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Trametes villosa Lignin Peroxidase (TvLiP): Genetic and Molecular Characterization
( Rita Terezinha De Oliveira Carneiro ),( Maiza Alves Lopes ),( Marilia Lordelo Cardoso Silva ),( Veronica Da Silva Santos ),( Volnei Brito De Souza ),( Aurizangela Oliveira De Sousa ),( Carlos Primin 한국미생물 · 생명공학회 2017 Journal of microbiology and biotechnology Vol.27 No.1
White-rot basidiomycetes are the organisms that decompose lignin most efficiently, and Trametes villosa is a promising species for ligninolytic enzyme production. There are several publications on T. villosa applications for lignin degradation regarding the expression and secretion of laccase and manganese peroxidase (MnP) but no reports on the identification and characterization of lignin peroxidase (LiP), a relevant enzyme for the efficient breakdown of lignin. The object of this study was to identify and partially characterize, for the first time, gDNA, mRNA, and the corresponding lignin peroxidase (TvLiP) protein from T. villosa strain CCMB561 from the Brazilian semiarid region. The presence of ligninolytic enzymes produced by this strain grown in inducer media was qualitatively and quantitatively analyzed by spectrophotometry, qPCR, and dye fading using Remazol Brilliant Blue R. The spectrophotometric analysis showed that LiP activity was higher than that of MnP. The greatest LiP expression as measured by qPCR occurred on the 7<sup>th</sup> day, and the ABSA medium (agar, sugarcane bagasse, and ammonium sulfate) was the best that favored LiP expression. The amplification of the TvLiP gene median region covering approximately 50% of the T. versicolor LPGIV gene (87% identity); the presence of Trp199, Leu115, Asp193, Trp199, and Ala203 in the translated amplicon of the T. villosa mRNA; and the close phylogenetic relationship between TvLiP and T. versicolor LiP all indicate that the target enzyme is a lignin peroxidase. Therefore, T. villosa CCMB561 has great potential for use as a LiP, MnP, and Lac producer for industrial applications.
Catalytic Properties of Lipases Immobilized onto Ultrasound-treated Chitosan Supports
Matheus Dorneles de Mello,Deborah Cordeiro,Lilian Terezinha Costa,Cristian Follmer 한국생물공학회 2013 Biotechnology and Bioprocess Engineering Vol.18 No.6
Ultrasound sonication has been utilized toproduce fragmentation of chitosan polymer and hence increasethe chitosan surface area, making it more accessible tointeractions with proteins. In this context, we haveinvestigated the catalytic properties of lipases from differentsources immobilized onto ultrasound-treated chitosan (ChiS)pre-activated with glutaraldehyde (ChiS-G). Atomic forcemicroscopy indicated that ChiS-G displays a more cohesiveframe without the presence of sheared/fragmented structureswhen compared with ChiS, which might be attributed tothe cross-linking of the polysaccharide chains. Theimmobilization efficiency onto ChiS-G and ChiS wereremarkably higher than using conventional beads. Incomparison with the free enzymes, lipases immobilizedonto ChiS show a slight increase of apparent Km anddecrease of apparent Vmax. On the other hand, immobilizationonto ChiS-G resulted in an increase of Vmax, even though aslight increase of Km was also observed. These datasuggest that the activation of chitosan with glutaraldehydehas beneficial effects on the activity of the immobilizedlipases. In addition, the immobilization of the lipases ontoChiS-G displayed the best reusability results: enzymesretained more than 50% of its initial activity after fourreuses, which might be attributed to the covalent attachmentof enzyme to activated chitosan. Overall, our findingsdemonstrate that the immobilization of lipases ontoultrasound-treated chitosan supports is an effective andlow-cost procedure for the generation of active immobilizedlipase systems, being an interesting alternative to conventionalchitosan beads.
Expression of Sara2 Human Gene in Erythroid Progenitors
Jardim, Denis Leonardo Fontes,Cunha, Anderson Ferreira Da,Duarte, Adriana Da Silva Santos,Santos, Camila Oresco Dos,Saad, Sara Terezinha Olalla,Costa, Fernando Ferreira Korean Society for Biochemistry and Molecular Biol 2005 Journal of biochemistry and molecular biology Vol.38 No.3
A human homologue of Sar1, named Sara2, was shown to be preferentially expressed during erythropoiesis in a culture stimulated by EPO. Previous studies, in yeast, have shown that secretion-associated and Ras-related protein (Sar1p) plays an essential role in protein transport from the endoplasmic reticulum to the Golgi apparatus. Here, we report the molecular analysis of Sara2 in erythroid cell culture. A 1250 bp long cDNA, encoding a 198 amino-acid protein very similar to Sar1 proteins from other organisms, was obtained. Furthermore, we also report a functional study of Sara2 with Real-time quantitative PCR analysis, demonstrating that expression of Sara2 mRNA increases during the initial stages of erythroid differentiation with EPO and that a two-fold increase in expression occurs following the addition of hydroxyurea (HU). In K562 cells, Sara2 mRNA was observed to have a constant expression and the addition of HU also up-regulated the expression in these cells. Our results suggest that Sara2 is an important gene in processes involving proliferation and differentiation and could be valuable for understanding the vesicular transport system during erythropoiesis.
Letícia Z. Oliveira,Vera F. M. Hossepian de Lima,Marcelo A. Levenhagen,Ricarda M. dos Santos,Terezinha I. Assumpção,José O. Jacomini,André F. C. de Andrade,Rubens P. de Arruda,Marcelo E. Beletti 대한수의학회 2011 Journal of Veterinary Science Vol.12 No.3
The objective of this study was to characterize acrosomal ultrastructure following discontinuous Percoll gradient centrifugation of cryopreserved bovine sperm. Semen was collected from six bulls of different breeds and three ejaculates per bull were evaluated. Frozen semen samples were thawed and the acrosomal region of sperm cells was evaluated by transmission electron microscopy (TEM) before (n = 18) and after (n = 18) Percoll centrifugation. The evaluation of 20 sperm heads from each of the 36 samples analyzed ensured that a large number of cells were investigated. The data were subjected to analysis of variance at a level of significance of 5%. Percoll centrifugation reduced the percentage of sperm exhibiting normal acrosomes (from 61.77 to 30.24%), reduced the percentage of sperm presenting atypical acrosome reactions (from 28.38 to 4.84%) and increased the percentage of sperm exhibiting damage in the acrosome (from 6.14 to 64.26%). The percentage of sperm with typical acrosome reactions was not significantly different before (3.70%) and after (0.67%) centrifugation. TEM distinguished four different types of acrosomal status and enabled ultrastructural characterization of acrosomal injuries. The percentage of sperm exhibiting normal acrosomes decreased and damage in the acrosome was the most frequent acrosomal injury with the Percoll gradient centrifugation protocol utilized.
CBCT-based assessment of root canal treatment using micro-CT reference images
Lamira, Alessando,Mazzi-Chaves, Jardel Francisco,Nicolielo, Laura Ferreira Pinheiro,Leoni, Graziela Bianchi,Silva-Sousa, Alice Correa,Silva-Sousa, Yara Terezinha Correa,Pauwels, Ruben,Buls, Nico,Jacob Korean Academy of Oral and Maxillofacial Radiology 2022 Imaging Science in Dentistry Vol.52 No.-
Purpose: This study compared the root canal anatomy between cone-beam computed tomography (CBCT) and micro-computed tomography (micro-CT) images before and after biomechanical preparation and root canal filling. Materials and Methods: Isthmus-containing mesial roots of mandibular molars(n=14) were scanned by micro-CT and 3 CBCT devices: 3D Accuitomo 170 (ACC), NewTom 5G (N5G) and NewTom VGi evo (NEVO). Two calibrated observers evaluated the images for 2-dimensional quantitative parameters, the presence of debris or root perforation, and filling quality in the root canal and isthmus. The kappa coefficient, analysis of variance, and the Tukey test were used for statistical analyses(α=5%). Results: Substantial intra-observer agreement (κ=0.63) was found between micro-CT and ACC, N5G, and NEVO. Debris detection was difficult using ACC (42.9%), N5G (40.0%), and NEVO (40%), with no agreement between micro-CT and ACC, N5G, and NEVO (0.05<κ<0.12). After biomechanical preparation, 2.4%-4.8% of CBCT images showed root perforation that was absent on micro-CT. The 2D parameters showed satisfactory reproducibility between micro-CT and ACC, N5G, and NEVO (intraclass correlation coefficient: 0.60-0.73). Partially filled isthmuses were observed in 2.9% of the ACC images, 8.8% of the N5G and NEVO images, and 26.5% of the micro-CT images, with no agreement between micro-CT and ACC, and poor agreement between micro-CT and N5G and NEVO. Excellent agreement was found for area, perimeter, and the major and minor diameters, while the roundness measures were satisfactory. Conclusion: CBCT images aided in isthmus detection and classification, but did not allow their classification after biomechanical preparation and root canal filling.
Tuane Krupek,Bruna Juliana Wanczinski Ferrari,Maria Angelica Raffaini Covas Pereira da Silva,Christiano Rodrigues Schamber,Dennis Armando Bertolini,Marcos Luciano Bruschi,Isolde Terezinha Santos Previ 한국식품영양과학회 2020 Journal of medicinal food Vol.23 No.5
The impact of oral supplementation with an effervescent glutamine formulation on the beneficial effects of antiretroviral therapies was evaluated in people living with HIV/AIDS. For this purpose, 12 HIV/AIDS carrier patients with CD4+ T cell counts <500, and who had received the same antiretroviral therapy for at least 1 year before starting this investigation were selected. The patients were required to dissolve the effervescent glutamine formulation (supplied in sachets) in water immediately before oral ingestion (12.4 g), once a day, after lunch or after dinner during 30 days. CD4+ T cell counts, complete blood cell counts, serum cytokines, and amino acids levels were quantified; biochemical and toxicological measurements were performed. The numbers of CD4+ T cells were increased (P < .05), and the serum C-reactive protein levels decreased (P < .01) after the administration of effervescent glutamine formulation. Serum levels of interferon-gamma inducible protein-10, RANTES, and macrophage inflammatory protein-1β were decreased after the treatment with effervescent glutamine formulation. No changes were observed in the serum levels of amino acids, hematological, toxicological, and biochemical parameters. In conclusion, the treatment during 30 days with effervescent glutamine formulation was well tolerated, promoted reduction of inflammation, and improved the beneficial effects of antiretroviral therapies in HIV/AIDS carrier patients.
CBCT-based assessment of root canal treatment using micro-CT reference images
Lamira Alessando,Mazzi-Chaves Jardel Francisco,Nicolielo Laura Ferreira Pinheiro,Leoni Graziela Bianchi,Silva-Sousa Alice Corrêa,Silva-Sousa Yara Terezinha Corrêa,Pauwels Ruben,Buls Nico,Jacobs Reinhi 대한영상치의학회 2022 Imaging Science in Dentistry Vol.52 No.3
Purpose: This study compared the root canal anatomy between cone-beam computed tomography (CBCT) and micro-computed tomography (micro-CT) images before and after biomechanical preparation and root canal filling. Materials and Methods: Isthmus-containing mesial roots of mandibular molars(n=14) were scanned by micro-CT and 3 CBCT devices: 3D Accuitomo 170 (ACC), NewTom 5G (N5G) and NewTom VGi evo (NEVO). Two calibrated observers evaluated the images for 2-dimensional quantitative parameters, the presence of debris or root perforation, and filling quality in the root canal and isthmus. The kappa coefficient, analysis of variance, and the Tukey test were used for statistical analyses(α=5%). Results: Substantial intra-observer agreement (κ=0.63) was found between micro-CT and ACC, N5G, and NEVO. Debris detection was difficult using ACC (42.9%), N5G (40.0%), and NEVO (40%), with no agreement between micro-CT and ACC, N5G, and NEVO (0.05<κ<0.12). After biomechanical preparation, 2.4%-4.8% of CBCT images showed root perforation that was absent on micro-CT. The 2D parameters showed satisfactory reproducibility between micro-CT and ACC, N5G, and NEVO (intraclass correlation coefficient: 0.60-0.73). Partially filled isthmuses were observed in 2.9% of the ACC images, 8.8% of the N5G and NEVO images, and 26.5% of the micro-CT images, with no agreement between micro-CT and ACC, and poor agreement between micro-CT and N5G and NEVO. Excellent agreement was found for area, perimeter, and the major and minor diameters, while the roundness measures were satisfactory. Conclusion: CBCT images aided in isthmus detection and classification, but did not allow their classification after biomechanical preparation and root canal filling.(