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161.
Giardia lamblia is a protozoan parasite with many characteristics common among eukaryotic cells, but lacking other features found in most eukaryotes. Cardiolipin is a phospholipid located exclusively in energy transducing membranes and it was identified in mitochondria, bacteria, hydrogenosomes and chloroplasts. In eukaryotes, cardiolipin is the only lipid that is synthesized in the mitochondria. Biochemical procedures (TLC, HPLC) and fluorescent tools (NAO) were applied in order to search for cardiolipin in G. lamblia. In addition, BLAST searches were used to find homologs of enzymes that participate in the cardiolipin synthesis. Cardiolipin synthase was searched in the Giardia genome, using Saccharomyces cerevisiae and Mycoplasma penetrans sequences as bait. However, a good match to G. lamblia related proteins was not found. Here we show that mitosomes of G. lamblia apparently do not contain cardiolipin, which raises the discussion for its endosymbiotic origin and for the previous proposal that Giardia mitosomes are modified mitochondria. 相似文献
162.
Sansom FM Riedmaier P Newton HJ Dunstone MA Müller CE Stephan H Byres E Beddoe T Rossjohn J Cowan PJ d'Apice AJ Robson SC Hartland EL 《The Journal of biological chemistry》2008,283(19):12909-12918
Legionella pneumophila is the predominant cause of Legionnaires disease, a severe and potentially fatal form of pneumonia. Recently, we identified an ecto-nucleoside triphosphate diphosphohydrolase (NTPDase) from L. pneumophila, termed Lpg1905, which enhances intracellular replication of L. pneumophila in eukaryotic cells. Lpg1905 is the first prokaryotic member of the CD39/NTPDase1 family of enzymes, which are characterized by the presence of five apyrase conserved regions and the ability to hydrolyze nucleoside tri- and diphosphates. Here we examined the substrate specificity of Lpg1905 and showed that apart from ATP and ADP, the enzyme catalyzed the hydrolysis of GTP and GDP but had limited activity against CTP, CDP, UTP, and UDP. Based on amino acid residues conserved in the apyrase conserved regions of eukaryotic NTPDases, we generated five site-directed mutants, Lpg1905E159A, R122A, N168A, Q193A, and W384A. Although the mutations E159A, R122A, Q193A, and W384A abrogated activity completely, N168A resulted in decreased activity caused by reduced affinity for nucleotides. When introduced into the lpg1905 mutant strain of L. pneumophila, only N168A partially restored the ability of L. pneumophila to replicate in THP-1 macrophages. Following intratracheal inoculation of A/J mice, none of the Lpg1905 mutants was able to restore virulence to an lpg1905 mutant during lung infection, thereby demonstrating the importance of NTPDase activity to L. pneumophila infection. Overall, the kinetic studies undertaken here demonstrated important differences to mammalian NTPDases and different sensitivities to NTPDase inhibitors that may reflect underlying structural variations. 相似文献
163.
Kátia Gisele Oliveira Rancura Michelli Rivero Montaño Robson Francisco Carvalho Cesar Martins Adriane Pinto Wasko Luciana Casaletti Alexandre Azevedo 《Acta zoologica》2008,89(1):29-36
This study presents data on myosin Va localization in the central nervous system of rainbow trout. We demonstrate, via immunoblots and immunocytochemistry, the expression of myosin Va in several neuronal populations of forebrain, midbrain, hindbrain and spinal cord. The neuronal populations that express myosin Va in trout constitute a very diverse group that do not seem to have many specific similarities such as neurotransmitters used, cellular size or length of their processes. The intensity of the immunoreactivity and the number of immunoreactive cells differ from region to region. Although there is a broad distribution of myosin Va, it is not present in all neuronal populations. This result is in agreement with a previous report, which indicated that myosin Va is approximately as abundant as conventional myosin II and kinesin, and it is broadly involved in neuronal motility events such as axoplasmatic transport. Furthermore, this distribution pattern is in accordance with what was shown in rats and mice; it indicates phylogenetic maintenance of the myosin Va main functions. 相似文献
164.
165.
Heme oxygenase expression in human placenta and placental bed: reduced expression of placenta endothelial HO-2 in preeclampsia and fetal growth restriction. 总被引:12,自引:0,他引:12
In this study we tested the hypothesis that expression of heme oxygenases HO-1 and HO-2, which are responsible for the production of carbon monoxide, are reduced in the placenta and placental bed of pregnancies complicated by preeclampsia (PE) and fetal growth restriction (FGR) compared with control third-trimester pregnancies. Placental protein expression was determined by Western blotting (n=10 in each group) and immunohistochemistry (controls n=18, PE n=19, FGR n=10). Extravillous trophoblast expression was determined by immunohistochemistry of placental bed biopsy samples (controls n=17, PE n=19, FGR n=10). Western blot analysis of placental homogenates showed no overall differences in HO-2 among groups. However, immunohistochemical analysis showed a reduction in HO-2 expression in endothelial cells in both abnormal groups (PE P<0.01; FGR P<0.0005 vs. control group) but no differences in villous trophoblast staining. HO-1 was undetectable by Western blotting in control and abnormal pregnancies and immunoreactivity was very low, suggesting that there is little HO-1 in the placenta. Within the placental bed, HO-2 but not HO-1 was detected on all populations of extravillous trophoblast, but expression of HO-2 or HO-1 did not change in PE or FGR. The reduced expression of HO-2 on endothelial cells in PE and FGR may be responsible for reduced placental blood flow in these conditions. The data do not show changes in HO in the placental bed in PE or FGR. 相似文献
166.
Characterization of the ethanol-inducible alc gene-expression system in Arabidopsis thaliana 总被引:4,自引:0,他引:4
Hairul A. Roslan Michael G. Salter Chris D. Wood Michael R. H. White Kevan P. Croft Frances Robson George Coupland John Doonan Patrick Laufs A. Brian Tomsett Mark X. Caddick 《The Plant journal : for cell and molecular biology》2001,28(2):225-235
Controlled expression of transgenes in plants is key to the characterization of gene function and the regulated manipulation of growth and development. The alc gene-expression system, derived from the filamentous fungus Aspergillus nidulans, has previously been used successfully in both tobacco and potato, and has potential for use in agriculture. Its value to fundamental research is largely dependent on its utility in Arabidopsis thaliana. We have undertaken a detailed function analysis of the alc regulon in A. thaliana. By linking the alcA promoter to beta-glucuronidase (GUS), luciferase (LUC) and green fluorescent protein (GFP) genes, we demonstrate that alcR-mediated expression occurs throughout the plant in a highly responsive manner. Induction occurs within one hour and is dose-dependent, with negligible activity in the absence of the exogenous inducer for soil-grown plants. Direct application of ethanol or exposure of whole plants to ethanol vapour are equally effective means of induction. Maximal expression using soil-grown plants occurred after 5 days of induction. In the majority of transgenics, expression is tightly regulated and reversible. We describe optimal strategies for utilizing the alc system in A. thaliana. 相似文献
167.
Natalia Cadaxo Rochael Luize Gon?alves Lima Sandra Maria Pereira de Oliveira Marcello André Barcinski Elvira Maria Saraiva Robson Queiroz Monteiro Lucia Helena Pinto-da-Silva 《Memórias do Instituto Oswaldo Cruz》2013,108(6):679-685
Leishmania parasites expose phosphatidylserine (PS) on their
surface, a process that has been associated with regulation of host''s immune
responses. In this study we demonstrate that PS exposure by metacyclic
promastigotes of Leishmania amazonensis favours blood
coagulation. L. amazonensis accelerates in vitro coagulation of
human plasma. In addition, L. amazonensis supports the assembly
of the prothrombinase complex, thus promoting thrombin formation. This process
was reversed by annexin V which blocks PS binding sites. During blood meal,
Lutzomyia longipalpis sandfly inject saliva in the bite
site, which has a series of pharmacologically active compounds that inhibit
blood coagulation. Since saliva and parasites are co-injected in the host during
natural transmission, we evaluated the anticoagulant properties of sandfly
saliva in counteracting the procoagulant activity of L.
amazonensis . Lu. longipalpis saliva reverses
plasma clotting promoted by promastigotes. It also inhibits thrombin formation
by the prothrombinase complex assembled either in phosphatidylcholine (PC)/PS
vesicles or in L. amazonensis . Sandfly saliva inhibits factor
X activation by the intrinsic tenase complex assembled on PC/PS vesicles and
blocks factor Xa catalytic activity. Altogether our results show that metacyclic
promastigotes of L. amazonensis are procoagulant due to PS
exposure. Notably, this effect is efficiently counteracted by sandfly
saliva. 相似文献
168.
Summary
Azotobacter chroococcum Fos 189 is a Tn1-induced mutant which, unlike the parent strain MCD1, does not fix nitrogen in air when provided with glucose or pyruvate as sole carbon sources. Fos 189 showed 5% of parental activity for phosphoenolpyruvate carboxylase though PEP synthetase activity was normal. The A. chroococcum phosphoenolpyruvate carboxylase (ppc) gene was isolated after complementation of an appropriate Escherichia coli mutant using a broad host range gene bank prepared from A. chroococcum genomic DNA. The gene was localised by transposon mutagenesis and subcloning on a minimum DNA fragment of 6.6 kb. Broad host range plasmids containing the A. chroococcum ppc gene complemented the mutation in Fos 189 thereby restoring aerotolerant nitrogen fixation. 相似文献
169.
The application of the Oribital-centred Force Field (OFF) to the calculation of the conformational behaviour of biomolecules is further considered. This method includes classical type potential functions between atom centres, but also specifies the location of non-core orbital lobes and their strength of electrostatic interaction with other lobes or atom centres. It is shown that the OFF method is not particularly advantageous for peptide systems, since although non-core orbital lobes are present with typical electrostatic strength, they do not dominate the potential surface. For other systems, however, of which sulphonamide is used as an example, the interactions between non-core orbitals are the major contribution, as was earlier shown to be the case for the nucleic acid backbone. An interesting test case is the dioxy system OXO, for which a priori the OFF approach may be inadequate because of an anomeric effect discussed by other authors interested in saccharide systems. The possibility that an intrinsic rotational potential may be required is considered but remains by no means obvious (the need for such potentials is normally absent when the OFF approach is used). Finally, the applications of the OFF approach are illustrated and tested by reference to the conformational analysis of 5′-AMP and to the computer-aided model building of DNA helices. 相似文献
170.
Non-Uniform Sampling has the potential to exploit the optimal resolution of high-field NMR instruments. This is not possible in 3D and 4D NMR experiments when using traditional uniform sampling due to the long overall measurement time. Nominally, uniformly sampled time domain data acquired to a maximum evolution time tmax can be extended to high resolution via a virtual maximum evolution time t*max while extrapolating with linear prediction or iterative soft thresholding (IST). At the high resolution obtainable with extrapolation of US data, however, the accuracy of peak positions is compromised as observed when comparing inter- and intra-residue peaks in a 3D HNCA experiment. However, the accuracy of peak positions is largely improved by spreading the same number of acquired time domain data points non-uniformly over a larger evolution time to an optimal tmax followed by extrapolation to a total t*max and processing the data with an appropriate reconstruction method, such as hmsIST. To explore the optimum value of experimentally measured tmax to be reached non-uniformly with a given number of sampling points we have created test situations of time-equivalent experiments and evaluate sensitivity and accuracy of peak positions. Here we use signal-to-maximum-noise ratio as the decisive measure of sensitivity. We find that both sensitivity and resolution are optimal when PoissonGap sampling to a tmax of about ½*T2 *. Digital resolution is further enhanced by extrapolating the range of acquired time domain data to 2*T2 * but without measuring experimental points beyond ½*T2 *. 相似文献