127 research outputs found
Automatic multi-banking of memory for microprocessors
A microprocessor system is provided with added memories to expand its address spaces beyond its address word length capacity by using indirect addressing instructions of a type having a detectable operations code and dedicating designated address spaces of memory to each of the added memories, one space to a memory. By decoding each operations code of instructions read from main memory into a decoder to identify indirect addressing instructions of the specified type, and then decoding the address that follows in a decoder to determine which added memory is associated therewith, the associated added memory is selectively enabled through a unit while the main memory is disabled to permit the instruction to be executed on the location to which the effective address of the indirect address instruction points, either before the indirect address is read from main memory or afterwards, depending on how the system is arranged by a switch
Compact artificial hand
A relatively simple, compact artificial hand, is described which includes hooks pivotally mounted on first frame to move together and apart. The first frame is rotatably mounted on a second frame to enable "turning at the wrist" movement without limitation. The second frame is pivotally mounted on a third frame to permit 'flexing at the wrist' movement. A hook-driving motor is fixed to the second frame but has a shaft that drives a speed reducer on the first frame which, in turn, drives the hooks. A second motor mounted on the second frame, turns a gear on the first frame to rotate the first frame and the hooks thereon. A third motor mounted on the third frame, turns a gear on a second frame to pivot it
Comparison of membrane proteins of Mycobacterium tuberculosis H37Rv and H37Ra strains
Background: The potential causes for variation in virulence between distinct M. tuberculosis strains are still not fully known. However, differences in protein expression are probably an important factor. In this study we used a labelfree quantitative proteomic approach to estimate differences in protein abundance between two closely related M. tuberculosis strains; the virulent H37Rv strain and its attenuated counterpart H37Ra. Results: We were able to identify more than 1700 proteins from both strains. As expected, the majority of the identified proteins had similar relative abundance in the two strains. However, 29 membrane-associated proteins were observed with a 5 or more fold difference in their relative abundance in one strain compared to the other. Of note, 19 membrane- and lipo-proteins had higher abundance in H37Rv, while another 10 proteins had a higher abundance in H37Ra. Interestingly, the possible protein-export membrane protein SecF (Rv2586c), and three ABCtransporter proteins (Rv0933, Rv1273c and Rv1819c) were among the more abundant proteins in M. tuberculosis H37Rv. Conclusion: Our data suggests that the bacterial secretion system and the transmembrane transport system may be important determinants of the ability of distinct M. tuberculosis strains to cause disease
Definition of novel cell envelope associated proteins in Triton X-114 extracts of Mycobacterium tuberculosis H37Rv
<p>Abstract</p> <p>Background</p> <p>Membrane- and membrane-associated proteins are important for the pathogenicity of bacteria. We have analysed the content of these proteins in virulent <it>Mycobacterium tuberculosis </it>H37Rv using Triton X-114 detergent-phase separation for extraction of lipophilic proteins, followed by their identification with high resolution mass spectrometry.</p> <p>Results</p> <p>In total, 1417 different proteins were identified. <it>In silico </it>analysis of the identified proteins revealed that 248 proteins had at least one predicted trans-membrane region. Also, 64 of the identified proteins were predicted lipoproteins, and 54 proteins were predicted as outer membrane proteins. Three-hundred-and-ninety-five of the observed proteins, including 91 integral membrane proteins were described for the first time. Comparison of abundance levels of the identified proteins was performed using the exponentially modified protein abundance index (emPAI) which takes into account the number of the observable peptides to the number of experimentally observed peptide ions for a given protein. The outcome showed that among the membrane-and membrane-associated proteins several proteins are present with high relative abundance. Further, a close examination of the lipoprotein LpqG (Rv3623) which is only detected in the membrane fractions of <it>M. tuberculosis </it>but not in <it>M. bovis</it>, revealed that the homologous gene in <it>M. bovis </it>lack the signal peptide and lipobox motif, suggesting impaired export to the membrane.</p> <p>Conclusions</p> <p>Altogether, we have identified a substantial proportion of membrane- and membrane-associated proteins of <it>M. tuberculosis </it>H37Rv, compared the relative abundance of the identified proteins and also revealed subtle differences between the different members of the <it>M. tuberculosis </it>complex.</p
Allergic sensitisation in tuberculosis patients at the time of diagnosis and following chemotherapy
<p>Abstract</p> <p>Background</p> <p>It is still a matter of debate whether there is an association between infection with <it>Mycobacterium tuberculosis </it>(<it>M. tuberculosis</it>) and allergy. Previously, we have shown higher levels of specific IgE to different inhalant allergens and total IgE in tuberculosis (TB) patients compared to controls. The objectives of this study were to evaluate a possible change in allergic sensitisation after successful TB treatment and to confirm the finding of our previous study of enhanced allergic sensitisation in TB patients compared to controls in a more controlled setting. Additionally, we wanted to determine the cytokine profile in the same groups and finally to evaluate the association between the presence of Bacillus Calmette-Guérin vaccination (BCG) scar and allergic sensitisation among the controls.</p> <p>Methods</p> <p>Sera were analysed for specific IgE to inhalant allergens (Phadiatop) and total IgE by the use of ImmunoCAP 1000 (Pharmacia Diagnostics). Thirteen different cytokines were also analysed in the sera by multiplex bead immunoassay (Luminex 100, Luminex Corporation), and clinical symptoms of allergy and BCG scar were reported in a questionnaire.</p> <p>Results</p> <p>A reduction in levels of specific and total IgE were observed after successful TB treatment. TB patients also had higher levels of specific and total IgE compared to healthy controls. Both interleukin (IL)-6 and interferon (IFN)γ were higher in TB patients compared to healthy controls. The levels of IL-6 were reduced after successful TB treatment. The presence of a BCG scar was associated with a reduced risk of developing allergic sensitisation.</p> <p>Conclusion</p> <p>We observed a reduced level of allergic sensitisation after successful TB treatment. TB patients seem to be more allergically sensitised than healthy controls, confirming our previous finding. Furthermore, we observed an inverse association between allergic sensitisation and visible BCG scar, which adds additional support to the hygiene hypothesis.</p
High accuracy mass spectrometry analysis as a tool to verify and improve gene annotation using Mycobacterium tuberculosis as an example
Background: While the genomic annotations of diverse lineages of the Mycobacterium tuberculosis complex are available, divergences between gene prediction methods are still a challenge for unbiased protein dataset generation. M. tuberculosis gene annotation is an example, where the most used datasets from two independent institutions (Sanger Institute and Institute of Genomic Research-TIGR) differ up to 12% in the number of annotated open reading frames, and 46% of the genes contained in both annotations have different start codons. Such differences emphasize the importance of the identification of the sequence of protein products to validate each gene annotation including its sequence coding area. Results: With this objective, we submitted a culture filtrate sample from M. tuberculosis to a highaccuracy LTQ-Orbitrap mass spectrometer analysis and applied refined N-terminal prediction to perform comparison of two gene annotations. From a total of 449 proteins identified from the MS data, we validated 35 tryptic peptides that were specific to one of the two datasets, representing 24 different proteins. From those, 5 proteins were only annotated in the Sanger database. In the remaining proteins, the observed differences were due to differences in annotation of transcriptional start sites. Conclusion: Our results indicate that, even in a less complex sample likely to represent only 10% of the bacterial proteome, we were still able to detect major differences between different gene annotation approaches. This gives hope that high-throughput proteomics techniques can be used to improve and validate gene annotations, and in particular for verification of high-throughput, automatic gene annotations.publishedVersio
Proteomic analysis of total cellular proteins of human neutrophils
This is an Open Access article distributed under the terms of the Creative Commons Attribution Licens
Evaluation of signal peptide prediction algorithms for identification of mycobacterial signal peptides using sequence data from proteomic methods
Secreted proteins play an important part in the pathogenicity of Mycobacterium tuberculosis, and are the primary source of vaccine and diagnostic candidates. A majority of these proteins are exported via the signal peptidase I-dependent pathway, and have a signal peptide that is cleaved off during the secretion process. Sequence similarities within signal peptides have spurred the development of several algorithms for predicting their presence as well as the respective cleavage sites. For proteins exported via this pathway, algorithms exist for eukaryotes, and for Gram-negative and Gram-positive bacteria. However, the unique structure of the mycobacterial membrane raises the question of whether the existing algorithms are suitable for predicting signal peptides within mycobacterial proteins. In this work, we have evaluated the performance of nine signal peptide prediction algorithms on a positive validation set, consisting of 57 proteins with a verified signal peptide and cleavage site, and a negative set, consisting of 61 proteins that have an N-terminal sequence that confirms the annotated translational start site. We found the hidden Markov model of SignalP v3.0 to be the best-performing algorithm for predicting the presence of a signal peptide in mycobacterial proteins. It predicted no false positives or false negatives, and predicted a correct cleavage site for 45 of the 57 proteins in the positive set. Based on these results, we used the hidden Markov model of SignalP v3.0 to analyse the 10 available annotated proteomes of mycobacterial species, including annotations of M. tuberculosis H37Rv from the Wellcome Trust Sanger Institute and the J. Craig Venter Institute (JCVI). When excluding proteins with transmembrane regions among the proteins predicted to harbour a signal peptide, we found between 7.8 and 10.5 % of the proteins in the proteomes to be putative secreted proteins. Interestingly, we observed a consistent difference in the percentage of predicted proteins between the Sanger Institute and JCVI. We have determined the most valuable algorithm for predicting signal peptidase I-processed proteins of M. tuberculosis, and used this algorithm to estimate the number of mycobacterial proteins with the potential to be exported via this pathway
Phosphoproteomics analysis of a clinical mycobacterium tuberculosis Beijing isolate : expanding the mycobacterial phosphoproteome catalog
CITATION: Fortuin, S., et al. 2015. Phosphoproteomics analysis of a clinical mycobacterium tuberculosis beijing isolate : expanding the mycobacterial phosphoproteome catalog. Frontiers in Microbiology, 6:6, doi:10.3389/fmicb.2015.00006.The original publication is available at www.frontiersin.orgReversible protein phosphorylation, regulated by protein kinases and phosphatases, mediates a switch between protein activity and cellular pathways that contribute to a large number of cellular processes. The Mycobacterium tuberculosis genome encodes 11 Serine/Threonine kinases (STPKs) which show close homology to eukaryotic kinases. This study aimed to elucidate the phosphoproteomic landscape of a clinical isolate of M. tuberculosis. We performed a high throughput mass spectrometric analysis of proteins extracted from an early-logarithmic phase culture. Whole cell lysate proteins were processed using the filter-aided sample preparation method, followed by phosphopeptide enrichment of tryptic peptides by strong cation exchange (SCX) and Titanium dioxide (TiO2) chromatography. The MaxQuant quantitative proteomics software package was used for protein identification. Our analysis identified 414 serine/threonine/tyrosine phosphorylated sites, with a distribution of S/T/Y sites; 38% on serine, 59% on threonine and 3% on tyrosine; present on 303 unique peptides mapping to 214 M. tuberculosis proteins. Only 45 of the S/T/Y phosphorylated proteins identified in our study had been previously described in the laboratory strain H37Rv, confirming previous reports. The remaining 169 phosphorylated proteins were newly identified in this clinical M. tuberculosis Beijing strain. We identified 5 novel tyrosine phosphorylated proteins. These findings not only expand upon our current understanding of the protein phosphorylation network in clinical M. tuberculosis but the data set also further extends and complements previous knowledge regarding phosphorylated peptides and phosphorylation sites in M. tuberculosis.http://journal.frontiersin.org/article/10.3389/fmicb.2015.00006/fullPublisher's versio
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