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101.
Cassandra A. Louw Michael H. Ludewig Gregory L. Blatch 《Protein expression and purification》2010,69(2):168-177
The heat shock protein 40 (Hsp40) family of proteins act as co-chaperones of the heat shock protein 70 (Hsp70) chaperone family, and together they play a vital role in the maintenance of cellular homeostasis. The Type III class of Hsp40s are diverse in terms of both sequence identity and function and have not been extensively characterised. The Trypanosoma brucei parasite is the causative agent of Human African Trypanosomiasis, and possesses an unusually large Hsp40 complement, consisting mostly of Type III Hsp40s. A novel T. brucei Type III Hsp40, Tbj1, was heterologously expressed, purified, and found to exist as a compact monomer in solution. Using polyclonal antibodies to the full-length recombinant protein, Tbj1 was found by Western analysis to be expressed in the T. brucei bloodstream-form. Tbj1 was found to be able to assist two different Hsp70 proteins in the suppression of protein aggregation in vitro, despite being unable to stimulate their ATPase activity. This indicated that while Tbj1 did not possess independent chaperone activity, it potentially functioned as a novel co-chaperone of Hsp70 in T. brucei. 相似文献
102.
Anna Bachmann Michaela Petter Ann-Kathrin Tilly Laura Biller Karin A. Uliczka Michael F. Duffy Egbert Tannich Iris Bruchhaus 《PloS one》2012,7(11)
Avoidance of antibody-mediated immune recognition allows parasites to establish chronic infections and enhances opportunities for transmission. The human malaria parasite Plasmodium falciparum possesses a number of multi-copy gene families, including var, rif, stevor and pfmc-2tm, which encode variant antigens believed to be expressed on the surfaces of infected erythrocytes. However, most studies of these antigens are based on in vitro analyses of culture-adapted isolates, most commonly the laboratory strain 3D7, and thus may not be representative of the unique challenges encountered by P. falciparum in the human host. To investigate the expression of the var, rif-A, rif-B, stevor and pfmc-2tm family genes under conditions that mimic more closely the natural course of infection, ex vivo clinical P. falciparum isolates were analyzed using a novel quantitative real-time PCR approach. Expression patterns in the clinical isolates at various time points during the first intraerythrocytic developmental cycle in vitro were compared to those of strain 3D7. In the clinical isolates, in contrast to strain 3D7, there was a peak of expression of the multi-copy gene families rif-A, stevor and pfmc-2tm at the young ring stage, in addition to the already known expression peak in trophozoites. Furthermore, most of the variant surface antigen families were overexpressed in the clinical isolates relative to 3D7, with the exception of the pfmc-2tm family, expression of which was higher in 3D7 parasites. Immunofluorescence analyses performed in parallel revealed two stage-dependent localization patterns of RIFIN, STEVOR and PfMC-2TM. Proteins were exported into the infected erythrocyte at the young trophozoite stage, whereas they remained inside the parasite membrane during schizont stage and were subsequently observed in different compartments in the merozoite. These results reveal a complex pattern of expression of P. falciparum multi-copy gene families during clinical progression and are suggestive of diverse functional roles of the respective proteins. 相似文献
103.
Ladwig F Stahl M Ludewig U Hirner AA Hammes UZ Stadler R Harter K Koch W 《Plant physiology》2012,158(4):1643-1655
Many membrane proteins are involved in the transport of nutrients in plants. While the import of amino acids into plant cells is, in principle, well understood, their export has been insufficiently described. Here, we present the identification and characterization of the membrane protein Siliques Are Red1 (SIAR1) from Arabidopsis (Arabidopsis thaliana) that is able to translocate amino acids bidirectionally into as well as out of the cell. Analyses in yeast and oocytes suggest a SIAR1-mediated export of amino acids. In Arabidopsis, SIAR1 localizes to the plasma membrane and is expressed in the vascular tissue, in the pericycle, in stamen, and in the chalazal seed coat of ovules and developing seeds. Mutant alleles of SIAR1 accumulate anthocyanins as a symptom of reduced amino acid content in the early stages of silique development. Our data demonstrate that the SIAR1-mediated export of amino acids plays an important role in organic nitrogen allocation and particularly in amino acid homeostasis in developing siliques. 相似文献
104.
Weber SS Joller N Küntzel AB Spörri R Tchang VS Scandella E Rösli C Ludewig B Hilbi H Oxenius A 《Journal of immunology (Baltimore, Md. : 1950)》2012,189(2):841-849
Abs confer protection from secondary infection with Legionella pneumophila, the causative agent of a severe form of pneumonia known as Legionnaires' disease. In this study, we demonstrate that Ab-mediated protection is effective across L. pneumophila serogroups, suggesting that Abs specific for conserved protein Ags are sufficient to mediate this protective effect. We used two independent methods to identify immunogenic L. pneumophila protein Ags, namely, the screening of a λ phage library representing the complete L. pneumophila genome and two-dimensional gel electrophoresis combined with Western blot analysis and protein spot identification by mass spectrometry. A total of 30 novel L. pneumophila B cell Ags were identified, the majority of which are located in or associated with the bacterial membrane, where they are accessible for Abs and, therefore, likely to be relevant for Ab-mediated protection against L. pneumophila. Selected B cell Ags were recombinantly expressed and tested in a vaccination protocol. Mice immunized with either single-protein Ags or an Ag combination showed reduced bacterial titers in bronchoalveolar lavage and lung after L. pneumophila challenge. To determine the clinical relevance of these findings, we tested Legionnaires' disease patient sera for reactivity with the identified L. pneumophila Ags. The recognized Ags were indeed conserved across host species, because Abs specific for all three selected Ags could be detected in patient sera, rendering the identified protein Ags potential vaccine candidates. 相似文献
105.
Semi-natural grassland soils are frequently fertilised for agricultural improvement. This practice often comes at a loss of
the indigenous flora while fast-growing nitrogen-responsive species, such as Lolium perenne, take over. Since soil microbial communities depend on plant root exudates for carbon and nitrogen sources, this shift in
vegetation is thought to influence soil microbial community structure. In this study, we investigated the influence of different
plant species, fertilisation and L. perenne ingression on microbial communities in soils from three semi-natural Irish grasslands. Bacterial and fungal community compositions
were determined by automated ribosomal intergenic spacer analysis, and community changes were linked to environmental factors
by multivariate statistical analysis. Soil type had a strong effect on bacterial and fungal communities, mainly correlated
to soil pH, as well as soil carbon and nitrogen status. Within each soil type, plant species composition was the main influencing
factor followed by nitrogen fertilisation and finally Lolium ingression in the acidic upland and mesotrophic grassland. In the alkaline grassland, however, Lolium ingression had a stronger effect than fertilisation. Our results suggest that a change in plant species diversity strongly
influences the microbial community structure, which may subsequently lead to significant changes in ecosystem functioning. 相似文献
106.
A new fluorogenic, lanthanide-based oligopeptide substrate for the detection of the zinc-dependent endoprotease thermolysin is described. Using time-resolved fluorescence measurement, a highly sensitive assay for thermolysin was developed with a 50 pM detection limit (3.5 fmol). 相似文献
107.
J. Susanne Hauswaldt Ann-Kathrin Ludewig Sabine Hagemann Heike Pröhl Miguel Vences 《Conservation Genetics》2009,10(6):1935-1937
We describe primers and PCR conditions to amplify nine new tetranucleotide loci and one new dinucleotide locus isolated from
the strawberry poison frog (Oophaga pumilio). In 21 individuals from Costa Rica, the number of alleles ranged from 4 to 16, observed heterozygosities from 40 to 100%,
and polymorphic information content ranged from 0.60 to 0.90 per locus. Evidence for linkage disequilibrium was found only
between two loci, but this pattern was not found in other populations tested. All primer pairs cross-amplified in Oophaga vicentei from Panama. 相似文献
108.
109.
Aquaporins and/or aquaglyceroporins regulate the permeability of plant membranes to water and small, uncharged molecules. Using molecular simulations with a plant plasma membrane aquaporin tetramer, the residues in the channel constriction region were identified as the crucial determinants of ammonia and urea conductance. The impact of these residues was experimentally verified using AtPIP2;1 pore mutants. Several, but not all, mutants with a NIP-like selectivity filter promoted yeast growth on urea or ammonia as sole sources of nitrogen. TIP-like mutants conducted urea but not NH(3), and a residue without direct contact to the pore lumen was critical for conduction in the mutants. 相似文献
110.