bluuebunny/crossref_metadata_2025_split huggingface dataset:
An analysis of the proteomic profile for Thermoanaerobacter tengcongensis under optimal culture conditions
Authors: Wang Jingqiang , Xue Yanfen , Feng Xiaoli , Li Xiaolei , Wang Hao , Li Wei , Zhao Caifeng , Cheng Xiaojie , Ma Yanhe , Zhou Peijin , Yin Jianning , Bhatnagar Aruni , Wang Rong , Liu Siqi 
Published: January 2004

AbstractThe genome of Thermoanaerobacter tengcongensis is estimated to encode 2588 theoretical proteins. In this study, we have vitalized approximately 46% of the theoretical proteome experimentally using a proteomic strategy that combines three different methods, shotgun digestion plus high‐performance liquid chromatography (HPLC) with ion‐trap tandem mass spectrometry (shotgun‐liquid chromatography (LC)/MS), one‐dimensional sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) plus HPLC with ion‐trap tandem mass spectrometry (one‐dimensional electrophoresis (1DE)‐LC/MS), and two‐dimensional gel electrophoresis plus matrix‐assisted laser desorption/ionization‐time of flight‐mass spectrometry (2DE‐MALDI‐TOF‐MS). Of the 1200 proteins identified, as few as 76 proteins were globally found by all three approaches, and notably, most of these proteins were in the soluble fraction. However, there were a number of unique proteins detected by one method only, suggesting that our strategy provides a means toward obtaining a comprehensive view of protein expression profile. Proteins from the major metabolic pathways are strongly represented on the map, and a number of these enzymes were identified by more than one proteomic method. Based upon the proteins identified in the present study, we are able to broaden the understanding of how T. tengcongensis survives under high temperature environment, whereas several of its properties can not be fully explained by genome data.
In vivo uniform 15N‐isotope labelling of plants: Using the greenhouse for structural proteomics
Authors: Ippel Johannes H. , Pouvreau Laurice , Kroef Toos , Gruppen Harry , Versteeg Geurt , van den Putten Peter , Struik Paul C. , van Mierlo Carlo P. M. 
Published: January 2004

AbstractIsotope labelling of proteins is important for progress in the field of structural proteomics. It enables the utilisation of the power of nuclear magnetic resonance spectroscopy (NMR) for the characterisation of the three‐dimensional structures and corresponding dynamical features of proteins. The usual approach to obtain isotopically labelled protein molecules is by expressing the corresponding gene in bacterial or yeast host organisms, which grow on isotope‐enriched media. This method has several drawbacks. Here, we demonstrate that it is possible to fully label a plant with 15N‐isotopes. The advantage of in vivo labelling of higher organisms is that all constituting proteins are labelled and become available as functional, post‐translationally modified, correctly folded proteins. A hydroponics set‐up was used to create the first example of a uniformly 15N‐labelled (> 98%) plant species, the potato plant (Solanum tuberosum L., cv. Elkana). Two plants were grown at low costs using potassium‐[15N]‐nitrate as the sole nitrogen source. At harvest time, a total of 3.6 kg of potato tubers and 1.6 kg of foliage, stolons and roots were collected, all of which were fully 15N‐labelled. Gram quantities of soluble 15N‐labelled proteins (composed mainly of the glycoprotein patatin and Kunitz‐type protease inhibitors) were isolated from the tubers. NMR results on the complete proteome of potato sap and on an isolated protease inhibitor illustrate the success of the labelling procedure. The presented method of isotope labelling is easily modified to label other plants. Its envisioned impact in the field of structural proteomics of plants is discussed.
C‐Terminal sequencing method for peptides and proteins by the reaction with a vapor of perfluoric acid in acetic anhydride
Authors: Miyazaki Kenji , Tsugita Akira 
Published: January 2004

AbstractA successive C‐terminal amino acid truncation reaction of peptides and proteins with a vapor generated from a low‐concentrated perfluoric acid in acetic anhydride is presented. The reaction products were analyzed with matrix‐assisted laser desorption/ionization‐time of flight mass‐spectrometry giving molecular mass ions of the C‐terminal truncated peptides or proteins from which the C‐terminal sequence information can be deduced. Acetylation reaction preceded the truncation reaction in order to protect the amino groups and other reactive groups in peptides and proteins, and after the truncation reaction, hydration reaction was carried out to afford cleaner mass spectra.
Proteome characterization of human T helper 1 and 2 cells
Authors: Rautajoki Kirsi , Nyman Tuula A. , Lahesmaa Riitta 
Published: January 2004

AbstractT helper (Th) cells can be polarized into two different main subtypes, Th1 and Th2 cells. Their activation is linked to the eradication of different pathogens and to dissimilar immunological dysfunctions, which implies differences also in their protein expression patterns. To identify these differences, CD4+ T cells were isolated from human cord blood, polarized in vitro to Th1 and Th2 and activated via CD3 and CD28. Cells were lysed, soluble proteins were separated with two‐dimensional electrophoresis and differing protein spots were identified with peptide mass fingerprinting. The expression of 14 proteins differed in Th1 and Th2 cells after both 7 and 14 days of polarization. Twelve of the proteins could be identified, most of which are new in this context. Two proteins were differentially modified in the two cell types. Especially, N‐terminal acetylation of cyclophilin A was stronger in Th1 than in Th2 cells. To compare the RNA and the protein levels of the identified genes, mRNA expression was measured with Affymetrix oligonucleotide microarrays (HG‐U133A). The mRNA and protein expression level correlated only in six cases out of eleven, which highlights the complementary roles that proteomics and transcriptomics have in the elucidation of biological phenomena.
Proteogenomic mapping as a complementary method to perform genome annotation
Authors: Jaffe Jacob D. , Berg Howard C. , Church George M. 
Published: January 2004

AbstractThe accelerated rate of genomic sequencing has led to an abundance of completely sequenced genomes. Annotation of the open reading frames (ORFs) (i.e., gene prediction) in these genomes is an important task and is most often performed computationally based on features in the nucleic acid sequence. Using recent advances in proteomics, we set out to predict the set of ORFs for an organism based principally on expressed protein‐based evidence. Using a novel search strategy, we mapped peptides detected in a whole‐cell lysate of Mycoplasma pneumoniae onto a genomic scaffold and extended these “hits” into ORFs bound by traditional genetic signals to generate a “proteogenomic map”. We were able to generate an ORF model for M. pneumoniae strain FH using proteomic data with a high correlation to models based on sequence features. Ultimately, we detected over 81% of the genomically predicted ORFs in M. pneumoniae strain M129 (the originally sequenced strain). We were also able to detect several new ORFs not originally predicted by genomic methods, various N‐terminal extensions, and some evidence that would suggest that certain predicted ORFs are bogus. Some of these differences may be a result of the strain analyzed but demonstrate the robustness of protein analysis across closely related genomes. This technique is a cost‐effective means to add value to genome annotation, and a prerequisite for proteome quantitation and in vivo interaction measures.
Global changes in gene expression observed at the transition from growth to stationary phase in Listeria monocytogenes ScottA batch culture
Authors: Weeks Mark E. , James David C. , Robinson Gary K. , Smales C. Mark 
Published: January 2004

AbstractListeria monocytogenes is a food‐borne Gram‐positive bacterium that is responsible for a variety of infections (worldwide) annually. The organism is able to survive a variety of environmental conditions and stresses, however, the mechanisms by which L. monocytogenes adapts to environmental change are yet to be fully elucidated. An understanding of the mechanism(s) by which L. monocytogenes survives unfavourable environmental conditions will aid in developing new food processing methods to control the organism in foodstuffs. We have utilized a proteomic approach to investigate the response of L. monocytogenes batch cultures to the transition from exponential to stationary growth phase. Proteomic analysis showed that batch cultures of L. monocytogenes perceived stress and began preparations for stationary phase much earlier (approximately A600 = 0.75, mid‐exponential) than predicted by growth characteristics alone. Global analysis of the proteome revealed that the expression levels of more than 50% of all proteins observed changed significantly over a 7–9 h period during this transition phase. We have highlighted ten proteins in particular whose expression levels appear to be important in the early onset of the stationary phase. The significance of these findings in terms of functionality and the mechanistic picture are discussed.
Comparison of gene expression between left atria and left ventricles from non‐diseased humans
Authors: Tsubakihara Masako , Williams Neal K. , Keogh Anne , dos Remedios Cristobal G. 
Published: January 2004

AbstractWe examine the reliability and accuracy of gene array technology in analyzing differences in gene expression between human non‐diseased left atrium and left ventricle. We have used cDNA gene arrays and validated those data by carefully designed quantitative real‐time polymerase chain reaction (PCR). We have identified pitfalls using cDNA gene array technology based on comparisons with other gene array studies and with changes reported for the levels of expression of the genes corresponding to these cDNAs. The high error rate reported here underscores the cautionary comments reported by others in this field.
Proteomics as a tool to improve investigation of substantial equivalence in genetically modified organisms: The case of a virus‐resistant tomato
Authors: Corpillo Davide , Gardini Giulia , Vaira Anna Maria , Basso Manuela , Aime Silvio , Accotto Gian Paolo , Fasano Mauro 
Published: January 2004

AbstractAt present, the so‐called “substantial equivalence” is the only widely accepted criterion for deciding whether or not a transgenic food is, from an alimentary point of view, to be considered totally correspondent to the “traditional” one from which it derives. Although never exactly defined, it deals with a comparison between the chemical composition of the two foods. A more in‐depth analysis can be performed by one of the most suitable methods that allows for the simultaneous screening of many components without prior identification, the analysis of the proteome. As a model for testing this kind of approach, we compared protein expression of two types of tomato plants, having the same genetic background, except for a virus resistance trait introduced by genetic engineering. When proteins extracted from seedlings of the two types were analyzed by two‐dimensional electrophoresis, no significant differences, either qualitative or quantitative, were detected, indicating that in this case the expression of major proteins was unmodified by the genetic manipulation. Fifteen proteins were identified by peptide mass fingerprinting.
A detergent‐ and cyanogen bromide‐free method for integral membrane proteomics: Application to Halobacterium purple membranes and the human epidermal membrane proteome
Authors: Blonder Josip , Conrads Thomas P. , Yu Li‐Rong , Terunuma Atsushi , Janini George M. , Issaq Haleem J. , Vogel Jonathan C. , Veenstra Timothy D. 
Published: January 2004

AbstractA simple and rapid method for characterizing hydrophobic integral membrane proteins and its utility for membrane proteomics using microcapillary liquid chromatography coupled on‐line with tandem mass spectrometry (μLC‐MS/MS) is described. The present technique does not rely on the use of detergents, strong organic acids or cyanogen bromide‐mediated proteolysis. A buffered solution of 60% methanol was used to extract, solubilize, and tryptically digest proteins within a preparation of Halobacterium (H.) halobium purple membranes. Analysis of the digested purple membrane proteins by μLC‐MS/MS resulted in the identification of all the predicted tryptic peptides of bacteriorhodopsin, including those that are known to be post‐translationally modified. In addition, 40 proteins from the purple membrane preparation were also identified, of which 80% are predicted to contain between 1 and 16 transmembrane domains. To evaluate the general applicability of the method, the same extraction, solubilization, and digestion conditions were applied to a plasma membrane fraction prepared from human epidermal sheets. A total of 117 proteins was identified in a single μLC‐MS/MS analysis, of which 55% are known to be integral or associated with the plasma membrane. Due to its simplicity, efficiency, and absence of MS interfering compounds, this technique can be used for the characterization of other integral membrane proteins and may be concomitantly applied for the analysis of membrane protein complexes or large‐scale proteomic studies of different membrane samples.
Nonreducing two‐dimensional gel electrophoresis for the detection of Bence Jones proteins in serum and urine
Authors: Miller Ingrid , Teinfalt Monika , Leschnik Michael , Wait Robin , Gemeiner Manfred 
Published: January 2004

AbstractNonreducing two‐dimensional gel electrophoresis (2‐DE) is described for the study of immunoglobulin disorders with asynchronous production of single chains. Unlike classical reducing 2‐DE, this method can distinguish between complex intact molecules and their free single chains (with different degrees of polymerization) and will thus be helpful for diagnosis of this type of disease. Examples are taken from canine patients, but the method may also be applied to both urine and serum specimens from other species. Nonreducing 2‐DE thus represents a useful tool complementary to classical 2‐DE, when further information about the appearance of free subunits or modifications of proteins are required, even in the presence of intact molecules.
Use of two‐dimensional gel electrophoresis proteome reference maps of dinoflagellates for species recognition of causative agents of harmful algal blooms
Authors: Chan Leo Lai , Hodgkiss Ivor John , Lu Songhui , Lo Samuel Chun‐Lap 
Published: January 2004

AbstractThe sample preparation procedures established for Prorocentrum triestinum were adapted to cover both thecate and athecate dinoflagellates. Further, whether trichloroacetic acid (TCA) precipitation can be used to fix and preserve the harmful or nuisance species from local waters that they infest was tested. Optimized technical procedures developed were used to generate proteome reference maps for eight other local causative species of harmful algal blooms (HABs): Prorocentrum micans, Prorocentrum minimum, Prorocentrum sigmoides, Prorocentrum dentatum, Scrippsiella trochoidea, Karenia longicanalis, Karenia digitata and Karenia mikimotoi; together with one American species Karenia brevis (Florida, USA). These proteome maps were used to test their ability for species recognition in a mixed culture of dinoflagellates and whether such investigations will provide a comparative view at a global level. Comparisons of proteome profiles were made (i) between closely related species within the same family; (ii) between distantly related species belonging to different types, i.e., gymnodinioids, prorocentroids or peridinioids, or (iii) between different groups, i.e., thecate (armored) dinoflagellate cells against athecate (naked or unarmored) dinoflagellate cells. Species‐specific two‐dimensional electrophoresis (2‐DE) protein profiles were observed in all ten species and it was possible to distinguish between even closely related species within the same family. To demonstrate the extent of reproducibility and usefulness of these 2‐DE reference maps, 2‐DE has been used to analyze three geographically distinct isolates of Prorocentrum dentatum, and to distinguish species composition in a mixed culture. Application of 2‐D PAGE analysis to differentiate between taxonomically confused strains of a single species could be a powerful taxonomic tool.
Immunostaining with dissociable antibody microarrays
Authors: Wang Yingjian 
Published: January 2004

AbstractThe availability of a large number of biological materials such as cDNA, antibodies, recombinant proteins, and tissues has promoted the development of microarray technologies that make use of these materials in high‐throughput screening assays. However, because microarray technologies have been less successful in examining proteins than DNA and mRNA , there is a need for improved protein microarray systems. To address this need, we developed an antibody microarray‐based immunostaining method that can analyze the properties of a large number of proteins simultaneously. In this method, antibodies are arrayed and immobilized on a solid support and cells bearing antigens of interest are attached to a second support. Apposition of the two supports allows the antibodies to dissociate from the array support and bind to the cellular antigens. After separation of the supports, antigen‐bound antibodies can be detected by standard secondary antibody techniques. These “dissociable” antibody arrays were used to detect both the expression and subcellular localization of a large number of specific proteins in various cultured cell types.
Identification of tumor‐associated proteins in oral tongue squamous cell carcinoma by proteomics
Authors: He Qing‐Yu , Chen Jia , Kung Hsiang‐Fu , Yuen Anthony Po‐Wing , Chiu Jen‐Fu 
Published: January 2004

AbstractOral tongue carcinoma is an aggressive tumor that particularly affects chronic smokers, drinkers and betel squid chewers. Patients often present symptoms at a late stage, and there is a high recurrence rate after treatment. In this article, we report the first proteomic analysis of oral tongue carcinoma to globally search for tumor related proteins. Apart from helping us to understand the molecular pathogenesis of the carcinoma, these proteins may also have potential clinical applications as biomarkers, enabling the tumor to be identified at an early stage in high risk individuals, treatment response to be predicted, and residual or recurrent carcinoma to be detected sooner after treatment. The protein expression profiles of ten oral tongue squamous cell carcinomas and their matched normal mucosal resection margins were examined by two‐dimensional gel electrophoresis and matrix‐assisted laser desorption/ionization‐time of flight mass spectroscopy. A number of tumor‐associated proteins including heat shock protein (HSP)60, HSP27, α B‐crystalline, ATP synthase β, calgranulin B, myosin, tropomyosin and galectin 1 were consistently found to be significantly altered in their expression levels in tongue carcinoma tissues, compared with their paired normal mucosae. The expression profile portrays a global protein alteration that appears specific to oral tongue cancer. The potential of utilizing these tumor related proteins for screening cancer and monitoring recurrence warrants further investigation.
Global analysis of the Ralstonia metallidurans proteome: Prelude for the large‐scale study of heavy metal response
Authors: Noël‐Georis Isabelle , Vallaeys Tatiana , Chauvaux Renaud , Monchy Sébastien , Falmagne Paul , Mergeay Max , Wattiez Ruddy 
Published: January 2004

AbstractA proteome map of Ralstonia metallidurans strain CH34 was constructed using two‐dimensional (2‐D) gel electrophoresis in combination with automated Edman degradation and mass spectrometry (MS). R. metallidurans CH34 is the type‐strain of a family of highly related strains characterized by their multiple resistance to millimolar amounts of heavy metals, conferred by two large plasmids. The protein content of this bacterium grown in minimal medium was separated by 2‐D gel electrophoresis using various pH gradients. Protein identification was carried out via N‐terminal amino acid sequencing, matrix assisted laser desorption/ionisation‐time of flight‐mass spectrometry (MALDI‐TOF‐MS) and tandem MS. So far, 224 different proteins were characterized from 352 protein spots. Although the proteome map is still not complete, one could appraise the importance of proteomics for genome analyses through (i) the identification of previously undetected open reading frames, (ii) the identification of proteins not encoded by the already sequenced genome fragments, (iii) the characterization of protein‐encoding genes spanning two different contigs, enabling their merging, and (iv) the precise delineation of the N‐terminus of several proteins. Finally, this map will prove a useful tool in the identification of proteins differentially expressed in the presence of different heavy metals.
Proteome comparative analysis of gynogenetic haploid and diploid embryos of goldfish (Carassius auratus)
Authors: Huang Lingyun , Li Bingxia , Luo Chen , Xie Jinyun , Chen Ping , Liang Songping 
Published: January 2004

AbstractRecently, it was found that in the gynogenetic haploid and diploid embryos of goldfish, which have exactly the same genome, the haploid condition results in obstruction of gene expression and abnormal development while the diploid embryos have normal gene expression and development. A diploid‐dependent regulatory apparatus was proposed to regulate gene expression. To study the difference at the protein expression level of the embryos of haploid and diploid in development, we extracted the total proteins of both the gynogenetic haploid and diploid embryos of goldfish in the same eye formation stage. Two‐dimensional polyacrylamide gel electrophoresis was used to separate proteins. The stained gel images were analyzed with the PDQUEST software. A part of protein spots that were differentially expressed in haploid and diploid embryos were identified by matrix assisted laser desorption/ionisation‐time of flight‐mass spectrometry and database analysis. Sixteen protein spots that were absolutely different (only expressed in diploid embryos but not in haploid embryos or vice versa) and 16 protein spots that were up‐ and downregulated were identified unambiguously, which include some proteins that are correlative with eyes development, nerve development, developing regulation, cell differentiation, and signal transduction. The different significantly gene expression during embryos developing between diploid and haploid is demonstrated.
Optimization of the first dimension for separation by two‐dimensional gel electrophoresis of basic proteins from human brain tissue
Authors: Pennington Kyla , McGregor Emma , Beasley Clare L. , Everall Ian , Cotter David , Dunn Michael J. 
Published: January 2004

AbstractA major cause of poor resolution in the alkaline pH range of two‐dimensional electrophoresis (2‐DE) gels is unsatisfactory separation of basic proteins in the first dimension. We have compared methods for the separation of basic proteins in the isoelectric focusing dimension of human brain proteins. The combined use of anodic cup‐loading and the hydroxyethyldisulphide containing solution (DeStreak) produced better resolution in both analytical and micropreparative protein loaded 2‐DE gels than the other methods investigated.