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Proteomic studies of the thylakoid membrane of Synechocystis sp. PCC 6803
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.
2005 (English)In: Proteomics, ISSN 1615-9853, Vol. 5, no 18, 4905-4916 p.Article in journal (Refereed) Published
Abstract [en]

Purified thylakoid membranes from the cyanobacterium Synechocystis sp. PCC 6803 were used for the first time in proteomic studies. The membranes were prepared by a combination of sucrose density centrifugation and aqueous polymer two-phase partitioning. In total, 76 different proteins were identified from 2- and 1-D gels by MALDI-TOF MS analysis. Twelve of the identified proteins have a predicted Sec/Tat signal peptide. Fourteen of the proteins were known, or predicted to be, integral membrane proteins. Among the proteins identified were subunits of the well-characterized thylakoid membrane constituents Photosystem I and II, ATP synthase, cytochrome b6f-complex, NADH dehydrogenase, and phycobilisome complex. In addition, novel thylakoid membrane proteins, both integral and peripheral were found, including enzymes involved in protein folding and pigment biosynthesis. The latter were the chlorophyll biosynthesis enzymes, light-dependent protochlorophyllide reductase and geranylgeranyl reductase as well as phytoene desaturase involved in carotenoid biosynthesis and a water-soluble carotenoid-binding protein. Interestingly, in view of the protein sorting mechanism in cyanobacteria, one of the two signal peptidases type I of Synechocystis was found in the thylakoid membrane, whereas the second one has been identified previously in the plasma membrane. Sixteen proteins are hypothetical proteins with unknown function.

Place, publisher, year, edition, pages
Weinheim: WILEY-VCH , 2005. Vol. 5, no 18, 4905-4916 p.
Keyword [en]
Gel electrophoresis, Mass spectrometry, Synechocystis 6803, Thylakoid membrane
Identifiers
URN: urn:nbn:se:su:diva-29229DOI: 10.1002/pmic.200500111OAI: oai:DiVA.org:su-29229DiVA: diva2:231747
Available from: 2009-08-17 Created: 2009-08-17 Last updated: 2009-09-04Bibliographically approved
In thesis
1. A model for membrane organization and protein sorting in the cyanobacterium Synechocystis sp. PCC 6803
Open this publication in new window or tab >>A model for membrane organization and protein sorting in the cyanobacterium Synechocystis sp. PCC 6803
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Cyanobacteria constitute a unique group of eubacteria, possessing outer and plasma membranes as well as internal thylakoid membranes, the site for both photosynthesis and respiration. The combination of sucrose density centrifugation and two-phase partitioning methods leads to purification of different membranes from the cyanobacterium Synechocystis sp. PCC 6803 (referred to as Synechocystis). The standard proteomics methods, based on 1D- and 2D-gel electrophoresis, followed by protein digestion and MALDI-TOF MS, led to identification of 76 thylakoid and 51 plasma membrane proteins. In order to increase the number of identified proteins a shotgun proteomics approach was employed. Proteins were digested in complex mixtures, followed by LC separation of obtained peptides coupled to MS/MS. This approach led to identification of 379 different thylakoid and plasma membrane proteins, of which 124 were integral membrane proteins. 

The complex membrane organization of Synechocystis requires a unique system for transport and sorting of proteins into extracytosolic cell compartments. Obtained gel-based and shotgun proteomics data as well as results of the multivariate sequence analysis of both soluble and integral membrane proteins allowed to suggest a new mechanism for membrane organization and protein sorting in Synechocystis. The plasma and thylakoid membranes are proposed to be dynamically connected and both soluble and integral membrane proteins are inserted into connection points.

The Synechocystis genome possesses two genes encoding leader peptidases. Proteomic studies revealed that Sll0716 is localized in the thylakoid membrane (LepT), whereas Slr1377 in the plasma membrane (LepP). The BN/SDS-PAGE of the total membranes from mutant LepT revealed that LepT is directly involved in the processing of several photosynthetic and lumenal subunits, required for the assembly and function of PSI and maintaining of the thylakoid membrane organization in Synechocystis.

 

 

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2009. 63 p.
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-29043 (URN)978-91-7155-917-3 (ISBN)
Public defence
2009-09-30, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 12 A, Stockholm, 10:00 (English)
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Note
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: In progress. Paper 5: In progress. Available from: 2009-09-08 Created: 2009-08-07 Last updated: 2011-05-03Bibliographically approved

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