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Identification and bioinformatic analysis of the membrane proteins of synechocystis sp. PCC 6803.

Wang Y, Xu W, Chitnis PR - Proteome Sci (2009)

Bottom Line: Bioinformatic analysis revealed that the identified proteins can be functionally classified into 14 distinct groups according to the cellular functions annotated by Cyanobase, including the two largest groups hypothetical and unknown, and photosynthesis and respiration.Homology analysis indicates that the identified membrane proteins are more conserved than the rest of the proteome.The proteins identified in this study combined with other published proteomic data provide the most comprehensive Synechocystis proteome catalog, which will serve as a useful reference for further detailed studies to address protein functions through both traditional gene-by-gene and systems biology approaches.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, 50011, USA. wychun@ucsd.edu

ABSTRACT

Background: The membranes of Synechocystis sp. PCC 6803 play a central role in photosynthesis, respiration and other important metabolic pathways. Comprehensive identification of the membrane proteins is of importance for a better understanding of the diverse functions of its unique membrane structures. Up to date, approximately 900 known or predicted membrane proteins, consisting 24.5% of Synechocystis sp. PCC 6803 proteome, have been indentified by large-scale proteomic studies.

Results: To resolve more membrane proteins on 2-D gels for mass spectrometry identification, we separated integral proteins from membrane associated proteins and collected them as the integral and peripheral fractions, respectively. In total, 95 proteins in the peripheral fraction and 29 proteins in the integral fraction were identified, including the 5 unique proteins that were not identified by any previous studies. Bioinformatic analysis revealed that the identified proteins can be functionally classified into 14 distinct groups according to the cellular functions annotated by Cyanobase, including the two largest groups hypothetical and unknown, and photosynthesis and respiration. Homology analysis indicates that the identified membrane proteins are more conserved than the rest of the proteome.

Conclusion: The proteins identified in this study combined with other published proteomic data provide the most comprehensive Synechocystis proteome catalog, which will serve as a useful reference for further detailed studies to address protein functions through both traditional gene-by-gene and systems biology approaches.

No MeSH data available.


Related in: MedlinePlus

Separation of integral membrane proteins by 2-DE. The integral proteins were separated using 18-cm IPG strips covering pH ranges 3–10 (nonlinear), and 4–7 for the first dimension, and then separated by 12–18% gradient SDS-PAGE gel in the second dimension. Each gel shown is a representative of the three replicates that has the best resolution. All the labeled protein spots in the 2-D gel were analyzed by mass spectrometry and identified using peptide mass fingerprinting.
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Figure 3: Separation of integral membrane proteins by 2-DE. The integral proteins were separated using 18-cm IPG strips covering pH ranges 3–10 (nonlinear), and 4–7 for the first dimension, and then separated by 12–18% gradient SDS-PAGE gel in the second dimension. Each gel shown is a representative of the three replicates that has the best resolution. All the labeled protein spots in the 2-D gel were analyzed by mass spectrometry and identified using peptide mass fingerprinting.

Mentions: To further enrich low abundant membrane proteins for the identification, we separated proteins on 2-D gels with multiple pH ranges. For the peripheral fraction, we used four 2-D gels covering pH ranges 3–10, 4–7, 4–5 and 5–6 (Figure 2). For the integral fraction, we used only two 2-D gels with pH range 3–10 and 4–7 (Figure 3) because the sample complexity in this fraction was relatively low compared with the peripheral fraction (Figure 1). Each gel had three replicates and only the protein spots presented in all replicates were chosen for PMF identification. In total, more than 600 protein spots were observed on 2-D gels for the peripheral fraction, and more than 200 protein spots were observed on 2-D gels for the integral fraction.


Identification and bioinformatic analysis of the membrane proteins of synechocystis sp. PCC 6803.

Wang Y, Xu W, Chitnis PR - Proteome Sci (2009)

Separation of integral membrane proteins by 2-DE. The integral proteins were separated using 18-cm IPG strips covering pH ranges 3–10 (nonlinear), and 4–7 for the first dimension, and then separated by 12–18% gradient SDS-PAGE gel in the second dimension. Each gel shown is a representative of the three replicates that has the best resolution. All the labeled protein spots in the 2-D gel were analyzed by mass spectrometry and identified using peptide mass fingerprinting.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2666656&req=5

Figure 3: Separation of integral membrane proteins by 2-DE. The integral proteins were separated using 18-cm IPG strips covering pH ranges 3–10 (nonlinear), and 4–7 for the first dimension, and then separated by 12–18% gradient SDS-PAGE gel in the second dimension. Each gel shown is a representative of the three replicates that has the best resolution. All the labeled protein spots in the 2-D gel were analyzed by mass spectrometry and identified using peptide mass fingerprinting.
Mentions: To further enrich low abundant membrane proteins for the identification, we separated proteins on 2-D gels with multiple pH ranges. For the peripheral fraction, we used four 2-D gels covering pH ranges 3–10, 4–7, 4–5 and 5–6 (Figure 2). For the integral fraction, we used only two 2-D gels with pH range 3–10 and 4–7 (Figure 3) because the sample complexity in this fraction was relatively low compared with the peripheral fraction (Figure 1). Each gel had three replicates and only the protein spots presented in all replicates were chosen for PMF identification. In total, more than 600 protein spots were observed on 2-D gels for the peripheral fraction, and more than 200 protein spots were observed on 2-D gels for the integral fraction.

Bottom Line: Bioinformatic analysis revealed that the identified proteins can be functionally classified into 14 distinct groups according to the cellular functions annotated by Cyanobase, including the two largest groups hypothetical and unknown, and photosynthesis and respiration.Homology analysis indicates that the identified membrane proteins are more conserved than the rest of the proteome.The proteins identified in this study combined with other published proteomic data provide the most comprehensive Synechocystis proteome catalog, which will serve as a useful reference for further detailed studies to address protein functions through both traditional gene-by-gene and systems biology approaches.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, 50011, USA. wychun@ucsd.edu

ABSTRACT

Background: The membranes of Synechocystis sp. PCC 6803 play a central role in photosynthesis, respiration and other important metabolic pathways. Comprehensive identification of the membrane proteins is of importance for a better understanding of the diverse functions of its unique membrane structures. Up to date, approximately 900 known or predicted membrane proteins, consisting 24.5% of Synechocystis sp. PCC 6803 proteome, have been indentified by large-scale proteomic studies.

Results: To resolve more membrane proteins on 2-D gels for mass spectrometry identification, we separated integral proteins from membrane associated proteins and collected them as the integral and peripheral fractions, respectively. In total, 95 proteins in the peripheral fraction and 29 proteins in the integral fraction were identified, including the 5 unique proteins that were not identified by any previous studies. Bioinformatic analysis revealed that the identified proteins can be functionally classified into 14 distinct groups according to the cellular functions annotated by Cyanobase, including the two largest groups hypothetical and unknown, and photosynthesis and respiration. Homology analysis indicates that the identified membrane proteins are more conserved than the rest of the proteome.

Conclusion: The proteins identified in this study combined with other published proteomic data provide the most comprehensive Synechocystis proteome catalog, which will serve as a useful reference for further detailed studies to address protein functions through both traditional gene-by-gene and systems biology approaches.

No MeSH data available.


Related in: MedlinePlus