cDNA cloning and 1.75 Å crystal structure determination of PPL2, an endochitinase and N-acetylglucosamine-binding hemagglutinin from Parkia platycephala seeds

Parkia platycephala lectin 2 was purified from Parkia platycephala (Leguminosae, Mimosoideae) seeds by affinity chromatography and RP-HPLC. Equilibrium sedimentation and MS showed that Parkia platycephala lectin 2 is a nonglycosylated monomeric protein of molecular mass 29 407 ± 15 Da, which contains six cysteine residues engaged in the formation of three intramolecular disulfide bonds. Parkia platycephala lectin 2 agglutinated rabbit erythrocytes, and this activity was specifically inhibited by N-acetylglucosamine. In addition, Parkia platycephala lectin 2 hydrolyzed β(1-4) glycosidic bonds linking 2-acetoamido-2-deoxy-β-d-glucopyranose units in chitin. The full-length amino acid sequence of Parkia platycephala lectin 2, determined by N-terminal sequencing and cDNA cloning, and its three-dimensional structure, established by X-ray crystallography at 1.75 Å resolution, showed that Parkia platycephala lectin 2 is homologous to endochitinases of the glycosyl hydrolase family 18, which share the (βα) 8 barrel topology harboring the catalytic residues Asp125, Glu127, and Tyr182. © 2006 The Authors.

Temporal evolution of sulfated zirconia sol-gel during dip coating: Analysis of mass drainage with time-varying refractive index

The formation of sulfated zirconia films from a sol-gel derived aqueous suspension is subjected to double-optical monitoring during batch dip coating. Interpretation of interferometric patterns, previously obscured by a variable refractive index, is now made possible by addition of its direct measurement by a polarimetric technique in real time. Significant sensitivity of the resulting physical thickness and refractive index curves (uncertainties of ±7 nm and ±0.005, respectively) to temporal film evolution is shown under different withdrawal speeds. As a first contribution to quantitative understanding of temporal film formation with varying nanostructure during dip coating, detailed analysis is directed to the stage of the process dominated by mass drainage, whose simple modeling with temporal t-1/2 dependence is verified experimentally. © 2006 Elsevier B.V. All rights reserved.

Detecção de Leptospira pomona em sêmen bovino por eletroforese capilar fluorescente

This study was performed in order to evaluate the detection limit of PCR with fluorescent capillary electrophoresis for Leptospira pomona diagnosis in bovine semen. Negative bovine semen samples were artificially contaminated with Leptospira pomona (10 0 to 10 7 bacteria/ ml) and DNA was extracted by phenol/chloroform protocol. DNA fragments visualization was done by three electrophoresis methods: under UV light in 2 % agarose gel, silver staining 8% polyacrylamide gel and fluorescent capillary electrophoresis. The detection limit of capillary electrophoresis for Leptospira pomona was 10 2bacteria/ml. Under UV light, in 2 % agarose gel, the detection limit was of 10 4 bacteria/ ml while for silver stained 8 % polyacrylamide gel it was 10 2 bacteria/ ml. PCR with fluorescent capillary electrophoresis is an efficient and rapid diagnostic test for DNA detection of Leptospira in bovine semen and this can be an important tool for herd and semen sanitary control in artificial insemination centers.

Effect of milk treatment on acidification, physicochemical characteristics, and probiotic cell counts in low fat yogurt

The effect of milk treatment (heat, high hydrostatic pressure - HHP, or combined heat and HHP) on acidification, physicochemical characteristics, and probiotic cell counts in low fat yogurt was studied. All samples were analyzed for fermentation time, pH, titratable acidity, total solids, water-holding capacity, syneresis, Hunter L*, a*, and b* values, Streptococcus thermophilus, Lactobacillus delbrueckii ssp bulgaricus, L. acidophilus, and Bifidobacterium longum. The application of HHP combined with thermal treatment resulted in yogurt gels with attractive physicochemical characteristics and high water-holding capacity. In addition to this, the milk treatment did not affect the probiotic bacteria growth. The balance of strains in the starter culture and level of inoculation influenced the yogurt fermentation and properties. The use of combined heat and HHP to treat milk before yogurt fermentation could be an alternative process for obtaining high quality, additive-free healthy products.

Synergy of DNA-bending nucleoid proteins and macromolecular crowding in condensing DNA

Many prokaryotic nucleoid proteins bend DNA and form extended helical protein-DNA fibers rather than condensed structures. On the other hand, it is known that such proteins (such as bacterial HU) strongly promote DNA condensation by macromolecular crowding. Using theoretical arguments, we show that this synergy is a simple consequence of the larger diameter and lower net charge density of the protein-DNA filaments as compared to naked DNA, and hence, should be quite general. To illustrate this generality, we use light-scattering to show that the 7kDa basic archaeal nucleoid protein Sso7d from Sulfolobus solfataricus (known to sharply bend DNA) likewise does not significantly condense DNA by itself. However, the resulting protein-DNA fibers are again highly susceptible to crowding-induced condensation. Clearly, if DNA-bending nucleoid proteins fail to condense DNA in dilute solution, this does not mean that they do not contribute to DNA condensation in the context of the crowded living cell. © 2007 World Scientific Publishing Company.

Waveguide features in self-patternable amine functionalized organic-inorganic hybrids

We report the use of organic-inorganic sol-gel derived poly(oxyehylene)/ siloxane hybrid doped with methacrylic acid modified zirconium (IV) n-propoxide for the fabrication of low cost waveguides trough direct UV laser writing. The organic-inorganic hybrids were processed as monoliths with size and shape control. The effective guiding region was identified and the number of modes was estimated via mode field analyses. A grating was successfully superimposed on the channel and the respective reflection spectrum was measured, enabling the determination of the guiding region dimension, the calculation of the effective refractive index of the guided mode. © 2007 IEEE.

Chalcopyrite and bornite differentially affect protein synthesis in planktonic cells of Acidithiobacillus ferrooxidans

Acidithiobacillus ferrooxidans is used in bioleaching industrial operations to recover metal ions from mineral sulfides. Chalcopyrite and bornite are copper sulfides that have the same elemental composition, but differ in their susceptibility to the bioleaching process. Our objective was to identify differentially expressed proteins in A. ferrooxidans LR cells exposed to chalcopyrite or bornite, as a sole energy source, for 24 hours. Compared to the control (without minerals), proteins were induced or repressed in planktonic cells after contact with chalcopyrite or bornite by 24 hours. These results demonstrated that the time of exposure to the copper minerals was enough to trigger distinct responses in the A. ferrooxidans metabolism. © 2007 Trans Tech Publications.

Polyethylene gel in the subcutaneous tissue of rats: Histopathologic and systemic evaluation

Purpose: To evaluate the histological and systemic response to subcutaneous injection of polyethylene gel in rats. Methods: Twenty-one white male rats were divided into 3 groups (G): G1 and G2 received subcutaneous polyethylene gel injection in the dorsal midline and were sacrificed at 30 and 60 postoperative days, respectively. G3 was not exposed to the polyethylene gel and was sacrificed after 60 days. Blood levels of lactate dehydrogenase (LDH), creatine kinase (CK), and alkaline phosphatase (ALP) were evaluated. The heart, kidney, liver, adrenal gland, injection site, and adjacent tissues were histologically examined. The results were submitted to statistical analysis. Results: There was no clinical evidence of extrusion, reduction of the injected volume, or abnormalities in the adjacent tissues. Blood levels of CK and LDH were normal and similar in all groups. ALP levels were significantly lower in G2 than in G1 and G3. The systemic organs were normal on histological examination in the 3 groups evaluated. Microscopically, the polyethylene gel was surrounded by a thin pseudocapsule formation and minimal inflammatory cell response, which decreased from G1 to G2. Conclusion: The subcutaneous injection of polyethylene gel in rats elicited minimal local inflammatory response and no systemic side effects. Copyright © 2008 Informa Healthcare USA, Inc.

Changes in amounts of total salivary gland proteins of Lutzomyia longipalpis (Diptera: Psychodidae) according to age and diet

Saliva plays important roles in facilitation of a bloodmeal, lubrication of mouthparts, and parasite transmission for some vector insects. Salivary composition changes during the lifetime of an insect, and differences in the salivary profile may influence its functions. In this report, the amount and profile of salivary gland protein of the American visceral leishmaniasis vector Lutzomyia longipalpis (Lutz & Neiva, 1912) were analyzed at different times of insect development and diet. Protein content from unfed female sand flies increased significantly with age, and a significant difference was observed in sugar-fed females during the first 10 d of adult life. Salivary protein content sharply decreased 1 d after blood feeding, with gradual increase in concentration the following days. SDS-polyacrylamide gel electrophoresis analysis revealed that most polypeptides present in the saliva of sugar-fed also were present in the saliva of blood-fed females. Understanding changes in sand fly's saliva contents at distinct days after emergence and the influence of a bloodmeal in this aspect may reveal the role played by saliva during leishmaniasis transmission. © 2008 Entomological Society of America.

Perfil eletroforético das proteínas de fase aguda em caprinos experimentalmente infectados com Trypanosoma evansi

Considered as one of the main agents of the tripanossomiases, Trypanosoma evansi causes a disease generically know as surra, with wide geographic occurence. This work has the aim to study the electrophoretic profile of the acute phase proteins of goats, experimentally infected with T. evansi. Ten crossbread female goats, around 4 months of age, clinically healthy and serum negative for the presence of antibodies anti-T. evansi (IFAT) were used. The animals were divided in two groups: six were inoculated (G1) intravenously with 2,38 × 10 6 tripomastigotes of T. evansi and four were kept as noninfected controls. The blood for serum was collected daily until the 14 days after inoculation (DAI), weekly up to the 98 th DAI and every two weeks up to the 364 th DAI. The serum proteins were separacted by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDSPAGE). Twenty-one proteins were found in the serum of the goats, eight were nominally identified; phosphorylase, transferrin, albumin, antitrypsin, acid glicoprotein, haptoglobin, hemoglobin, and light chain immunoglobulin.

A preliminary differential proteomic analysis of the periplasmic fraction in Acidithiobacillus ferrooxidans planktonic and attached cells exposed to chalcopyrite

The Acidithiobacillus ferrooxidans periplasmic space is known to have proteins involved in the respiratory chains. There are no reports about the expression of the periplasmic proteins in A. ferrooxidans cells attached to chalcopyrite. In this preliminary work, it was compared the periplasmic protein profiles of A. ferrooxidans planktonic and attached cells after exposure to chalcopyrite for 2 hours. The bacterial response to chalcopyrite was investigated by a proteomic approach (two- dimensional gel electrophoresis and mass spectrometry). Four proteins differentially expressed between planktonic and attached cells after exposure to chalcopyrite were identified. Two of these proteins, repressed in chalcopyrite- attached cells, were both identified as superoxide dismutase, whereas the single strand binding protein (SSB) and the PspA/IM30 protein were induced. These results showed that A. ferrooxidans chalcopyrite- attached and planktonic cells show differential expression of the periplasmic proteins and that a proteomic approach can provide a valuable tool to detect proteins related to the A. ferrooxidans response to attachment to the mineral substrates. © (2009) Trans Tech Publications.

Experimental infection of commercial layers using a Salmonella enterica serovar Gallinarum strain: Leukogram and serum acute-phase protein concentrations

The aim of the present study was to evaluate white blood cell counts and serum protein profiles of commercial layers experimentally infected with Salmonella Gallinarum (SG) in order to better understand the pathophysiology of the disease caused by this bacterium. 180 five-day-old commercial layers were divided into 3 groups (G); G1 and G2 received 0.2 mL of inoculate containing 3.3x10 8 CFU or 3.3×10 5 CFU SG resistant to nalidix acid (Nal r)/mL, respectively, directly into their crops. G3 group did not receive the inoculum. Birds were sacrificed 24 hours before (T1) and 24 hours after the infection (T2), and three (T3), five (T4), seven (T5), and ten (T6) days after the administration of the inoculum. White blood cell counts were carried out in a Neubauer hemocytometer and in blood smears. Serum protein concentrations, including acute-phase proteins, were determined using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Data were submitted to analysis of variance, and means were compared by Tukey's test (P <0.05). G1 and G2 groups presented higher leukocyte counts on T4 and T5, respectively, due to the increase of circulating lymphocytes and heterophils, with a significant difference relative to G3. In electrophoresis, an increase in the serum levels of ceruloplasmin, haptoglobin, and hemopexin and a decrease in transferrin, which are acute-phase proteins, was verified. IgA serum levels did not change; however, IgG concentration increased during the infection. In conclusion, the results provide information for the better understanding of the pathophysiology of fowl typhoid.

Influence of the coloring agent concentration on bleaching gel and pulp chamber temperatures during dental bleaching

This study evaluated the Influence of the coloring agent concentration on the temperature of the gel layer and pulp chamber during dental bleaching with an LED/laser light source. Ten human incisors and a digital thermometer with K-type thermocouples were used. Using a high-speed spherical diamond bur, endodontic access was gained through openings on the lingual faces until pulp chamber was exposed. One end of the thermocouple was placed on the labial surface (immersed in bleaching gel) and the other end in the pulp chamber. The same 10 specimens were used in the 12 groups, according to the type and concentration of bleaching gel. Each bleaching gel was used in four different concentrations: manipulated without coloring, with normal quantity recommended by the manufacturer, with double the recommended amount of coloring, and with triple the recommended amount of coloring. The temperature rise was measured every 30 seconds for three minutes with a K-type thermocouple. The data were analyzed by ANOVA to examine the concentration and type of bleaching gel. This test was followed by Tukey's test, which was performed Independently for the gel at the labial surface and the pulp chamber (a = 5%). For both surfaces, values of p = 0.00 were obtained for all factors and for the Interaction between them. The varying concentrations of coloring agent produced statistically significant differences in terms of temperature increase for both the gel layer and the pulp chamber during activation.

Antimicrobial analysis of chlorhexidine gel and intracanal medicaments against microorganisms inoculated in root canals

The aim of this paper was to evaluate the antimicrobial activity of 2% chlorhexidine gel (CLX) associated with various intracanal medicaments against Candida albicans and Enterococcus faecalis inoculated in root canals. Thirty six human single-rooted teeth were contaminated with C.albicans and E.faecalis. The canals were instrumented using 2% CLX gel and were divided into three groups according to the intracanal medicaments (ICM) used. Group 1: calcium hydroxide paste [Ca(OH)], Group 2: 2% chlorhexidine gel (CLX) and Group 3: 2% CLX gel + Ca(OH). The root canal collections were performed after 21 days of contamination (control collection), after instrumentation (1st collection), after 14 days of intracanal medicament (2nd collection) and 7 days after medicament removal (3rd collection). The microbiological samples were plated in culture media and incubated for 48 hours. The results were submitted to Kruskal-Wallis test (P ≤ 0.05). It was verified that the instrumentation with CLX reduced the number of CFU/ml significantly when compared with the confirmation collection (control). However, the use of the ICM was only capable to eliminate completely the microorganisms in the root canals without difference statistics between them. Although the use of 2% chlorherixidine gel reduces the number of microorganisms significantly, only the ICM calcium hydroxide and calcium hydroxide associated with chlorhexidine are able to eliminate these microorganisms completely.