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Search Results: 1 - 10 of 330204 matches for " Mary S. Lipton "
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An Empirical Strategy for Characterizing Bacterial Proteomes across Species in the Absence of Genomic Sequences
Joshua E. Turse,Matthew J. Marshall,James K. Fredrickson,Mary S. Lipton,Stephen J. Callister
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0013968
Abstract: Global protein identification through current proteomics methods typically depends on the availability of sequenced genomes. In spite of increasingly high throughput sequencing technologies, this information is not available for every microorganism and rarely available for entire microbial communities. Nevertheless, the protein-level homology that exists between related bacteria makes it possible to extract biological information from the proteome of an organism or microbial community by using the genomic sequences of a near neighbor organism. Here, we demonstrate a trans-organism search strategy for determining the extent to which near-neighbor genome sequences can be applied to identify proteins in unsequenced environmental isolates. In proof of concept testing, we found that within a CLUSTAL W distance of 0.089, near-neighbor genomes successfully identified a high percentage of proteins within an organism. Application of this strategy to characterize environmental bacterial isolates lacking sequenced genomes, but having 16S rDNA sequence similarity to Shewanella resulted in the identification of 300–500 proteins in each strain. The majority of identified pathways mapped to core processes, as well as to processes unique to the Shewanellae, in particular to the presence of c-type cytochromes. Examples of core functional categories include energy metabolism, protein and nucleotide synthesis and cofactor biosynthesis, allowing classification of bacteria by observation of conserved processes. Additionally, within these core functionalities, we observed proteins involved in the alternative lactate utilization pathway, recently described in Shewanella.
Proteomic Detection of Non-Annotated Protein-Coding Genes in Pseudomonas fluorescens Pf0-1
Wook Kim,Mark W. Silby,Sam O. Purvine,Julie S. Nicoll,Kim K. Hixson,Matt Monroe,Carrie D. Nicora,Mary S. Lipton,Stuart B. Levy
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0008455
Abstract: Genome sequences are annotated by computational prediction of coding sequences, followed by similarity searches such as BLAST, which provide a layer of possible functional information. While the existence of processes such as alternative splicing complicates matters for eukaryote genomes, the view of bacterial genomes as a linear series of closely spaced genes leads to the assumption that computational annotations that predict such arrangements completely describe the coding capacity of bacterial genomes. We undertook a proteomic study to identify proteins expressed by Pseudomonas fluorescens Pf0-1 from genes that were not predicted during the genome annotation. Mapping peptides to the Pf0-1 genome sequence identified sixteen non-annotated protein-coding regions, of which nine were antisense to predicted genes, six were intergenic, and one read in the same direction as an annotated gene but in a different frame. The expression of all but one of the newly discovered genes was verified by RT-PCR. Few clues as to the function of the new genes were gleaned from informatic analyses, but potential orthologs in other Pseudomonas genomes were identified for eight of the new genes. The 16 newly identified genes improve the quality of the Pf0-1 genome annotation, and the detection of antisense protein-coding genes indicates the under-appreciated complexity of bacterial genome organization.
Comparative Bacterial Proteomics: Analysis of the Core Genome Concept
Stephen J. Callister, Lee Ann McCue, Joshua E. Turse, Matthew E. Monroe, Kenneth J. Auberry, Richard D. Smith, Joshua N. Adkins, Mary S. Lipton
PLOS ONE , 2008, DOI: 10.1371/journal.pone.0001542
Abstract: While comparative bacterial genomic studies commonly predict a set of genes indicative of common ancestry, experimental validation of the existence of this core genome requires extensive measurement and is typically not undertaken. Enabled by an extensive proteome database developed over six years, we have experimentally verified the expression of proteins predicted from genomic ortholog comparisons among 17 environmental and pathogenic bacteria. More exclusive relationships were observed among the expressed protein content of phenotypically related bacteria, which is indicative of the specific lifestyles associated with these organisms. Although genomic studies can establish relative orthologous relationships among a set of bacteria and propose a set of ancestral genes, our proteomics study establishes expressed lifestyle differences among conserved genes and proposes a set of expressed ancestral traits.
Genome Sequence of the Deltaproteobacterial Strain NaphS2 and Analysis of Differential Gene Expression during Anaerobic Growth on Naphthalene
Raymond J. DiDonato Jr.,Nelson D. Young,Jessica E. Butler,Kuk-Jeong Chin,Kim K. Hixson,Paula Mouser,Mary S. Lipton,Robert DeBoy,Barbara A. Methé
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0014072
Abstract: Anaerobic polycyclic hydrocarbon (PAH) degradation coupled to sulfate reduction may be an important mechanism for in situ remediation of contaminated sediments. Steps involved in the anaerobic degradation of 2-methylnaphthalene have been described in the sulfate reducing strains NaphS3, NaphS6 and N47. Evidence from N47 suggests that naphthalene degradation involves 2-methylnaphthalene as an intermediate, whereas evidence in NaphS2, NaphS3 and NaphS6 suggests a mechanism for naphthalene degradation that does not involve 2-methylnaphthalene. To further characterize pathways involved in naphthalene degradation in NaphS2, the draft genome was sequenced, and gene and protein expression examined.
Genome-Scale Modeling of Light-Driven Reductant Partitioning and Carbon Fluxes in Diazotrophic Unicellular Cyanobacterium Cyanothece sp. ATCC 51142
Trang T. Vu ,Sergey M. Stolyar ,Grigoriy E. Pinchuk,Eric A. Hill,Leo A. Kucek,Roslyn N. Brown,Mary S. Lipton,Andrei Osterman,Jim K. Fredrickson,Allan E. Konopka,Alexander S. Beliaev ,Jennifer L. Reed
PLOS Computational Biology , 2012, DOI: 10.1371/journal.pcbi.1002460
Abstract: Genome-scale metabolic models have proven useful for answering fundamental questions about metabolic capabilities of a variety of microorganisms, as well as informing their metabolic engineering. However, only a few models are available for oxygenic photosynthetic microorganisms, particularly in cyanobacteria in which photosynthetic and respiratory electron transport chains (ETC) share components. We addressed the complexity of cyanobacterial ETC by developing a genome-scale model for the diazotrophic cyanobacterium, Cyanothece sp. ATCC 51142. The resulting metabolic reconstruction, iCce806, consists of 806 genes associated with 667 metabolic reactions and includes a detailed representation of the ETC and a biomass equation based on experimental measurements. Both computational and experimental approaches were used to investigate light-driven metabolism in Cyanothece sp. ATCC 51142, with a particular focus on reductant production and partitioning within the ETC. The simulation results suggest that growth and metabolic flux distributions are substantially impacted by the relative amounts of light going into the individual photosystems. When growth is limited by the flux through photosystem I, terminal respiratory oxidases are predicted to be an important mechanism for removing excess reductant. Similarly, under photosystem II flux limitation, excess electron carriers must be removed via cyclic electron transport. Furthermore, in silico calculations were in good quantitative agreement with the measured growth rates whereas predictions of reaction usage were qualitatively consistent with protein and mRNA expression data, which we used to further improve the resolution of intracellular flux values.
Transcriptional and Translational Regulatory Responses to Iron Limitation in the Globally Distributed Marine Bacterium Candidatus Pelagibacter ubique
Daniel P. Smith,Joshua B. Kitner,Angela D. Norbeck,Therese R. Clauss,Mary S. Lipton,Michael S. Schwalbach,Laura Steindler,Carrie D. Nicora,Richard D. Smith,Stephen J. Giovannoni
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0010487
Abstract: Iron is recognized as an important micronutrient that limits microbial plankton productivity over vast regions of the oceans. We investigated the gene expression responses of Candidatus Pelagibacter ubique cultures to iron limitation in natural seawater media supplemented with a siderophore to chelate iron. Microarray data indicated transcription of the periplasmic iron binding protein sfuC increased by 16-fold, and iron transporter subunits, iron-sulfur center assembly genes, and the putative ferroxidase rubrerythrin transcripts increased to a lesser extent. Quantitative peptide mass spectrometry revealed that sfuC protein abundance increased 27-fold, despite an average decrease of 59% across the global proteome. Thus, we propose sfuC as a marker gene for indicating iron limitation in marine metatranscriptomic and metaproteomic ecological surveys. The marked proteome reduction was not directly correlated to changes in the transcriptome, implicating post-transcriptional regulatory mechanisms as modulators of protein expression. Two RNA-binding proteins, CspE and CspL, correlated well with iron availability, suggesting that they may contribute to the observed differences between the transcriptome and proteome. We propose a model in which the RNA-binding activity of CspE and CspL selectively enables protein synthesis of the iron acquisition protein SfuC during transient growth-limiting episodes of iron scarcity.
Fluctuations in Species-Level Protein Expression Occur during Element and Nutrient Cycling in the Subsurface
Michael J. Wilkins, Kelly C. Wrighton, Carrie D. Nicora, Kenneth H. Williams, Lee Ann McCue, Kim M. Handley, Chris S. Miller, Ludovic Giloteaux, Alison P. Montgomery, Derek R. Lovley, Jillian F. Banfield, Philip E. Long, Mary S. Lipton
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0057819
Abstract: While microbial activities in environmental systems play a key role in the utilization and cycling of essential elements and compounds, microbial activity and growth frequently fluctuates in response to environmental stimuli and perturbations. To investigate these fluctuations within a saturated aquifer system, we monitored a carbon-stimulated in situ Geobacter population while iron reduction was occurring, using 16S rRNA abundances and high-resolution tandem mass spectrometry proteome measurements. Following carbon amendment, 16S rRNA analysis of temporally separated samples revealed the rapid enrichment of Geobacter-like environmental strains with strong similarity to G. bemidjiensis. Tandem mass spectrometry proteomics measurements suggest high carbon flux through Geobacter respiratory pathways, and the synthesis of anapleurotic four carbon compounds from acetyl-CoA via pyruvate ferredoxin oxidoreductase activity. Across a 40-day period where Fe(III) reduction was occurring, fluctuations in protein expression reflected changes in anabolic versus catabolic reactions, with increased levels of biosynthesis occurring soon after acetate arrival in the aquifer. In addition, localized shifts in nutrient limitation were inferred based on expression of nitrogenase enzymes and phosphate uptake proteins. These temporal data offer the first example of differing microbial protein expression associated with changing geochemical conditions in a subsurface environment.
Establishing the Proteome of Normal Human Cerebrospinal Fluid
Steven E. Schutzer,Tao Liu,Benjamin H. Natelson,Thomas E. Angel,Athena A. Schepmoes,Samuel O. Purvine,Kim K. Hixson,Mary S. Lipton,David G. Camp II,Patricia K. Coyle,Richard D. Smith,Jonas Bergquist
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0010980
Abstract: Knowledge of the entire protein content, the proteome, of normal human cerebrospinal fluid (CSF) would enable insights into neurologic and psychiatric disorders. Until now technologic hurdles and access to true normal samples hindered attaining this goal.
Top-Down Characterization of the Post-Translationally Modified Intact Periplasmic Proteome from the Bacterium Novosphingobium aromaticivorans
Si Wu,Roslyn N. Brown,Samuel H. Payne,Da Meng,Rui Zhao,Nikola Toli?,Li Cao,Anil Shukla,Matthew E. Monroe,Ronald J. Moore,Mary S. Lipton,Ljiljana Pa?a-Toli?
International Journal of Proteomics , 2013, DOI: 10.1155/2013/279590
Abstract: The periplasm of Gram-negative bacteria is a dynamic and physiologically important subcellular compartment where the constant exposure to potential environmental insults amplifies the need for proper protein folding and modifications. Top-down proteomics analysis of the periplasmic fraction at the intact protein level provides unrestricted characterization and annotation of the periplasmic proteome, including the post-translational modifications (PTMs) on these proteins. Here, we used single-dimension ultra-high pressure liquid chromatography coupled with the Fourier transform mass spectrometry (FTMS) to investigate the intact periplasmic proteome of Novosphingobium aromaticivorans. Our top-down analysis provided the confident identification of 55 proteins in the periplasm and characterized their PTMs including signal peptide removal, N-terminal methionine excision, acetylation, glutathionylation, pyroglutamate, and disulfide bond formation. This study provides the first experimental evidence for the expression and periplasmic localization of many hypothetical and uncharacterized proteins and the first unrestrictive, large-scale data on PTMs in the bacterial periplasm. 1. Introduction The periplasm of Gram-negative bacteria is a hydrated gel located between the cytoplasmic and outer membranes and is comprised of peptidoglycan (cell wall), proteins, carbohydrates, and small solutes [1–3]. The periplasm is a dynamic subcellular compartment important for trafficking of molecules into and out of cells, maintaining cellular osmotic balance, envelope structure, responding to environmental cues and stresses, electron transport, xenobiotic metabolism, and protein folding and modification [4]. The periplasm provides a good model system to study protein biogenesis, composition, sorting, and modification at the molecular level. Indeed, it is analogous in many ways to the endoplasmic reticulum of eukaryotic cells in terms of transport, folding, and quality control [3]. Localization to the periplasm and beyond often involves an N-terminal secretion signal that targets the protein for translocation across the cytoplasmic membrane via the general secretory pathway [5]. These secretion signals (also known as signal peptides) are cleaved by signal peptidases located in the cytoplasmic membrane [6]. Thus, it is expected that signal peptide cleavage is a common modification in the periplasmic proteome. Compared to the cytoplasm, the periplasm is more vulnerable to changes in pH, temperature, and osmolarity in the external environment [4, 7, 8]. For structural stability in
Regulatory Response to Carbon Starvation in Caulobacter crescentus
Leticia Britos,Eduardo Abeliuk,Thomas Taverner,Mary Lipton,Harley McAdams,Lucy Shapiro
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0018179
Abstract: Bacteria adapt to shifts from rapid to slow growth, and have developed strategies for long-term survival during prolonged starvation and stress conditions. We report the regulatory response of C. crescentus to carbon starvation, based on combined high-throughput proteome and transcriptome analyses. Our results identify cell cycle changes in gene expression in response to carbon starvation that involve the prominent role of the FixK FNR/CAP family transcription factor and the CtrA cell cycle regulator. Notably, the SigT ECF sigma factor mediates the carbon starvation-induced degradation of CtrA, while activating a core set of general starvation-stress genes that respond to carbon starvation, osmotic stress, and exposure to heavy metals. Comparison of the response of swarmer cells and stalked cells to carbon starvation revealed four groups of genes that exhibit different expression profiles. Also, cell pole morphogenesis and initiation of chromosome replication normally occurring at the swarmer-to-stalked cell transition are uncoupled in carbon-starved cells.
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