Analysis of Automatically Generated Peptide Mass Fingerprints of Cellular Proteins and Antigens from Helicobacter pylori 26695 Separated by Two-dimensional Electrophoresis

Krah, Alexander; Schmidt, Frank; Becher, Dörte; Schmid, Monika S.; Albrecht, Dirk; Rack, Axel; Büttner, Knut; Jungblut, Peter R.

doi:10.1074/mcp.m300077-mcp200

Cited by 27 publications

(18 citation statements)

References 27 publications

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“…The identification of the GroEL protein with pI 5.55, MW 58264.4, Gene/ORF-TIGR HP0010 and access number P42383 (ExPASy -Swiss-Prot and TrEMBL® -http://us.expasy.org/sprot), at the reference proteome map 3-10, is preliminary information that indicates this protein could become a target for subsequent research. These results are similar to data described by KRAH et al 4 who reported that the pattern expression of GroEL was different in gastrointestinal diseases.…”

Section: Abstract: Helicobacter Pylori; Proteome; Bidimensional Electsupporting

confidence: 83%

Comparative analysis of two-dimensional electrophoresis maps (2-DE) of Helicobacter pylori from Brazilian patients with chronic gastritis and duodenal ulcer: a preliminary report

Pereira

Martins

Winck

et al. 2006

Rev. Inst. Med. trop. S. Paulo

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SUMMARYHelicobacter pylori is a bacterium recognized as the major cause of peptic ulcer and chronic gastritis. Recently, a proteomebased approach was developed to investigate pathogenic factors related to H. pylori. In this preliminary study, H. pylori strains were isolated from gastric biopsies of patients with chronic gastritis and duodenal ulcers. A partial proteomic analysis of H. pylori strains was performed by bacterial lyses and proteins were separated by two-dimensional gel electrophoresis (2-DE). A comparative analysis was performed to verify a differential protein expression between these two 2-DE maps. These data should be useful to clarify the role of different proteins related to bacterial pathogenesis. This study will be completed using a larger number of samples and protein identification of H. pylori by MALDI-TOF mass spectrometry. KEYWORDS:Helicobacter pylori; Proteome; Bidimensional electrophoresis; Chronic gastritis; Peptic ulcers.Helicobacter pylori is a pathogenic bacterium associated with the etiopathogenesis of universally spread chronic gastritis, peptic ulcers, gastric adenocarcinoma and MALT-lymphoma, mainly in developing countries 1,5 . Several of its biological aspects have not been revealed yet, in spite of molecular studies about this bacterium. A strategy to improve these studies consists in analyzing the whole protein expressed by the organism in its microenvironment. This is defined as proteome and the methodology is based around the technique of two-dimensional electrophoresis, which is directed towards display of proteins expressed inside gels, followed by protein identification using Peptide Mass Fingerprinting mass spectrometry (MALDI-TOF-MS) 7,8,9 .Recently, some reports have described a comparison among 2-DE proteome maps of clinical isolates obtained from patients who had peptic ulcers and chronic gastritis. It has been found that the main proteins have a different expression in these 2-DE maps, including GroEL (a protein that increases urease enzyme activity) and SodF (an enzyme involved in free radical and hydrogen peroxide catabolism) 2,3 . In this work, we carried out a comparative analysis of 2-DE maps of H. pylori from Brazilian patients, in order to find molecular targets related to its pathogenic mechanism in clinical isolates derived from duodenal ulcers and gastritis.H. pylori strains were isolated from gastric mucosa biopsies of two patients who had not been previously treated, submitted to upper gastrointestinal endoscopy at Gastrocentro, UNICAMP. Bacterial isolation and protein extraction were performed according to our previous reports 6,8 .The samples were submitted to isoelectrofocusing (IEF) solubilized with 8M urea, 4% CHAPS, 70 mM DTT, ampholine linear pH gradient 4-7 (pH 4-7L) 1.5% and 0.001% bromophenol blue. At this stage, 64 μg of each protein sample were applied to polyacrylamide dried strips, pH gradient 4-7 L (immobilized pH gradient -IPG). The first dimension of 2-DE was performed on an Amersham Biosciences Electrophoresis System accumulating 96 ...

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Section: Abstract: Helicobacter Pylori; Proteome; Bidimensional Electsupporting

confidence: 83%

Comparative analysis of two-dimensional electrophoresis maps (2-DE) of Helicobacter pylori from Brazilian patients with chronic gastritis and duodenal ulcer: a preliminary report

Pereira

Martins

Winck

et al. 2006

Rev. Inst. Med. trop. S. Paulo

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“…TsaA belongs to the five immunodominant H. pylori proteins (58). Pentameric arrangements of homodimers of TsaA were described in H. pylori cytoplasm (59,60). In our study, TsaA was found in homooligomeric form but also in a complex including proteins JHP1030 and PepQ.…”

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confidence: 60%

Two-dimensional Blue Native/SDS Gel Electrophoresis of Multiprotein Complexes from Helicobacter pylori

Pyndiah

Lasserre

Ménard

et al. 2007

Molecular & Cellular Proteomics

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The study of protein interactions constitutes an important domain to understand the physiology and pathogenesis of microorganisms. The two-dimensional blue native/SDS-PAGE was initially reported to analyze membrane protein complexes. In this study, both cytoplasmic and membrane complexes of a bacterium, the strain J99 of the gastric pathogen Helicobacter pylori, were analyzed by this method. It was possible to identify 34 different proteins grouped in 13 multiprotein complexes, 11 from the cytoplasm and two from the membrane, either previously reported partially or totally in the literature. Besides complexes involved in H. pylori physiology, this method allowed the description of interactions involving known pathogenic factors such as (i) urease with the heat shock protein GroEL or with the putative ketol-acid reductoisomerase IlvC and (ii) the cag pathogenicity island CagA protein with the DNA gyrase GyrA as well as insight on the partners of TsaA, a peroxide reductase/stress-dependent molecular chaperone. The two-dimensional blue native/ SDS-PAGE combined with mass spectrometry is a potential tool to study the differences in complexes isolated in various situations and also to study the interactions between bacterial and eucaryotic cell proteins. Molecular & Cellular Proteomics 6:193-206, 2007.

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“…2) (the pattern for fresh cultures is freely available at http://www .mpiib-berlin.mpg.de/2D-PAGE/) in agreement with previous low-resolution data (7). Peptide mass fingerprinting (9,10) allowed the identification of all 16 detectable rod-shape-specific proteins and 11 of the 14 coccoid-specific proteins (Table 1).…”

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confidence: 99%

Lack of Stage-Specific Proteins in Coccoid Helicobacter pylori Cells

et al. 2004

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Analysis of Automatically Generated Peptide Mass Fingerprints of Cellular Proteins and Antigens from Helicobacter pylori 26695 Separated by Two-dimensional Electrophoresis

Cited by 27 publications

References 27 publications

Comparative analysis of two-dimensional electrophoresis maps (2-DE) of Helicobacter pylori from Brazilian patients with chronic gastritis and duodenal ulcer: a preliminary report

Comparative analysis of two-dimensional electrophoresis maps (2-DE) of Helicobacter pylori from Brazilian patients with chronic gastritis and duodenal ulcer: a preliminary report

Two-dimensional Blue Native/SDS Gel Electrophoresis of Multiprotein Complexes from Helicobacter pylori

Lack of Stage-Specific Proteins in Coccoid Helicobacter pylori Cells

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