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Yellow fever virus (YFV) replication is highly dependent on host cell factors. YFV NS4B is reported to be involved in viral replication and immune evasion. Here interactions between NS4B and human proteins were determined using a GST pull-down assay and analyzed using 1-DE and LC-MS/MS. We present a total of 207 proteins confirmed using Scaffold 3 Software. Cyclophilin A (CypA), a protein that has been shown to be necessary for the positive regulation of flavivirus replication, was identified as a possible NS4B partner. 59 proteins were found to be significantly increased when compared with a negative control, and CypA exhibited the greatest difference, with a 22-fold change. Fisher's exact test was significant for 58 proteins, and the p value of CypA was the most significant (0.000000019). The Ingenuity Systems software identified 16 pathways, and this analysis indicated sirolimus, an mTOR pathway inhibitor, as a potential inhibitor of CypA. Immunofluorescence and viral plaque assays showed a significant reduction in YFV replication using sirolimus and cyclosporine A (CsA) as inhibitors. Furthermore, YFV replication was strongly inhibited in cells treated with both inhibitors using reporter BHK-21-rep-YFV17D-LucNeoIres cells. Taken together, these data suggest that CypA-NS4B interaction regulates YFV replication. Finally, we present the first evidence that YFV inhibition may depend on NS4B-CypA interaction.

Background: Chronic inflammation during morbid obesity significantly alters cutaneous tissue. Large weight loss achieved after bariatric surgery minimizes or halts damage caused by metabolic syndrome, but further deteriorates the clinical condition of skin. Postbariatric skin flaccidity produces major difficulties to plastic surgery. In this study, we analyzed differences in protein composition of the skin between patients with morbid obesity and those after large weight loss and established correlations between differentially expressed proteins and clinical characteristics of postbariatric skin tissue, to improve body contouring surgery techniques. Methods: Skin fragments were removed from the abdomen of 32 patients, who were allocated into 3 groups: morbidly obese, large weight loss without surgery, and postbariatric surgery. Samples were subjected to proteomic analysis, and the protein profiles of the groups were compared. Six differentially expressed proteins of clinical interest were validated by immunohistochemistry and statistical analysis. Results: Comparative analyses confirmed differences in protein profile of the skin between morbidly obese and large weight loss groups. A persistent increase in inflammatory markers such as haptoglobin was observed in all groups and decrease in the expression of collagen XIV, which regulates the physical properties of cutaneous tissue, was observed in the postbariatric group. Conclusions: High expression of haptoglobin associated with the decrease of Collagen XIV, vinculin, and periplakin in the groups after major weight losses, mainly postbariatric, confirm that the inflammatory lesion remains active in the skin and causes changes in its structural organization, with serious repercussions on its clinical characteristics and physical properties.

A abordagem proteômica tem permitido estudos em larga escala da expressão proteica em diferentes tecidos e fluidos corporais, em condições e/ou momentos distintos. O recente progresso de metodologias nessa área tem aberto novas oportunidades para obtenção de informações relevantes sobre processos normais e anormais que ocorrem no organismo humano. No presente artigo, é feita uma revisão das principais técnicas proteômicas e de suas aplicações no estudo de doenças humanas. Proteomic approach has allowed large-scale studies of protein expression in different tissues and body fluids in discrete conditions and/or time points. Recent advances of methodologies in this field have opened new opportunities to obtain relevant information on normal and abnormal processes occurring in the human body. In the current report, the main proteomics techniques and their application to human disease study are reviewed.

Several body fluids have been evaluated as new sources for cancer biomarker discovery. In this context, salivary and serum proteomics seem promising diagnostic and predictive tools for head and neck diseases. In the present study, we performed a proteomic analysis of saliva and serum from patients presenting head and neck squamous cell carcinoma (HNSCC) and compared the results before and after therapy. In saliva of cancer patients, we observed an altered protein profile, including over-expression of PLUNC and zinc-alpha-2-glycoprotein. Both proteins may contribute to control tumor growth and, therefore, represent targets for new analysis. We also detected serotransferrin and a modified transthyretin form with altered levels in serum from patients. Comparing preoperative and postreatment samples, the results showed that the protein profile after treatment reverted to a pattern closer to those observed for controls. These results add information on the role of secreted proteins in the cancer process and emphasize the potential of saliva and serum analysis for diagnosis and monitoring of HNSCC.

Submitted by Guilherme Lemeszenski (guilherme@nead.unesp.br) on 2013-08-22T18:40:35Z No. of bitstreams: 1 S1516-84842002000100008.pdf: 137534 bytes, checksum: 783607fb7d42658bb8c0d0806b89b579 (MD5) Made available in DSpace on 2013-08-22T18:40:35Z (GMT). No. of bitstreams: 1 S1516-84842002000100008.pdf: 137534 bytes, checksum: 783607fb7d42658bb8c0d0806b89b579 (MD5) Previous issue date: 2002-03-01 Made available in DSpace on 2013-09-30T18:42:13Z (GMT). No. of bitstreams: 2 S1516-84842002000100008.pdf: 137534 bytes, checksum: 783607fb7d42658bb8c0d0806b89b579 (MD5) S1516-84842002000100008.pdf.txt: 14137 bytes, checksum: e0f29560d29843bce428d31110c9f618 (MD5) Previous issue date: 2002-03-01 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T14:01:19Z No. of bitstreams: 2 S1516-84842002000100008.pdf: 137534 bytes, checksum: 783607fb7d42658bb8c0d0806b89b579 (MD5) S1516-84842002000100008.pdf.txt: 14137 bytes, checksum: e0f29560d29843bce428d31110c9f618 (MD5) Made available in DSpace on 2014-05-20T14:01:19Z (GMT). No. of bitstreams: 2 S1516-84842002000100008.pdf: 137534 bytes, checksum: 783607fb7d42658bb8c0d0806b89b579 (MD5) S1516-84842002000100008.pdf.txt: 14137 bytes, checksum: e0f29560d29843bce428d31110c9f618 (MD5) Previous issue date: 2002-03-01 Alterações genéticas em que a mutação de aminoácidos nas globinas afeta a estrutura da molécula tornando-a instável são classificadas como hemoglobinas instáveis. Devido à grande diversidade dos pontos de mutações por substituições e deleções de aminoácidos, as formas de instabilização se apresentam muito variadas. A hemoglobina Köln é a variante instável descrita com maior freqüência na literatura e a terceira descoberta no Brasil, as outras são Hb Niterói e Hb Hasharon. Anemia moderada, icterícia e presença de urina escura caracterizam as manifestações clínicas da Hb Köln. em programa de triagem neonatal identificamos uma criança com suspeita de heterozigose para hemoglobina Köln, confirmada por procedimentos eletroforéticos e HPLC. Avaliações por diferentes metodologias laboratoriais e estudo familiar auxiliam no diagnóstico precoce, possibilitando minimizar os sintomas decorrentes da hemoglobina anormal e a realização do aconselhamento genético e educacional destas alterações hereditárias. Hemoglobinopathies are a diverse group of inherited recessive disorders that include thalassemias and sickle cell disease. They were the first genetic diseases to be characterized at the molecular level and consequently have been used repeatedly as a prototype for the development of new techniques of mutation detection. A major group of the structural hemoglobins exhibit the property of instability in solution, resulting from their altered molecular structures. Mutation of amino acids in the globin affects the structure of the molecule turning it unstable and they are classified as unstable hemoglobins. Due to the great diversity of the mutation points, substitutions and deletions of amino acids, the unstable forms are very varied. Hemoglobin Köln is the unstable variant most described in literature and the third discovered in Brazil, the others are Hb Niterói and Hb Hasharon. Moderate anemia, jaundice and the presence of dark urine characterize Hb Köln s clinical manifestations. In a neonatal screening program we identified a child suspected of having hemoglobin Köln, which was confirmed by electrophoretic procedures and HPLC. Evaluations by different laboratory examinations and the family study aided in a precocious diagnosis, thereby facilitating to minimize the current symptoms of the abnormal hemoglobin, and thus the family received support with genetic and educational counseling of these hereditary alterations. UNESP IBILCE Departamento de Biologia FAMERP Departamento de Biologia Molecular UNESP IBILCE Departamento de Biologia

Made available in DSpace on 2013-08-28T14:07:40Z (GMT). No. of bitstreams: 1 WOS000278383600001.pdf: 960234 bytes, checksum: 9cd26a9750ec13e67eaf9019e1f48e7b (MD5) Made available in DSpace on 2013-09-30T18:12:07Z (GMT). No. of bitstreams: 2 WOS000278383600001.pdf: 960234 bytes, checksum: 9cd26a9750ec13e67eaf9019e1f48e7b (MD5) WOS000278383600001.pdf.txt: 69473 bytes, checksum: e718db9d53c8e8829f735f82524e401f (MD5) Previous issue date: 2010-05-04 Submitted by Vitor Silverio Rodrigues (vitorsrodrigues@reitoria.unesp.br) on 2014-05-20T14:00:48Z No. of bitstreams: 2 WOS000278383600001.pdf: 960234 bytes, checksum: 9cd26a9750ec13e67eaf9019e1f48e7b (MD5) WOS000278383600001.pdf.txt: 69473 bytes, checksum: e718db9d53c8e8829f735f82524e401f (MD5) Made available in DSpace on 2014-05-20T14:00:48Z (GMT). No. of bitstreams: 2 WOS000278383600001.pdf: 960234 bytes, checksum: 9cd26a9750ec13e67eaf9019e1f48e7b (MD5) WOS000278383600001.pdf.txt: 69473 bytes, checksum: e718db9d53c8e8829f735f82524e401f (MD5) Previous issue date: 2010-05-04 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Rede Proteoma do Estado de São Paulo Ludwig Institute for Cancer Research Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Background: The development and progression of cancer depend on its genetic characteristics as well as on the interactions with its microenvironment. Understanding these interactions may contribute to diagnostic and prognostic evaluations and to the development of new cancer therapies. Aiming to investigate potential mechanisms by which the tumor microenvironment might contribute to a cancer phenotype, we evaluated soluble paracrine factors produced by stromal and neoplastic cells which may influence proliferation and gene and protein expression.Methods: The study was carried out on the epithelial cancer cell line (Hep-2) and fibroblasts isolated from a primary oral cancer. We combined a conditioned-medium technique with subtraction hybridization approach, quantitative PCR and proteomics, in order to evaluate gene and protein expression influenced by soluble paracrine factors produced by stromal and neoplastic cells.Results: We observed that conditioned medium from fibroblast cultures (FCM) inhibited proliferation and induced apoptosis in Hep-2 cells. In neoplastic cells, 41 genes and 5 proteins exhibited changes in expression levels in response to FCM and, in fibroblasts, 17 genes and 2 proteins showed down-regulation in response to conditioned medium from Hep-2 cells (HCM). Nine genes were selected and the expression results of 6 down-regulated genes (ARID4A, CALR, GNB2L1, RNF10, SQSTM1, USP9X) were validated by real time PCR.Conclusions: A significant and common denominator in the results was the potential induction of signaling changes associated with immune or inflammatory response in the absence of a specific protein. Sch Med FAMERP, Dept Mol Biol, Sao Jose do Rio Preto, Brazil São Paulo State Univ UNESP, IBILCE, Dept Biol, Sao Jose do Rio Preto, Brazil Sch Med FAMERP, Dept Otorhinolaryngol & Head & Neck Surg, Sao Jose do Rio Preto, Brazil Univ São Paulo, Inst Biociencias, Dept Genet & Evolutionary Biol, São Paulo, Brazil Arnaldo Vieira de Carvalho Hosp, Dept Head & Neck Surg, São Paulo, Brazil Univ São Paulo, Sch Med, Dept Surg, Div Head & Neck Surg, BR-09500900 São Paulo, Brazil Heliopolis Hosp, Dept Head & Neck Surg, São Paulo, Brazil UNESP, Fac Engn Ilha Solteria, Dept Biol & Zootechny, Ilha Solteira, Brazil São Paulo State Univ UNESP, IBILCE, Dept Biol, Sao Jose do Rio Preto, Brazil UNESP, Fac Engn Ilha Solteria, Dept Biol & Zootechny, Ilha Solteira, Brazil FAPESP: 04/12054-9 FAPESP: 06/60162-0 FAPESP: 04/14846-0 FINEP: 01.07.0290.00

Made available in DSpace on 2022-04-28T20:43:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-07-01 Miliin, a new thiol-dependent serine protease purified from the latex of Euphorbia milii possesses a molecular weight of 79 kDa, an isoelectric point of 4.3 and is optimally active at 60°C in the pH range of and 7.5-11.0. Activity tests indicate that milliin is a thiol-dependent serine protease. © 2008 Bentham Science Publishers Ltd. Molecular Biophysics Graduate Program IBILCE-UNESP, Campinas, SP Center for Structural Molecular Biology Brazilian Synchrotron Light Laboratory, Campinas, SP Faculty of Pharmacy University of São Paulo, Ribeirão Preto, SP Department of Molecular Biology FAMERP, São José do Rio Preto, SP Department of Physics IBILCE-UNESP, São Paulo Department of Biophysics UNIFESP, São Paulo Department of Chemistry and Environmental Sciences IBILCE-UNESP, São José do Rio Preto SP DQCA IBILCE-UNESP, Rua Cristovão Colombo 2265, São José do Rio Preto SP, CEP 15054-000 Molecular Biophysics Graduate Program IBILCE-UNESP, Campinas, SP Department of Physics IBILCE-UNESP, São Paulo Department of Chemistry and Environmental Sciences IBILCE-UNESP, São José do Rio Preto SP DQCA IBILCE-UNESP, Rua Cristovão Colombo 2265, São José do Rio Preto SP, CEP 15054-000

ResumoA abordagem proteômica tem permitido estudos em larga escala da expressão proteica em diferentes tecidos e fluidos corporais, em condições e/ou momentos distintos. O recente progresso de metodologias nessa área tem aberto novas oportunidades para obtenção de informações relevantes sobre processos normais e anormais que ocorrem no organismo humano. No presente artigo, é feita uma revisão das principais técnicas proteômicas e de suas aplicações no estudo de doenças humanas.SummaryProteomic approach has allowed large-scale studies of protein expression in different tissues and body fluids in discrete conditions and/or time points. Recent advances of methodologies in this field have opened new opportunities to obtain relevant information on normal and abnormal processes occurring in the human body. In the current report, the main proteomics techniques and their application to human disease study are reviewed.

The eukaryotic translation initiation factor 3, subunit L (eIF3L) is one of the subunits of the eIF3 complex, an accessory protein of the Polymerase I enzyme and may have an important role in the Flavivirus replication by interaction with a viral non-structural 5 protein. Considering the importance of eIF3L in a diversity of cellular functions, we have produced the recombinant full-length eIF3L protein in Escherichia coli and performed spectroscopic and in silico analyses to gain insights into its hydrodynamic behavior and structure. Dynamic light scattering showed that eIF3L behaves as monomer when it is not interacting with other molecular partners. Circular dichroism experiments showed a typical spectrum of α-helical protein for eIF3L, which is supported by sequence-based predictions of secondary structure and the 3D in silico model. The molecular docking with the K subunit of the eIF3 complex revealed a strong interaction. It was also predicted several potential interaction sites in eIF3L, indicating that the protein is likely capable of interacting with other molecules as experimentally shown in other functional studies. Moreover, bioinformatics analyses showed approximately 8 putative phosphorylation sites and one possible N-glycosylation site, suggesting its regulation by post-translational modifications. The production of the eIF3L protein in E. coli and structural information gained in this study can be instrumental for target-based drug design and inhibitors against Flavivirus replication and to shed light on the molecular mechanisms involved in the eukaryotic translation initiation.