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91.
Takayuki K. Nemoto Yuko Ohara-Nemoto Gustavo Arruda Bezerra Yu Shimoyama Shigenobu Kimura 《The Journal of biological chemistry》2016,291(11):5913-5925
Exopeptidases, including dipeptidyl- and tripeptidylpeptidase, are crucial for the growth of Porphyromonas gingivalis, a periodontopathic asaccharolytic bacterium that incorporates amino acids mainly as di- and tripeptides. In this study, we identified a novel exopeptidase, designated acylpeptidyl oligopeptidase (AOP), composed of 759 amino acid residues with active Ser615 and encoded by PGN_1349 in P. gingivalis ATCC 33277. AOP is currently listed as an unassigned S9 family peptidase or prolyl oligopeptidase. Recombinant AOP did not hydrolyze a Pro-Xaa bond. In addition, although sequence similarities to human and archaea-type acylaminoacyl peptidase sequences were observed, its enzymatic properties were apparently distinct from those, because AOP scarcely released an N-acyl-amino acid as compared with di- and tripeptides, especially with N-terminal modification. The kcat/Km value against benzyloxycarbonyl-Val-Lys-Met-4-methycoumaryl-7-amide, the most potent substrate, was 123.3 ± 17.3 μm−1 s−1, optimal pH was 7–8.5, and the activity was decreased with increased NaCl concentrations. AOP existed predominantly in the periplasmic fraction as a monomer, whereas equilibrium between monomers and oligomers was observed with a recombinant molecule, suggesting a tendency of oligomerization mediated by the N-terminal region (Met16–Glu101). Three-dimensional modeling revealed the three domain structures (residues Met16–Ala126, which has no similar homologue with known structure; residues Leu127–Met495 (β-propeller domain); and residues Ala496–Phe736 (α/β-hydrolase domain)) and further indicated the hydrophobic S1 site of AOP in accord with its hydrophobic P1 preference. AOP orthologues are widely distributed in bacteria, archaea, and eukaryotes, suggesting its importance for processing of nutritional and/or bioactive oligopeptides. 相似文献
92.
Mauro Maciel de Arruda Fabiano Borges Figueiredo Andreza Pain Marcelino José Ronaldo Barbosa Guilherme Loureiro Werneck Elza Ferreira Noronha Gustavo Adolfo Sierra Romero 《Memórias do Instituto Oswaldo Cruz》2016,111(3):168-173
In Brazil, human and canine visceral leishmaniasis (CVL) caused byLeishmania
infantum has undergone urbanisation since 1980, constituting a public
health problem, and serological tests are tools of choice for identifying infected
dogs. Until recently, the Brazilian zoonoses control program recommended
enzyme-linked immunosorbent assays (ELISA) and indirect immunofluorescence assays
(IFA) as the screening and confirmatory methods, respectively, for the detection of
canine infection. The purpose of this study was to estimate the accuracy of ELISA and
IFA in parallel or serial combinations. The reference standard comprised the results
of direct visualisation of parasites in histological sections, immunohistochemical
test, or isolation of the parasite in culture. Samples from 98 cases and 1,327
noncases were included. Individually, both tests presented sensitivity of 91.8% and
90.8%, and specificity of 83.4 and 53.4%, for the ELISA and IFA, respectively. When
tests were used in parallel combination, sensitivity attained 99.2%, while
specificity dropped to 44.8%. When used in serial combination (ELISA followed by
IFA), decreased sensitivity (83.3%) and increased specificity (92.5%) were observed.
Serial testing approach improved specificity with moderate loss in sensitivity. This
strategy could partially fulfill the needs of public health and dog owners for a more
accurate diagnosis of CVL. 相似文献
93.
94.
Development of a scaffoldless three-dimensional engineered nerve using a nerve-fibroblast co-culture 总被引:1,自引:0,他引:1
Jennifer Baltich Leah Hatch-Vallier Aaron M. Adams Ellen M. Arruda Lisa M. Larkin 《In vitro cellular & developmental biology. Animal》2010,46(5):438-444
Nerve grafts are often required to replace tissue damaged by disease, surgery, or extensive trauma. Limitations such as graft
availability, donor site morbidity, and immune rejection have led investigators to develop strategies to engineer nerve tissue.
The goal of this study was to fabricate a scaffoldless three-dimensional (3D) nerve construct using a co-culture of fetal
nerve cells with a fibroblast monolayer and allow the co-culture to remodel into a 3D construct with an external fibroblast
layer and an internal core of interconnected neuronal cells. Primary fibroblasts were seeded on laminin-coated plates and
allowed to form a confluent monolayer. Neural cells isolated from E-15 spinal cords were seeded on top of the fibroblast monolayer
and allowed to form a networked monolayer across the monolayer of fibroblasts. Media shifts initiated contraction of the fibroblast
monolayer and a remodeling of the co-culture into a 3D construct held statically in place by the two constraint pins. Immunohistochemistry
using S100 (Schwann cell), β3-tubulin, DAPI, and collagen I indicated an inner core of nerve cells surrounded by an external
layer of fibroblasts. Conduction velocities of the 3D nerve and control (fibroblast-only) constructs were measured in vitro
and compared to in vivo measures of neonatal sciatic nerve. The conduction velocities of the nerve constructs were comparable
to 24-d-old neonatal nerve. The presence of Schwann cells and the ability to conduct neuronal signals in vitro suggest the
scaffoldless 3D nerve constructs will be a viable option for nerve repair. 相似文献
95.
Chiarella AP Arruda C Pina A Costa TA Ferreira RC Calich VL 《Microbes and infection / Institut Pasteur》2007,9(9):1078-1088
Protective immunity in paracoccidioidomycosis (PCM) is believed to be mediated by cellular immunity, but the role of T cell subsets has never been investigated. The aim of this study was to characterize the function of CD4+ and CD8+ T cells in the immunity developed by susceptible, intermediate and resistant mice after P. brasiliensis infection. In susceptible mice, depletion of CD4+ T cells did not alter disease severity and anergy of cellular immunity but diminished antibody production. Anti-CD8 treatment led to increased fungal loads, but restored DTH reactivity. In resistant mice, both CD4+ and CD8+ T cells control fungal burdens and cytokines although only the former regulate DTH reactions and antibody production. In the intermediate strain, deficiency of whole T and CD8+ T cells but not of CD4+ T or B cells led to increased mortality rates. Thus, in pulmonary PCM: (a) irrespective of the host susceptibility pattern, fungal loads are mainly controlled by CD8+ T cells, whereas antibody production and DTH reactions are regulated by CD4+ T cells; (c) CD4+ T cells play a protective role in the resistant and intermediate mouse strains, whereas in susceptible mice they are deleted or anergic; (d) genetic resistance to PCM is associated with concomitant CD4+ and CD8+ T cell immunity secreting type 1 and type 2 cytokines. 相似文献
96.
97.
Lysine degradation through the saccharopine pathway has been shown only in plants and animals. Here, we show that bacteria possess the genes encoding lysine-ketoglutarate reductase (LKR) and saccharopine dehydrogenase (SDH). In Silicibacter, the contiguous lkr and sdh genes are interspersed, in another frame, by a polypeptide of unknown function. The bacterial enzyme does not contain the 110-amino-acid interdomain (ID) that intersperses the LKR and SDH domains of the plant enzyme. The ID was found in Cyanobacteria interspersing polypeptides without similarities and activities of LKR and SDH. The LKR/SDH bifunctional polypeptide of animals and plants may have arisen from a α-proteobacterium with a configuration similar to that of Silicibacter, whereas the ID in the plant enzyme may have been inherited from Cyanobacteria. 相似文献
98.
Genetically modified sugarcane for bioenergy generation 总被引:2,自引:0,他引:2
Arruda P 《Current opinion in biotechnology》2012,23(3):315-322
Sugarcane breeding has significantly progressed over the past 30 years, but attempts to further increase crop yield have been limited due to the complexity of the sugarcane genome. An alternative to boost the crop yield is the introduction of genes encoding desirable traits in the elite sugarcane cultivars. Genetically modified sugarcane with increased yield and pest and disease resistance has already proven its value not only by the increased sugar content but also for the improvement of the crop performance. However, transgene stability is still a challenge since transgene silencing seems to occur in a large proportion of genetically modified sugarcane plants. In addition, regulatory issues associated with the crop propagation model will also be a challenge to the commercial approval of genetically modified sugarcane. 相似文献
99.
da Silva BR de Freitas VA Nascimento-Neto LG Carneiro VA Arruda FV de Aguiar AS Cavada BS Teixeira EH 《Peptides》2012,36(2):315-321
Antimicrobial peptides, molecules produced in many different organisms, have high biocidal activity against several microorganisms. However, several questions about these molecules remain unclear. Therefore, this report details a systematic survey of the literature on the use of antimicrobial peptides against oral pathogens and indicates which peptides and microorganisms are most extensively studied. Articles were located using the PubMed and Science Direct databases with the following inclusion criteria: publication date between 2002 and 2011; keywords "biofilm OR biological film OR biological layer OR bacterial growth" AND "peptide" AND "oral cavity OR mouth OR buccal mucosa OR oral mucosa OR mouth mucosa"; and abstract in English. A total of 73 articles were selected after refinement of the data. An increase in publications focusing on the use of antimicrobial peptides against oral microorganisms was observed. In addition, the peptides produced by cells of the oral mucosa (defensins, LL-37 and histatins) as well as Streptococcus mutans (among cariogenic bacteria) and Porphyromonas gingivalis (among periodontal bacteria) were the most studied subjects. It was concluded that the use of antimicrobial peptides as a tool for microbial control is of increasing importance, likely due to its widespread use, mechanism of action, and low rates of bacterial resistance. 相似文献
100.
da Silva Pinto L Gonçales RA Conceição FR Knabah PF Borsuk S Campos VF Arruda FV Leite FP 《Applied microbiology and biotechnology》2012,95(5):1235-1241
Bacillus sphaericus produces a two-chain binary toxin composed of BinA (42 kDa) and BinB (51 kDa), which are deposited as parasporal crystals during sporulation. The toxin is highly active against Culex larvae and Aedes and Anopheles mosquitoes, which are the principal vectors for the transmission of malaria, yellow fever, encephalitis, and dengue. The use of B. sphaericus and Bacillus thuringiensis in mosquito control programs is limited by their sedimentation in still water. In this study, the binA and binB genes were cloned and the recombinant BinAB protein was expressed in three strains of Escherichia coli. These recombinant strains were used in a toxicity assay against Culex quinquefasciatus larvae. The highest expression level was achieved when both proteins were expressed in a single operon construct. The BinAB protein expressed in the E. coli Arctic strain showed higher larvicidal activity than either of the recombinant proteins from the E. coli Ril or pLysS strains. Furthermore, it had the highest oviposition attraction (49.1%, P?0.05). These data suggest that biologically active recombinant BinA and BinB toxins might be useful in mosquito control programs, delivered by inactivated bacterial cells or in traps. 相似文献