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Proteomics-based method for the assessment of marine pollution using liquid chromatography coupled with two-dimensional electrophoresis
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
2007 (English)In: Journal of Proteome Research, ISSN 1535-3893, E-ISSN 1535-3907, Vol. 6, no 6, 2094-2104 p.Article in journal (Refereed) Published
Abstract [en]

Using a proteomic approach, we have developed a new method for the assessment of marine pollution that generates highly reproducible protein expression patterns and it is simple and scalable. The protocol is based on applying liquid chromatography (LC) coupled with two-dimensional electrophoresis (2-DE) to analyze changes in the protein expression pattern after exposure to marine pollution. The digestive gland of the sentinel “blue mussel” (Mytilus edulis) was batch-processed through a simple cell fractionation followed by ion-exchange chromatography and 2-DE. The selection of ligands, elution method, and small volume design was carefully considered to define a protocol that could be mainly robotized. A pilot study with samples collected from different Gothenburg harbor areas indicated that the clean area could be distinguished from the polluted ones based on a protein expression pattern (PES) composed of 13 proteins. Principal component analysis (PCA) and hierarchical clustering confirmed that the PES was sufficient to discriminate polluted and unpolluted areas and to provide a spatial gradient from the polluted source. Several proteins from the PES were identified by electrospray ionization tandem mass spectrometry (ESI−MS/MS), and they are involved in β-oxidation, amino acid metabolism, detoxification, protein degradation, organelle biogenesis, and protein folding. In the near future, this methodology could show potential advantages to assess marine pollution and could become a stable platform to elucidate ecotoxicological questions.

Place, publisher, year, edition, pages
2007. Vol. 6, no 6, 2094-2104 p.
Keyword [en]
Amino Acid Sequence, Animals, Cell Fractionation, Chromatography; Ion Exchange/methods, Electrophoresis; Gel; Two-Dimensional/methods, Molecular Sequence Data, Mytilus edulis/*chemistry, Oceans and Seas, Proteins/*analysis, Proteomics/*methods, Spectrometry; Mass; Electrospray Ionization, Water Pollution; Chemical
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:su:diva-11250DOI: 10.1021/pr060689sISI: 000246893500004OAI: oai:DiVA.org:su-11250DiVA: diva2:177769
Available from: 2008-01-10 Created: 2008-01-10 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Development and application of a proteomic approach to the assessment of pollution in the marine environment
Open this publication in new window or tab >>Development and application of a proteomic approach to the assessment of pollution in the marine environment
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Today, assessment of the health of coastal waters is recognized as being important for both the conservation of nature and well-being of humans. Anthropogenic pollution has been the focus of extensive research for some time and a variety of programs for the monitoring and assessment of environmental pollution have been developed. Determination of the levels of pollution in sensitive ‘sentinels’ such as mussels, allows monitoring of these levels in a given area over a prolonged period of time. Furthermore, the biological effects of pollution are reflected in a series of biomarkers, none of which provides a general picture of the sentinel’s state of health and all of which are individually specific for certain pollutants and influenced by both biotic and abiotic factors.

In an attempt to improve biomonitoring of marine pollution, we have developed two proteomic approaches here. In the first portion of the thesis, a proteomic analysis was performed on peroxisomes isolated from mussels exposed either to one of three model anthropogenic pollutants, or two different types of crude oil, or from mussels exposed to the Prestige oil spill. Application of two-dimensional electrophoresis (2-DE) provided protein expression signatures (PES) for exposure to these different pollutants.Furthermore, several individual protein components of these PES could be putatively identified.

In the second portion of this work, such analysis of subproteomes was developed further in order to improve the applicability of this approach to biomonitoring. A simple fractionation procedure in combination with liquid chromatography and 2-DE provided samples from mussels residing in different regions of a pollution gradient around the harbor of Gothenburg, as well as from mussels exposed to two types of fuel oil similar to that of the Prestige that were suitable for environmental proteomics. In addition, we constructed a model for this approach that can be cross-validated in the future and applied to assess sources of fuel oil pollution in connection with biomonitoring programs.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2009. 58 p.
Keyword
proteomics, mussels, peroxisomes, two-dimensional electrophoresis (2-DE), difference gel electrophoresis (DIGE), bisphenol A (BPA), diallyl phthalate (DAP), 2, 2′, 4, 4′-tetrabromodiphenyl ether (PBDE-47), fuel oil, Prestige oil spill
National Category
Environmental Sciences Pharmacology and Toxicology
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-26150 (URN)978-91-7155-801-5 (ISBN)
Public defence
2009-04-17, Magnélisalen , Kemiska övningslaboratoriet, Svante Arrhenius väg 12A, Stockholm University, Stockholm, 14:00 (English)
Opponent
Supervisors
Available from: 2009-03-26 Created: 2009-03-10 Last updated: 2009-04-02Bibliographically approved
2. Proteomics in biomarker research: Insights into the effects of aging and environment on biological systems
Open this publication in new window or tab >>Proteomics in biomarker research: Insights into the effects of aging and environment on biological systems
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Proteomics is the global analysis of proteins that covers a broad range of technologies aimed at determining the identity and quantity of proteins expressed in the cell, their three-dimensional structure and interaction partners. In contrast to genome, proteome reflects more accurately on the dynamic state of the cell, tissue, or an organism. Therefore much is expected from proteomics to yield better disease markers for early diagnosis and therapy monitoring, as well as biomarkers that would indicate environmental exposure or provide prediction of biological age.

In this thesis, I have developed and applied robust and sensitive subproteomic approaches to study the effect of aging as well as and environmental pollution using different animal models.

In the first part, a high-throughput proteomic method based on liquid chromatography coupled to 2-dimensional gel electrophoresis (LC/2-DE) was developed. The usefulness of this method has been demonstrated by applying it to the assessment of marine pollution in a field experiment.

Next, I have utilized this subproteomic approach to study the effect of aging in mouse kidney of both genders. As a result, a protein expression signature of aging kidney was obtained, revealing gender-dependent alterations in proteome profiles of aging mouse kidney.

In order to further reduce the dynamic range of protein expression and increase the sensitivity of proteomic analysis, I have applied a shotgun mass spectrometry-based proteomic approach using isobaric tags for relative and absolute quantification (iTRAQ) coupled to liquid chromatography and tandem mass spectrometry (LC-MS/MS) to study age-related differences in peroxisome-enriched fractions from mouse liver. Only eight proteins showed statistically significant difference in expression (p<0.05) with moderate folds. This study indicates that age-depended changes in the liver proteome are minimal, suggesting that its proteome is efficiently maintained until certain age.

Finally, in the context of aging studies and the role of peroxisomes in aging, I have tested the utility of cell-penetrating peptides (CPPs) as agents for protein delivery into acatalasemic peroxisomes using yeast as a model. The results obtained suggest that CPPs may be suitable for the delivery of antioxidants to peroxisomes and in future could provide a tool for the protein therapy of age-related diseases.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2011. 63 p.
Keyword
proteomics, biomarker, peroxisomes, 2-dimensional gel electrophoresis (2-DE), mass spectrometry (MS), aging, marine pollution
National Category
Biochemistry and Molecular Biology Environmental Sciences Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-56483 (URN)978-91-7447-209-7 (ISBN)
Public defence
2011-05-27, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 14:00 (English)
Opponent
Supervisors
Note
At the time of the doctoral defense, the following publications were unpublished and had a status as follows: Paper 3: Submitted, Paper 4: Submitted.Available from: 2011-05-05 Created: 2011-04-18 Last updated: 2011-05-10Bibliographically approved

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