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41.
Dorner GT Garhofer G Kiss B Polska E Polak K Riva CE Schmetterer L 《American journal of physiology. Heart and circulatory physiology》2003,285(2):H631-H636
The purpose of the present study was to investigate the contribution of basal nitric oxide (NO) on retinal vascular tone in humans. In addition, we set out to elucidate the role of NO in flicker-induced retinal vasodilation in humans. Twelve healthy young subjects were studied in a three-way crossover design. Subjects received an intravenous infusion of either placebo or NG-monomethyl-L-arginine (L-NMMA; 3 or 6 mg/kg over 5 min), an inhibitor of NO synthase. Thereafter, diffuse luminance flicker was consecutively performed for 16, 32, and 64 s at a frequency of 8 Hz. The effect of L-NMMA on retinal arterial and venous diameter was assessed under resting conditions and during the hyperemic flicker response. Retinal vessel diameter was measured with a Zeiss retinal vessel analyzer. L-NMMA significantly reduced arterial diameter (3 mg/kg: -2%; 6 mg/kg: -4%, P < 0.001) and venous diameter (3 mg/kg: -5%; 6 mg/kg: -8%, P < 0.001). After placebo infusion, flicker induced a significant increase in retinal vessel diameter (P < 0.001). At a flicker duration of 64 s, arterial diameter increased by 4% and venous diameter increased by 3%. L-NMMA did not abolish these hyperemic responses but blunted venous vasodilation (P = 0.017) and arterial vasodilation (P = 0.02) in response to flicker stimulation. Our data indicate that NO contributes to basal retinal vascular tone in humans. In addition, NO appears to play a role in flicker-induced vasodilation of the human retinal vasculature. 相似文献
42.
The structure of the yeast DNA-dependent RNA polymerase I (RNA Pol I), prepared by cryo-negative staining, was studied by electron microscopy. A structural model of the enzyme at a resolution of 1.8 nm was determined from the analysis of isolated molecules and showed an excellent fit with the atomic structure of the RNA Pol II Delta4/7. The high signal-to-noise ratio (SNR) of the stained molecular images revealed a conformational flexibility within the image data set that could be recovered in three-dimensions after implementation of a novel strategy to sort the "open" and "closed" conformations in our heterogeneous data set. This conformational change mapped in the "wall/flap" domain of the second largest subunit (beta-like) and allows a better accessibility of the DNA-binding groove. This displacement of the wall/flap domain could play an important role in the transition between initiation and elongation state of the enzyme. Moreover, a protrusion was apparent in the cryo-negatively stained model, which was absent in the atomic structure and was not detected in previous 3D models of RNA Pol I. This structure could, however, be detected in unstained views of the enzyme obtained from frozen hydrated 2D crystals, indicating that this novel feature is not induced by the staining process. Unexpectedly, negatively charged molybdenum compounds were found to accumulate within the DNA-binding groove, which is best explained by the highly positive electrostatic potential of this region of the molecule, thus, suggesting that the stain distribution reflects the overall surface charge of the molecule. 相似文献
43.
Simerská P Kuzma M Pisvejcová A Weignerová L Macková M Riva S Kren V 《Folia microbiologica》2003,48(3):329-337
4-Nitrophenyl alpha-D-galactopyranosyl-(1-->3)-6-O-acetyl-alpha-D-galactopyranoside was prepared in a transglycosylation reaction catalyzed by alpha-D-galactosidase from Talaromyces flavus using 4-nitrophenyl alpha-D-galactopyranoside as a glycosyl donor and 4-nitrophenyl 6-O-acetyl-alpha-D-galactopyranoside as an acceptor. 4-Nitrophenyl 6-O-acetyl-alpha-D-galactopyranoside and 4-nitrophenyl 6-O-acetyl-beta-D-galactopyranoside were prepared in a regioselective enzymic transesterification in pyridine-acetone catalyzed by the lipase PS from Burkholderia cepacia. A series of water-miscible organic solvents (acetone, acetonitrile, dimethylformamide, dimethyl sulfoxide, 1,4-dioxane, 2-methoxyethanol, pyridine, 2-methylpropan-2-ol, tetrahydrofuran, propargyl alcohol) were used as co-solvents in this enzymic reaction. Their influence on the activity and stability of the alpha-galactosidase from T. flavus was established. 2-Methylpropan-2-ol and acetone (increasing the solubility of the modified substrate acceptors and displaying the minimum impairment of the activity and stability of the enzyme) were used as co-solvents in transglycosylation reactions. 相似文献
44.
Franzoni L Lücke C Pérez C Cavazzini D Rademacher M Ludwig C Spisni A Rossi GL Rüterjans H 《The Journal of biological chemistry》2002,277(24):21983-21997
Retinoid-binding proteins play an important role in regulating transport, storage, and metabolism of vitamin A and its derivatives. The solution structure and backbone dynamics of rat cellular retinol-binding protein type I (CRBP) in the apo- and holo-form have been determined and compared using multidimensional high resolution NMR spectroscopy. The global fold of the protein is consistent with the common motif described for members of the intracellular lipid-binding protein family. The most relevant difference between the NMR structure ensembles of apo- and holoCRBP is the higher backbone disorder, in the ligand-free form, of some segments that frame the putative entrance to the ligand-binding site. These comprise alpha-helix II, the subsequent linker to beta-strand B, the hairpin turn between beta-strands C and D, and the betaE-betaF turn. The internal backbone dynamics, obtained from 15N relaxation data (T1, T2, and heteronuclear nuclear Overhauser effect) at two different fields, indicate several regions with significantly higher backbone mobility in the apoprotein, including the betaC-betaD and betaE-betaF turns. Although apoCRBP contains a binding cavity more shielded than that of any other retinoid carrier, conformational flexibility in the portal region may assist retinol uptake. The stiffening of the backbone in the holoprotein guarantees the stability of the complex during retinol transport and suggests that targeted retinol release requires a transiently open state that is likely to be promoted by the acceptor or the local environment. 相似文献
45.
Bischler N Brino L Carles C Riva M Tschochner H Mallouh V Schultz P 《The EMBO journal》2002,21(15):4136-4144
The spatial distribution of four subunits specifically associated to the yeast DNA-dependent RNA polymerase I (RNA pol I) was studied by electron microscopy. A structural model of the native enzyme was determined by cryo-electron microscopy from isolated molecules and was compared with the atomic structure of RNA pol II Delta 4/7, which lacks the specific polypeptides. The two models were aligned and a difference map revealed four additional protein densities present in RNA pol I, which were characterized by immunolabelling. A protruding protein density named stalk was found to contain the RNA pol I-specific subunits A43 and A14. The docking with the atomic structure showed that the stalk protruded from the structure at the same site as the C-terminal domain (CTD) of the largest subunit of RNA pol II. Subunit A49 was placed on top of the clamp whereas subunit A34.5 bound at the entrance of the DNA binding cleft, where it could contact the downstream DNA. The location of the RNA pol I-specific subunits is correlated with their biological activity. 相似文献
46.
SNPper: retrieval and analysis of human SNPs 总被引:4,自引:0,他引:4
MOTIVATION: Single Nucleotide Polymorphisms (SNPs) are an increasingly important tool for the study of the human genome. SNPs can be used as markers to create high-density genetic maps, as causal candidates for diseases, or to reconstruct the history of our genome. SNP-based studies rely on the availability of large numbers of validated, high-frequency SNPs whose position on the chromosomes is known with precision. Although large collections of SNPs exist in public databases, researchers need tools to effectively retrieve and manipulate them. RESULTS: We describe the implementation and usage of SNPper, a web-based application to automate the tasks of extracting SNPs from public databases, analyzing them and exporting them in formats suitable for subsequent use. Our application is oriented toward the needs of candidate-gene, whole-genome and fine-mapping studies, and provides several flexible ways to present and export the data. The application has been publicly available for over a year, and has received positive user feedback and high usage levels. 相似文献
47.
Human chromosomes 9, 12, and 15 contain the nucleation sites of stress-induced nuclear bodies 下载免费PDF全文
Denegri M Moralli D Rocchi M Biggiogera M Raimondi E Cobianchi F De Carli L Riva S Biamonti G 《Molecular biology of the cell》2002,13(6):2069-2079
We previously reported the identification of a novel nuclear compartment detectable in heat-shocked HeLa cells that we termed stress-induced Src-activated during mitosis nuclear body (SNB). This structure is the recruitment center for heat shock factor 1 and for a number of RNA processing factors, among a subset of Serine-Arginine splicing factors. In this article, we show that stress-induced SNBs are detectable in human but not in hamster cells. By means of hamster>human cell hybrids, we have identified three human chromosomes (9, 12, and 15) that are individually able to direct the formation of stress bodies in hamster cells. Similarly to stress-induced SNB, these bodies are sites of accumulation of hnRNP A1-interacting protein and heat shock factor 1, are usually associated to nucleoli, and consist of clusters of perichromatin granules. We show that the p13-q13 region of human chromosome 9 is sufficient to direct the formation of stress bodies in hamster>human cell hybrids. Fluorescence in situ hybridization experiments demonstrate that the pericentromeric heterochromatic q12 band of chromosome 9 and the centromeric regions of chromosomes 12 and 15 colocalize with stress-induced SNBs in human cells. Our data indicate that human chromosomes 9, 12, and 15 contain the nucleation sites of stress bodies in heat-shocked HeLa cells. 相似文献
48.
Gene expression of ABC proteins in hepatocellular carcinoma,perineoplastic tissue,and liver diseases 总被引:11,自引:0,他引:11
Bonin S Pascolo L Crocé LS Stanta G Tiribelli C 《Molecular medicine (Cambridge, Mass.)》2002,8(6):318-325
BACKGROUND: The development of hepatocellular carcinoma (HCC) is a frequent event during the natural history of cirrhosis. Effective treatment is, however, hampered by drug resistance related to the expression of multidrug resistance (MDR) proteins belonging to the ABC family transporters. Studying expression of genes coding for these proteins may help to explain the potential sensitivity of HCC to chemotherapy. MATERIAL AND METHODS: The expression of MRP1, MRP2, MRP3, MDR1, and MDR3 was investigated by quantitative RT-PCR analyses in paraffin-embedded tissues obtained from 9 cases of HCC, 16 cases of cirrhosis, 10 cases of chronic extrahepatic cholestasis, and 16 cases of normal liver. In HCC cases, gene expression was assessed both in neoplastic and perineoplastic tissue after microscopically assisted microdissection. RESULTS: MRP1 was significantly and similarly overexpressed in HCC and perineoplastic tissue. MRP2 and MDR1 were also increased in HCC, but the level of expression did not correlate with that of perineoplastic tissue. The level of expression was either reduced or normal in cirrhotic liver and during chronic cholestasis. Expression of MDR3 was unchanged in all conditions investigated. CONCLUSIONS: The genetic expression of multi-drug resistance proteins, in particular MRP1, MRP2, and MDR1, is increased during HCC. In the case of MRP1, the extent of expression is similar in neoplastic and perineoplastic tissue, but this is not the case for MRP2 and MDR1. The assessment of ABC protein expression pattern may provide important information for the diagnosis and treatment of HCC. 相似文献
49.
Chagas disease, caused by the parasite protozoan Trypanosoma cruzi, is characterised by a variable clinical course, from symptomless cases to severe chronic disease with cardiac and/or gastrointestinal involvement. This variability has been attributed both to differences in the host response and to genomic heterogeneity of the parasite. This article reviews the evidence in favour of an important role of the genetic constitution of T. cruzi in determining the clinical characteristics of Chagas disease and discusses the basis of the 'Clonal-Histotropic Model' for the pathogenesis of this disease. 相似文献
50.
Fraceto LF Pinto Lde M Franzoni L Braga AA Spisni A Schreier S de Paula E 《Biophysical chemistry》2002,99(3):229-243
We examined the effect of uncharged lidocaine on the structure and dynamics of egg phosphatidylcholine (EPC) membranes at pH 10.5 in order to assess the location of this local anesthetic in the bilayer. Changes in the organization of small unilamellar vesicles were monitored either by electron paramagnetic resonance (EPR)-in the spectra of doxyl derivatives of stearic acid methyl esters labeled at different positions in the acyl chain (5-, 7-, 12- and 16-MeSL)-or by fluorescence, with pyrene fatty-acid (4-, 6-, 10- and 16-Py) probes. The largest effects were observed with labels located at the upper positions of the fatty-acid acyl-chain. Dynamic information was obtained by 1H-NMR. Lidocaine protons presented shorter longitudinal relaxation times (T(1)) values due to their binding, and consequent immobilization to the membrane. In the presence of lidocaine the mobility of all glycerol protons of EPC decreased, while the choline protons revealed a higher degree of mobility, indicating a reduced participation in lipid-lipid interactions. Two-dimensional Nuclear Overhauser Effect experiments detected contacts between aromatic lidocaine protons and the phospholipid-choline methyl group. Fourier-transform infrared spectroscopy spectra revealed that lidocaine changes the access of water to the glycerol region of the bilayer. A "transient site" model for lidocaine preferential location in EPC bilayers is proposed. The model is based on the consideration that insertion of the bulky aromatic ring of the anesthetic into the glycerol backbone region causes a decrease in the mobility of that EPC region (T(1) data) and an increased mobility of the acyl chains (EPR and fluorescence data). 相似文献