1 than IgG2a antibodies; • The fusion protein also induced the expression of the TNF and IL-17 cytokines; • Mice inoculated with MBP:PLD:CP40 presented a 57.14% survival.
  • ItemArtigo
    Probability distribution to obtain the characteristic passage time for different tri-stable potentials
    (2022-11-15) Drigo Filho, Elso [UNESP]; Chahine, Jorge [UNESP]; Araujo, Marcelo Tozo; Ricotta, Regina Maria; Universidade Estadual Paulista (UNESP); UNILAGO; CEETEPS
    The purpose of this work is to explore the kinetics of the transition probability distribution obtained as a solution to the Fokker–Planck equation (FPE) for a system described by a potential function (free energy) that has three regions of minimum, a tri-stable potential. The Fokker–Planck equation is rewritten as a Schrödinger equation (SE) and this allows the use of the Supersymmetric Quantum Mechanics formalism (SQM) associated to the variational method to obtain the analytical approximated spectrum. From the solutions obtained the probability distribution is evaluated which allows the determination of the characteristic passage time between the three potential minima. The results show the dependence between the diffusion process and the relative depth between of the central and the lateral minima of the potential.
  • ItemArtigo
    Kinetic and structural studies of Mycobacterium tuberculosis dihydroorotate dehydrogenase reveal new insights into class 2 DHODH inhibition
    (2023-07-01) Teixeira, Olívia; Martins, Ingrid Bernardes Santana [UNESP]; Froes, Thamires Quadros; de Araujo, Alexandre Suman [UNESP]; Nonato, Maria Cristina; Universidade de São Paulo (USP); Universidade Federal do Rio de Janeiro (UFRJ); Universidade Estadual Paulista (UNESP)
    Tuberculosis (TB) is a leading cause of death worldwide. TB represents a serious public health threat, and it is characterized by high transmission rates, prevalence in impoverished regions, and high co-infection rates with HIV. Moreover, the serious side effects of long-term treatment that decrease patient adherence, and the emergence of multi-resistant strains of Mycobacterium tuberculosis, the causing agent of TBs, pose several challenges for its eradication. The search for a new TB treatment is necessary and urgent. Dihydroorotate dehydrogenase (DHODH) is responsible for the stereospecific oxidation of (S)-dihydroorotate (DHO) to orotate during the fourth and only redox step of the de novo pyrimidine nucleotide biosynthetic pathway. DHODH has been considered an attractive target against infectious diseases. As a first step towards exploiting DHODH as a drug target against TB, we performed a full kinetic characterization of both bacterial MtDHODH and its human ortholog (HsDHDOH) using both substrates coenzyme Q0 (Q0) and vitamin K3 (K3). MtDHODH follows a ping-pong mechanism of catalysis and shares similar catalytic parameters with the human enzyme. Serendipitously, Q0 was found to inhibit MtDHODH (KI (Q0) = 138 ± 31 μM). To the best of our knowledge, Q0 is the first non-orotate like dihydroorotate-competitive inhibitor for class 2 DHODHs ever described. Molecular dynamics simulations along with in silico solvent mapping allowed us to successfully probe protein flexibility and correlate it with the druggability of binding sites. Together, our results provide the starting point for the design of a new generation of potent and selective inhibitors against MtDHODH.
  • ItemArtigo
    Out-of-plane deformability and its coupling with electrostatics in biomembranes
    (2023-03-31) Wilke, Natalia; Alvares, Dayane S. [UNESP]; Universidad Nacional de Córdoba; Universidade Estadual Paulista (UNESP)
    Cell membranes are quasi-bidimensional soft systems formed by multipoles in an ordered array that can be polarized in an electric field. Consequently, electrostatic potentials emerge inside membranes, and membranes respond to external electric fields. From a mechanical perspective, membranes can be easily compressed-expanded, laterally deformed, and curved. Bending is particularly easy, and this kind of deformation translates to changes in the relative positions of the negative and positive charges, leading to strain gradient-induced polarization. Conversely, an external electric field gradient will exert a bending stress that translates to mechanical membrane deformation. These phenomena are described through membrane flexoelectricity. Here, we describe this property in lipid bilayers and cell membranes and summarize the studies in the field with emphasis on the effects promoted by membrane asymmetry.
  • ItemArtigo
    Electrochemical sensor based on epoxy-functionalized BEA nanozeolite and graphene oxide modified glassy carbon electrode for bisphenol E determination
    (2023-01-01) Miller, Alex H. [UNESP]; Nguyen, Huong Thi-Thanh; Nery, José G. [UNESP]; Fielding, Alistair J.; Liverpool John Moores University; Universidade Estadual Paulista (UNESP); University of York
    An epoxy-functionalized beta type nanozeolite (BEA)/graphene oxide nanocomposite modified glassy carbon electrode (GCE/BEA/APTMS/GA/GO/NF) has been created for the differential pulse voltammetric determination of bisphenol E (BPE). The modified electrode presented an enhanced current response in comparison with bare GCE. A linear dependence of anodic peak current (Ip) and scan rate (ν) was observed, which showed that the electrochemical process was adsorption-controlled. Differential pulse voltammetry (DPV) was employed and optimized for the sensitive determination of BPE. Under the optimized conditions, the anodic peak current was linearly proportional to BPE concentration in the range between 0.07 and 4.81 µM, with a correlation coefficient of 0.995 and limit of detection 0.056 μM (S/N = 3). The electrode showed good repeatability and storage stability, and a low response to interfering compounds. Comparison was made to the determination of bisphenol A. To confirm the electrode analytical performance, recovery tests were performed, and deviations lower than 10% were found. The BEA zeolite-GO nanocomposite proved to be a promising sensing platform for bisphenol determination. Graphical abstract: [Figure not available: see fulltext.].
  • ItemArtigo
    Self-assembly of dengue virus empty capsid-like particles in solution
    (2023-03-17) Neves-Martins, Thais C.; Mebus-Antunes, Nathane C.; Neto, Carlos H.G.; Barbosa, Glauce M.; Almeida, Fabio C.L.; Caruso, Icaro P. [UNESP]; Da Poian, Andrea T.; Universidade Federal do Rio de Janeiro (UFRJ); Universidade Estadual Paulista (UNESP)
    Nucleocapsid (NC) assembly is an essential step of the virus replication cycle. It ensures genome protection and transmission among hosts. Flaviviruses are human viruses for which envelope structure is well known, whereas no information on NC organization is available. Here we designed a dengue virus capsid protein (DENVC) mutant in which a highly positive spot conferred by arginine 85 in α4-helix was replaced by a cysteine residue, simultaneously removing the positive charge and restricting the intermolecular motion through the formation of a disulfide cross-link. We showed that the mutant self-assembles into capsid-like particles (CLP) in solution without nucleic acids. Using biophysical techniques, we investigated capsid assembly thermodynamics, showing that an efficient assembly is related to an increased DENVC stability due to α4/α4′ motion restriction. To our knowledge, this is the first time that flaviviruses’ empty capsid assembly is obtained in solution, revealing the R85C mutant as a powerful tool to understand the NC assembly mechanism.
  • ItemArtigo
    The influence of pH on the structure and stability of the Grb2 dimer reveals changes in the inter-domain and molecular interaction: Could it be a modulation mechanism?
    (2023-04-01) Tedesco, Jéssica A. [UNESP]; Dias, Raphael V.R. [UNESP]; Casteluci, Giovana [UNESP]; Pedro, Renan P. [UNESP]; de Oliveira, Leandro C. [UNESP]; Caruso, Ícaro P. [UNESP]; Melo, Fernando A. [UNESP]; Universidade Estadual Paulista (UNESP); Universidade Federal do Rio de Janeiro (UFRJ)
    Cancer cells present an increased replicative potential as a hallmark. The increased replication leads to a higher intracellular pH. Grb2, an adapter protein, is mainly involved in several types of cancers due to its role in signaling pathways responsible for cell growth and proliferation. At pH 7, we observed a more compact structure, as seen by DLS and 1H NMR relaxation experiments, with high cooperativity within domains. On the other hand, we observed an increase in disordered structures at pH 8, with relative independence between domains characterized by higher melting temperatures and enthalpy of unfolding. CD and DLS corroborate with these observations at pH 8, conferring more flexibility among the domains, followed by lower unfolding cooperativity and increased hydrodynamic diameter at higher pH. In addition, 15N-HSQC chemical shift perturbations experiments showed significant differences in the positions of several amino acids spread on the Grb2 structure when pH was changed, which agrees with the previous results. Finally, the molecular dynamic analysis demonstrates that Grb2 presents a movement pattern where both SH3 domains move toward the center of the protein at pH 7. On the contrary, the pattern changes its direction at pH 8, where domains move outside the center of the protein, conferring a more elongated structure at higher pH. So, Grb2 presents significant structural and dynamic changes modulated by pH. If considering the role of Grb2 in cell signaling upstream, these conformational changes could be a critical mechanistic behavior of this protein, preventing/disrupting the stability of the cell signaling pathways related to cancer.
  • ItemArtigo
    An adsorption agent based on chitosan-zeolite composite: environmental and radioactive liquid waste remediation
    (2023-01-30) de Araujo, Leandro Goulart; Medeiros, Vinicius Litrenta [UNESP]; Guarnieri, Guilherme de Paula [UNESP]; da Silva, Danilo Antonio [UNESP]; Watanabe, Tamires; Marumo, Júlio Takehiro; Nery, José Geraldo [UNESP]; Nuclear and Energy Research Institute (IPEN); Universidade Estadual Paulista (UNESP)
    In this article, we present a chitosan-zeolite composite, which was synthesized and used as an adsorbent material for caesium (Cs) removal from aqueous media and real liquid radioactive organic waste (LROW). The compound was characterized by X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron microscopy techniques. The physicochemical characterization indicates the production of a composite. Adsorption experiments were first performed using the prepared solutions contaminated with Cs using full factorial design with two variables of interest: initial Cs concentration (Cs0) and adsorbent dosage (mg L−1). The results indicated a high caesium removal rate with removal values above 93% and adsorption capacity of up to 10 mg g−1. With the best experimental conditions according to our experimental domain, time was evaluated and equilibrium was reached in 180 min. Finally, the adsorbent material was tested as an adsorbent for Cs, Am, and U from LROW. When in contact with LORW, the removal rates (%) were 21.51 (137Cs), 26.39 (241Am), and 20.26 (U (total)). Although lower, this material indicated that it has the potential to be used for multi-elemental adsorption.
  • ItemArtigo
    Interphase chromosomes of the Aedes aegypti mosquito are liquid crystalline and can sense mechanical cues
    (2023-12-01) Contessoto, Vinícius G. [UNESP]; Dudchenko, Olga; Aiden, Erez Lieberman; Wolynes, Peter G.; Onuchic, José N.; Di Pierro, Michele; Rice University; Universidade Estadual Paulista (UNESP); Baylor College of Medicine; Northeastern University
    We use data-driven physical simulations to study the three-dimensional architecture of the Aedes aegypti genome. Hi-C maps exhibit both a broad diagonal and compartmentalization with telomeres and centromeres clustering together. Physical modeling reveals that these observations correspond to an ensemble of 3D chromosomal structures that are folded over and partially condensed. Clustering of the centromeres and telomeres near the nuclear lamina appears to be a necessary condition for the formation of the observed structures. Further analysis of the mechanical properties of the genome reveals that the chromosomes of Aedes aegypti, by virtue of their atypical structural organization, are highly sensitive to the deformation of the nuclei. This last finding provides a possible physical mechanism linking mechanical cues to gene regulation.
  • ItemArtigo
    Syntheses of novel topical hemostatic agents based on pectin biopolymer aiming hemorrhage control: Thromboelastographic studies and non-homogeneous swelling effect
    (2022-12-01) Santisteban, Oscar A.N. [UNESP]; Sacco, Guilherme M. [UNESP]; Guarnieri, Guilherme De Paula [UNESP]; Godoy, Moacir Fernandes de; Azevedo, Eduardo Ribeiro de; Nery, José Geraldo [UNESP]; Universidade Estadual Paulista (UNESP); School of Medicine of São José do Rio Preto; Universidade de São Paulo (USP)
    Topical hemostatic agents based on pectin biopolymer were synthesized and characterized by X-ray diffraction (XRD), Scanning Electronic Microscopy (SEM), Phase Contrast Microscopy, Infrared spectroscopy (FT-IR), Differential Scanning Calorimetry (DSC) and Solid State 13C MAS NMR. The pectin biofilms were crosslinked with CaCl2 using different concentrations of Ca2+ solutions ( 4.5 ×10-2 M, 2.7 ×102 M, 4.5 ×102 M). 13C MAS NMR and FT-IR data revealed that crosslinking with Ca2+ was successfully achieved, in line with the formation of egg-box type structures. The anti-hemorragic effect caused by the interaction of the red blood cells with the pectin biofilm interface was showed in details by phase contrast microscopy. The images showed that higher Ca2+ concentration (0.45 ×102 M) positively favor the coagulation cascade by facilitating the migration and an orderly stacking of the red blood cells into the pectin biofilm scaffold. The hemostatic action of the prepared pectin biofilms were analyzed by Thromboelastography (TEG). The results indicated that the hemostatic action of the biofilms is as effective as that of the hybrid materials reported in the literature, indicating that these pectin biofilms have potential to become safe topic hemostat. It was also observed a non-homogeneous swelling effect caused by the interaction between the pectin biofilms interface and blood cells which induced a rapid bending and a bucking instability that swollen the pectin network.
  • ItemArtigo
    Transcriptomic Profiling of Rectus Abdominis Muscle in Women with Gestational Diabetes-Induced Myopathy: Characterization of Pathophysiology and Potential Muscle Biomarkers of Pregnancy-Specific Urinary Incontinence
    (2022-11-01) Alves, Fernanda Cristina Bergamo [UNESP]; Oliveira, Rafael Guilen de [UNESP]; Reyes, David Rafael Abreu [UNESP]; Garcia, Gabriela Azevedo [UNESP]; Floriano, Juliana Ferreira [UNESP]; Shetty, Raghavendra Hallur Lakshmana [UNESP]; Mareco, Edson Assunção; Dal-Pai-Silva, Maeli [UNESP]; Payão, Spencer Luiz Marques; Souza, Fátima Pereira de [UNESP]; Witkin, Steven S.; Sobrevia, Luis [UNESP]; Barbosa, Angélica Mércia Pascon [UNESP]; Rudge, Marilza Vieira Cunha [UNESP]; Universidade Estadual Paulista (UNESP); Pravara Institute of Medical Sciences; University of Western São Paulo (UNOESTE); Faculdade de Medicina de Marília (FAMEMA); Weill Cornell Medicine; Universidade de São Paulo (USP); Pontificia Universidad Católica de Chile; Universidad de Sevilla; University of Queensland; University of Groningen; School of Medicine and Health Sciences
    Gestational diabetes mellitus (GDM) is recognized as a “window of opportunity” for the future prediction of such complications as type 2 diabetes mellitus and pelvic floor muscle disorders, including urinary incontinence and genitourinary dysfunction. Translational studies have reported that pelvic floor muscle disorders are due to a GDM-induced-myopathy (GDiM) of the pelvic floor muscle and rectus abdominis muscle (RAM). We now describe the transcriptome profiling of the RAM obtained by Cesarean section from GDM and non-GDM women with and without pregnancy-specific urinary incontinence (PSUI). We identified 650 genes in total, and the differentially expressed genes were defined by comparing three control groups to the GDM with PSUI group (GDiM). Enrichment analysis showed that GDM with PSUI was associated with decreased gene expression related to muscle structure and muscle protein synthesis, the reduced ability of muscle fibers to ameliorate muscle damage, and the altered the maintenance and generation of energy through glycogenesis. Potential genetic muscle biomarkers were validated by RT-PCR, and their relationship to the pathophysiology of the disease was verified. These findings help elucidate the molecular mechanisms of GDiM and will promote the development of innovative interventions to prevent and treat complications such as post-GDM urinary incontinence.
  • ItemArtigo
    Structure-Function Relationship in Heterodimeric Neurotoxin PLA2s from Viperidae Snakes Inhabiting Europe, South America, and Asia Functional Importance of the Nontoxic Components
    (Springer, 2016-01-01) Georgieva, Dessislava; Arni, Raghuvir K. [UNESP]; Betzel, Christian; Calvete, J. J.; Gopalakrishnakone, P.; Univ Hamburg; Universidade Estadual Paulista (UNESP)
    Snake venom heterodimeric non-covalent phospholipase A(2) (PLA(2)) complexes (ncHdPLA(2)s) are neurotoxins encountered in the venoms of Viperinae and Crotalinae snakes. In contrast to their monomeric counterparts, they have a sophisticated mechanism of action in order to avoid nonspecific binding to wrong targets and to increase the efficiency of the pharmacological attack. ncHdPLA(2)s consist of a toxic and enzymatically active basic PLA(2) and an acidic and catalytically inactive PLA(2) protein. The main function of the acidic subunit is to direct the toxic component to the correct targets on the cell membrane and in this way to avoid binding to phospholipids, which are not important for the pharmacological effect of the toxin. The nontoxic component of ncHdPLA(2)s is multifunctional. It can modulate the toxicity and catalytic activity of PLA(2). The structure and function of ncHdPLA(2)s from the venoms of snakes inhabiting Europe, South America, and Asia are similar. There exists a high identity at the levels of primary and three-dimensional structures of ncHdPLA(2)s from snakes inhabiting widely separated regions of the world. Although a substantial progress has been made during the last years in understanding the structure and biological action of ncHdPLA(2)s, a number of questions still remain to be answered.
  • ItemArtigo
    Three-Dimensional Structures and Mechanisms of Snake Venom Serine Proteinases, Metalloproteinases, and Phospholipase A(2)s
    (Springer, 2016-01-01) Coronado, M. A. [UNESP]; Moraes, F. R. de [UNESP]; Ullah, A. [UNESP]; Masood, R. [UNESP]; Santana, V. S. [UNESP]; Mariutti, R. [UNESP]; Brognaro, H. [UNESP]; Georgieva, Dessislava; Murakami, M. T.; Betzel, Christian; Arni, Raghuvir K. [UNESP]; Calvete, J. J.; Gopalakrishnakone, P.; Universidade Estadual Paulista (UNESP); Univ Hamburg; Natl Ctr Res Energy & Mat
    High-resolution crystal structures provide accurate information of the positions of the atoms which can be used to understand substrate specificity, secondary binding sites, and catalytic mechanisms. Detailed structural information and mechanisms of serine proteinases, metalloproteinases, and phospholipases A(2)s are presented.
  • ItemArtigo
    A Computational–Experimental Investigation of the Molecular Mechanism of Interleukin-6-Piperine Interaction
    (2022-07-01) Povinelli, Ana Paula Ribeiro; Zazeri, Gabriel; Jones, Alan M.; Cornélio, Marinônio Lopes [UNESP]; Science and Technology of Mato Grosso; University of Birmingham; Universidade Estadual Paulista (UNESP)
    Herein, we elucidate the biophysical aspects of the interaction of an important protein, Interleukin-6 (IL6), which is involved in cytokine storm syndrome, with a natural product with anti-inflammatory activity, piperine. Despite the role of piperine in the inhibition of the transcriptional protein NF-κB pathway responsible for activation of IL6 gene expression, there are no studies to the best of our knowledge regarding the characterisation of the molecular interaction of the IL6-piperine complex. In this context, the characterisation was performed with spectroscopic experiments aided by molecular modelling. Fluorescence spectroscopy alongside van’t Hoff analyses showed that the complexation event is a spontaneous process driven by non-specific interactions. Circular dichroism aided by molecular dynamics revealed that piperine caused local α-helix reduction. Molecular docking and molecular dynamics disclosed the microenvironment of interaction as non-polar amino acid residues. Although piperine has three available hydrogen bond acceptors, only one hydrogen-bond was formed during our simulation experiments, reinforcing the major role of non-specific interactions that we observed experimentally. Root mean square deviation (RMSD) and hydrodynamic radii revealed that the IL6-piperine complex was stable during 800 ns of simulation. Taken together, these results can support ongoing IL6 drug discovery efforts.
  • ItemArtigo
    Biophysical studies of the interaction of hRSV Non-Structural 1 protein with natural flavonoids and their acetylated derivatives by spectroscopic techniques and computational simulations
    (2022-12-15) Ottenio de Lourenço, Isabella [UNESP]; Toscano Pedroso Quintino, Evelyn [UNESP]; Henrique Pereira, Matheus [UNESP]; Sprengel Lima, Caroline [UNESP]; Campos Araújo, Gabriela [UNESP]; Octávio Regasini, Luis [UNESP]; Alves de Melo, Fernando [UNESP]; Pereira de Souza, Fátima [UNESP]; Andres Fossey, Marcelo [UNESP]; Putinhon Caruso, Ícaro [UNESP]; Universidade Estadual Paulista (UNESP); Federal University of Rio de Janeiro
    Human respiratory syncytial virus (hRSV) infections are one of the most causes of acute lower respiratory tract infections in children and elderly. The development of effective antiviral therapies or preventive vaccines against hRSV is not available yet. Thus, it is necessary to search for protein targets to combat this viral infection, as well as potential ways to block them. Non-Structural 1 (NS1) protein is an important factor for viral replication success since reduces the immune response by interacting with proteins in the type I interferon pathway. The influence of NS1 on the cell's immune response denotes the potential of its inhibition, being a possible target of treatment against hRSV infection. Here, it was studied the interaction of hRSV NS1 with natural flavonoids chrysin, morin, kaempferol, and myricetin and their mono-acetylated chrysin and penta-acetylated morin derivatives using spectroscopic techniques and computational simulations. The fluorescence data indicate that the binding affinities are on the order of 105 M−1, which are directly related to the partition coefficient of each flavonoid with Pearson's correlation coefficients of 0.76–0.80. The thermodynamic analysis suggests that hydrophobic interactions play a key role in the formation of the NS1/flavonoid complexes, with positive values of enthalpy and entropy changes. The computational approach proposes that flavonoids bind in a region of NS1 formed between the C-terminal α3-helix and the protein core, important for its biological function, and corroborate with experimental data revealing that hydrophobic contacts are important for the binding. Therefore, the present study provides relevant molecular details for the development of a possible new strategy to fight infections caused by hRSV.
  • ItemArtigo
    Aqueous mixtures of cornstarch and Pluronic® F127 studied by experimental and computational techniques
    (2022-08-01) Baldassa, Monize A.B. [UNESP]; Dias, Raphael V.R. [UNESP]; Oliveira, Leandro C. [UNESP]; Feitosa, Eloi; Universidade Estadual Paulista (UNESP); Federal Technological University of Paraná (UTFPR)
    The possibility of interaction between cornstarch (CS) and amphiphilic molecules, such as the micelle-forming triblock copolymer Pluronic® F127 (F127), also known by Poloxamer 407, indicates that CS-F127 aqueous mixtures can regulate either the starch solubility or the copolymer micellization. Herein experimental and computational techniques were used to investigate CS-F127 aqueous mixtures aiming to highlight the role of these compounds on the molecular complexation. Dynamic light scattering results show that CS in water is highly polydisperse, while the F127 concentration and temperature influence the micellization process and the interaction with CS. Circular dichroism data of CS supernatants indicate the existence of small helical-like granules (Dh ≈ 800 nm) in the CS-F127 mixed aqueous solutions at 25 °C. UV–Vis spectrophotometry shows a small absorption band around 267 or 275 nm characteristic of micelles, granules, or molecular complexes, while FTIR and X-ray diffractometry indicate negligible structural changes. Lugol iodine tests at 25 °C show that both the precipitate and supernatant in the mixtures undergo some structural changes also indicating molecular complexation. Molecular dynamic simulations show the formation of stabilized inclusion complexes (V-amylose), where the propylene oxide segment of the copolymer inside the amylose helix and the ethylene oxide branches facing the aqueous media. These results together reveal weak CS-F127 interactions, evidencing a small solubility of CS both in the absence and presence of F127 as a solubilizing agent. Furthermore, moderate CS amounts do not change the F127 micelle structure.
  • ItemArtigo
    Riboflavin, a Potent Neuroprotective Vitamin: Focus on Flavivirus and Alphavirus Proteases
    (2022-07-01) Eberle, Raphael J.; Olivier, Danilo S.; Amaral, Marcos S.; Pacca, Carolina C.; Nogueira, Mauricio L.; Arni, Raghuvir K. [UNESP]; Willbold, Dieter; Coronado, Monika A.; Forschungszentrum Jülich GmbH; Universitätsstraße; Federal University of Tocantins; Federal University of Mato Grosso do Sul; FACERES Medical School; Faculdade de Medicina de São José do Rio Preto-FAMERP; University of Texas Medical Branch; Universidade Estadual Paulista (UNESP); Forchungszentrum Jülich
    Several neurotropic viruses are members of the flavivirus and alphavirus families. Infections caused by these viruses may cause long-term neurological sequelae in humans. The continuous emergence of infections caused by viruses around the world, such as the chikungunya virus (CHIKV) (Alphavirus genus), the zika virus (ZIKV) and the yellow fever virus (YFV) (both of the Flavivirus genus), warrants the development of new strategies to combat them. Our study demonstrates the inhibitory potential of the water-soluble vitamin riboflavin against NS2B/NS3pro of ZIKV and YFV and nsP2pro of CHIKV. Riboflavin presents a competitive inhibition mode with IC50 values in the medium µM range of 79.4 ± 5.0 µM for ZIKV NS2B/NS3pro and 45.7 ± 2.9 µM for YFV NS2B/NS3pro. Against CHIKV nsP2pro, the vitamin showed a very strong effect (93 ± 5.7 nM). The determined dissociation constants (KD) are significantly below the threshold value of 30 µM. The ligand binding increases the thermal stability between 4 °C and 8 °C. Unexpectedly, riboflavin showed inhibiting activity against another viral protein; the molecule was also able to inhibit the viral entry of CHIKV. Molecular dynamics simulations indicated great stability of riboflavin in the protease active site, which validates the repurposing of riboflavin as a promising molecule in drug development against the viruses presented here.