Molecular cloning of a cDNA encoding a pollen extracellular protein as a potential source of a pollen allergen in Brassica rapa

Gastritis caused by infection with Helicobacter pylori is one of the most common infectious diseases worldwide. There are data on the epidemiology, pathophysiology and histology of this disease that show that Helicobacter pylori gastritis plays an important role in gastric carcinogenesis. However, we must remember that only a very few among those infected with Helicobacter pylori will develop gastric cancer. Hence, one of the main targets of future research will be to identify individuals who carry a greater risk for developing gastric cancer and may therefore benefit from eradication of Helicobacter pylori in terms of gastric cancer prevention.     

A cellulase complex in culture filtrates of Penicillium citrinum

During growth in a liquid medium that contained a single soluble or an insoluble cellulosic carbon source Penicillium citrinum released a complex of cellulase enzymes into the medium. A temperature of 30 degrees C was best for cellulase production. Presence of carbon-containing compounds, particularly glucose, inhibited cellulase activity. The enzyme complex was separated by gel filtration followed by ion-exchange chromatography into 11 components, 4 of high molecular weight and 7 of low molecular weight. One of the components (Bb) had the character of C1 cellulase enzyme. When the components were combined they released more reducing sugars from cullulosic substrates than when they were used singly.     

Transport of a novel complex in the cytoplasmic matrix of Chlamydomonas flagella

Proteins necessary for maintenance and function of eukaryotic flagella are synthesized in the cell body. Transport of the inner dynein arm subunit p28(IDA4) in Chlamydomonas flagella requires the activity of the kinesin KHP1(FLA10), a protein inactive at restrictive temperature in fla10, a temperature-dependent mutant of flagellar assembly. To identify other molecules involved in active transport of inner dynein arms within flagella we searched for polypeptides of the cytoplasmic matrix of flagella that fulfill two conditions: they bind to p28 and require the activity of KHP1 to be present in flagella. We found that the cytoplasmic matrix of flagella contains a previously unidentified "17S" complex of at least 13 polypeptides that in part is associated with p28. The 17S complex is present at permissive but not at restrictive temperature in fla10 flagella. It also turns over in the cytoplasmic matrix more frequently than dynein arms within the axoneme. This evidence suggests that the 17S complex transports precursors of inner dynein arms within flagella.     

Solution conformation and dynamics of an extracellular polysaccharide isolated from Bradyrhyzobium as deduced from 1H-NMR off resonance ROESY and 13C-NMR relaxation measurements

The conformational and dynamical features of an extracellular branched deacetylated polysaccharide isolated from Bradyrhizobium (Chamaecytisus proliferus) have been investigated by homo and heteronuclear NMR methods. 1H-NMR cross relaxation rates have been obtained for this polysaccharide through regular NOESY and ROESY spectra as well as by modern off resonance ROESY techniques. Local proton-proton correlation times as well as interproton distances have been obtained. 13C-NMR relaxation parameters (T1, T2, NOE) have also been measured at two different magnetic fields and interpreted using different approximations based on the Lipari and Szabo model free approach. The analysis of the data indicates the existence of important flexibility for the different linkages of the polysaccharide. Motions in the range of several ns contribute to the relaxation of the macromolecule, although faster internal motions in the 600-800 ps time scales are also present. These time scales indicate that segmental motions as well as internal motions around the glycosidic linkages are the major sources of relaxation for this molecule at 299 K.     

Lipase from a Brazilian strain of Penicillium citrinum

A single intratracheal instillation of porcine pancreatic elastase (PPE, 100 U/Kg) induces in rabbits bronchial secretory cell metaplasia as well as emphysematous changes. The mucus hypersecretion and the marked reduction of ciliated cells matched by a high percentage of atypical cilia are responsible for the delayed mucociliary clearance in this model. S-Carboxymethylcysteine lysine salt (SCMC-LYS, 0.35 g/Kg b.w.), given per os daily for 10 days starting 2 days before elastase administration, significantly ameliorated the mucociliary clearance. The pharmacological treatment did not modify the degree of secretory cell metaplasia and the percentage of atypical cilia, or prevent the alveolar wall destruction. At TEM examination, the morphological aspects of secretion occurring in bronchial tree of PPE-treated animals were rarely visible in the PPE + SCMC-LYS treated group. The beneficial effect of SCMC-LYS on mucociliary clearance may be ascribed to an antisecretagogue effect of this drug through elastase inhibition and to a reduction of mucus viscosity.     

Identification of a putative membrane-interacting domain of CTP:phosphocholine cytidylyltransferase from rat liver

A putative membrane-interacting domain of CTP:phosphocholine cytidylyltransferase (CT) was identified using two peptide-specific antibodies. One antibody (SA2) was raised against the N-terminus of CT (amino acid residues 1-17) and the other antibody (SA209) against an alpha-helical domain of the enzyme (amino acid residues 247-257). Both antibodies quantitatively immunoprecipitated CT from rat liver cytosol and showed specificity towards CT when octylglucoside extracts of rat liver cytosol were assessed by Western blot analysis. However, further experiments revealed that the antibodies had different characteristics. Whereas the antibody directed against the N-terminus of CT (SA2) did not influence CT/membrane interaction, the new antibody (SA209) against the alpha-helical domain of the enzyme interfered with this interaction. Our results provide experimental evidence that the alpha-helical domain (amino acid residues 228-287) of CT may serve as a membrane-interacting domain.     

Heat shock induction by a misassembled cytoplasmic membrane protein complex in Escherichia coli

We analysed the effects of the overproduction of parts or all of a multisubunit ATP-binding cassette (ABC) transporter, the MalFGK2 complex, involved in the uptake of maltose and maltodextrins in Escherichia coli. We found that production of the MalF protein alone was inducing the phtrA promoter, which is under the control of a recently discovered sigma factor, sigma24, involved in the response to extracytoplasmic stresses. The production level, stability and localization of MalF were not altered when produced without its partners, suggesting that the protein was correctly inserted in the membrane. Our results indicate that a large periplasmic loop located between the third and fourth transmembrane segment of MalF, the L3 loop, is responsible for phtrA induction: (i) deleted MalF proteins with no L3 loop or with a L3 loop lacking 120 amino acids do not induce the phtrA promoter; (ii) the export to the periplasm of the L3 loop alone or fused to MalE induces the phtrA promoter. Moreover, the proteolytic sensitivity of MalF is different when it is produced alone and when MalF and MalG are produced together, suggesting a change in the conformation and/or accessibility of MalF. These results suggest that some inner membrane proteins can be sensed outside the cytoplasm by a quality control apparatus or by the export machinery. Moreover, the observation of the phtrA induction by MalF could be a useful new tool for studying the insertion and assembly of the MalFGK2 complex.     

Cholinergic modulation of extracellular ATP-induced cytoplasmic calcium concentrations in cochlear outer hair cells

Outer hair cells (OHC) of the mammalian cochlea modulate the inner hair cell (IHC) mechanoelectrical transduction of sound. They are contacted by synapsing efferent neurons from the CNS, their main efferent neurotransmitter being acetylcholine (ACh). OHC function and in particular their control of [Ca2+]i is highly important and is modulated by ACh and also by other substances including extracellular (EC) ATP. OHC carry at their efferent synapse a not yet completely identified neuronal type of ionotropic ACh receptor (AChR), with an unusual pharmacology, which is, in vivo and in vitro, reversibly blocked by alpha-bungarotoxin (alpha-bgtx). The AChR mediates a fast influx of Ca2+ into OHC which, in turn, activates a closeby located outwardly-directed Ca(2+)-dependent K(+)-channel, thus shortly hyperpolarizing the cell. A cloned homomeric alpha 9 nAChR mimicks the function and pharmacology of this receptor. We here report results from a study designed to observe only slower effects triggered by EC ATP and the ACh-AChR system. EC presence of ATP at OHC increases [Ca2+]i by activating both P2x and P2y purinoceptors and also by indirect activation of OHC L-type Ca(2+)-channels. The L-type channel activation is responsible for a large part of the [Ca2+]i increase. Simultaneous EC presence of ACh and ATP at OHC was found to depress ATP-induced effects on OHC [Ca2+]i, an effect that is completely blocked in the presence of alpha-bgtx. Our observations suggest that the ACh-AChR system is involved in the modulation of the observed EC ATP-triggered events; possibly the OHC AChR is able to act both in its well known rapid ionotropic way, but also, perhaps after modification in a slower, metabotropic way interfering with the EC ATP-induced [Ca2+]i increase.     

Immunomodulatory effects of a polysaccharide from Tamarindus indica

Using a streptavidin/biotin labeling technique, we unintentionally cloned a gene encoding a biotin carboxyl carrier protein, a subunit of biotin-dependent enzymes, from a genomic library of Streptococcus mutans strain UT-041. In colony lifts, the clone reacted positively to the streptavidin-containing detection system but could not be detected in Southern blot analysis. The amino acid sequence of the gene product, deduced from its nucleotide sequence, demonstrated all the features common to biotin carboxyl carrier proteins from other bacteria, indicating that the biotin carboxyl carrier protein in the clone had produced a "false-positive" (DNA probe-independent) reaction by binding to the streptovidin. To circumvent this problem with the detection system in gene probing in the future, we recommend that all positive clones be screened by direct incubation with streptavidin-alkaline phosphatase (SA-AP) in the absence of biotin-labeled probe DNA. Clones binding to SA-AP would be considered false positives.     

The liver as a source of somatomedin

Somatomedin (Sm) activity (measured by [35S] utake in chick embryo cartilage) was determined in serum samples simultaneously drawn from the hepatic vein, portal vein, femoral artery and demoral vein of seventeen anaesthezied normal adult dogs). A pool of human serum was taken as reference (Sm = 1 U/ml). Sm levels in the peripheral vein of dogs were 0.38 +/- 0.03 U/ml). Mean +/- SEM). Sm activity was greater in the hepatic vein (0.48 U/ml) than in the other vessels (0.36, 0.39, 0.38 U/ml), and the paired differences were significant (P less than 0.002 to P less than 0.05). In three dogs which received b-GH (20 IU/day), the Sm levels were significantly increased after nine days in the femoural vein (P less than 0.05) and in the hepatic vein (P less than 0.05). The validity of the assay is discussed; a possible interference of NEFA in the assay is eliminated. The difference of Sm levels between hepatic and portal veins, related to hepatic flow measured in seven of these dogs, indicate an important production of Sm by the liver.     

Protoplasts of Cosmarium as a potential protein source

Cosmarium turpinii, a fast-growing desmid alga, transforms into protoplasts in 4 h when incubated in a mineral medium + 0.4 M mannitol + 0.5% Cellulysin.     

Behavioral anchors as a source of bias in rating.

Behavioral anchors may affect the way that raters process information about ratees, and may in some cases be a source of bias in rating. This study tested the hypothesis that the presence of behavioral anchors that closely matched behaviors actually observed by raters would bias performance ratings. Subjects (N?=?180) viewed videotaped lectures and rated them, using scales that contained examples of either good or bad performance that had actually occurred on the tapes, but that were not representative of the ratee's overall performance. One half of the subjects read the scales before viewing the lectures; the remaining subjects read the scales only after viewing the lectures. There was a significant scale effect, but no Scale?×?Order interaction; ratings were biased in the direction of unrepresentative anchors. These results suggest that behavioral anchors can be a source of bias in ratings and they may lead to biased recall, but they probably do not bias the observation and encoding of ratee behavior. Our results suggest that behaviorally anchored scales are not necessarily more objective or less prone to bias than are scales without behavioral anchors. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Preferential release of ATP and its extracellular catabolism as a source of adenosine upon high- but not low-frequency stimulation of rat hippocampal slices

The release of adenosine and ATP evoked by electrical field stimulation in rat hippocampal slices was investigated with the following two patterns of stimulation: (1) a brief, high-frequency burst stimulation (trains of stimuli at 100 Hz for 50 ms applied every 2 s for 1 min), to mimic a long-term potentiation (LTP) stimulation paradigm, and (2) a more prolonged (3 min) and low-frequency (5 Hz) train stimulation, to mimic a long-term depression (LTD) stimulation paradigm. The release of ATP was greater at a brief, high-frequency burst stimulation, whereas the release of [3H]adenosine was slightly greater at a more prolonged and low-frequency stimulation. To investigate the source of extracellular adenosine, the following two pharmacological tools were used: alpha, beta-methylene ADP (AOPCP), an inhibitor of ecto-5'-nucleotidase, to assess the contribution of the catabolism of released adenine nucleotides as a source of extracellular adenosine, and S-(4-nitrobenzyl)-6-thioinosine (NBTI), an inhibitor of adenosine transporters, to assess the contribution of the release of adenosine, as such, as a source of extracellular adenosine. At low-frequency stimulation, NBTI inhibited by nearly 50% the evoked outflow of [3H]-adenosine, whereas AOPCP inhibited [3H]adenosine outflow only marginally. In contrast, at high-frequency stimulation, AOPCP inhibited by 30% the evoked release of [3H]adenosine, whereas NBTI produced a 40% inhibition of [3H]adenosine outflow. At both frequencies, the kinetics of evoked [3H]adenosine outflow was affected in different manners by AOPCP and NBTI; NBTI mainly depressed the rate of evoked [3H]adenosine outflow, whereas AOPCP mainly inhibited the later phase of evoked [3H]adenosine accumulation. These results show that there is a simultaneous, but quantitatively different, release of ATP and adenosine from rat hippocampal slices stimulated at frequencies that can induce plasticity phenomena such as LTP (100 Hz) or LTD (5 Hz). The source of extracellular adenosine is also different according to the frequency of stimulation; i.e., at a brief, high-frequency stimulation there is a greater contribution of released adenine nucleotides for the formation of extracellular adenosine than at a low frequency with a more prolonged stimulation.     

Influence of salivary components and extracellular polysaccharide synthesis from sucrose on the attachment of Streptococcus mutans 6715 to hydroxyapatite surfaces

The adsorption of (3)H-labeled Streptococcus mutans 6715 cells to disks of hydroxyapatite (HA) was studied. The number of streptococci that adsorbed was logarithmically related to the concentration of cells available up to at least 2 x 10(8) per ml; equilibrium occurred within 45 min. Assay reliability was verified by direct scanning electron microscopic counts. Untreated HA disks exposed to buffered saline (PBS)-suspended streptococci at a concentration of 1.1 x 10(8) per ml absorbed 3.2 x 10(6) cells per cm(2); approximately 3% of the surface area was, therefore, occupied by adsorbed organisms. The presence of adsorbed salivary components on HA reduced the number of attaching S. mutans cells by half. When S. mutans cells were suspended in saliva to mimic conditions existing in the mouth, the number of streptococci adsorbing to saliva-treated HA was reduced more than 30-fold compared to untreated HA. Approximately one-half of the streptococci adsorbed to untreated or to saliva-treated HA disks could be desorbed over a 4-h period with 0.067 M phosphate buffer. S. mutans cells exposed to sucrose to permit extracellular polysaccharide synthesis before or during adsorption attached in fewer numbers to both saliva-treated and untreated HA than PBS-treated organisms. When S. mutans cells adsorbed on untreated HA were exposed to sucrose, fewer organisms could be desorbed; thus, in situ polysaccharide synthesis promoted their more firm attachment to untreated HA. However, when saliva-suspended streptococci were adsorbed to saliva-treated HA surfaces, exposure to sucrose before or subsequent to adsorption did not promote more firm attachment. Evidently, the powerful adherence-inhibiting and desorptive effects of salivary components overshadowed any promoting effects attributable to glucan synthesis from sucrose. Similarly, no differences were noted in the desorption of S. mutans cells from human teeth after exposure to sucrose, glucose, or PBS relative to a strain of Streptococcus mitis (S. mitior). Thus, no evidence was obtained to support the hypothesis that glucan synthesis from sucrose was essential for, or promoted, the attachment of S. mutans cells to HA surfaces exposed to saliva or to the smooth surfaces of human teeth.     

Use of polyfilament suture as a source of microsuture

We present a case of reconstruction of the anterior leaflet in mitral valve prolapse and subacute bacterial endocarditis in which the resected prolapsing segment of the posterior leaflet was used as patch material. Competence of the valve was achieved with no recurrence of infection. Quadrangular resection of the posterior leaflet supplies presumably viable patch material for valve repair, which is particularly useful in bacterial endocarditis and when pliability is required.