Isolation of Outer Membrane of Synechocystis sp. PCC 6803 and Its Proteomic Characterization

In this report, we describe a newly developed method for isolating outer membranes from Synechocystis sp. PCC 6803 cells. The purity of the outer membrane fraction was verified by immunoblot analysis using antibodies against membrane-specific marker proteins. We investigated the protein composition...

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Veröffentlicht in:Molecular & cellular proteomics 2004-06, Vol.3 (6), p.586-595
Hauptverfasser: Huang, Fang, Hedman, Erik, Funk, Christiane, Kieselbach, Thomas, Schröder, Wolfgang P, Norling, Birgitta
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Sprache:eng
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Zusammenfassung:In this report, we describe a newly developed method for isolating outer membranes from Synechocystis sp. PCC 6803 cells. The purity of the outer membrane fraction was verified by immunoblot analysis using antibodies against membrane-specific marker proteins. We investigated the protein composition of the outer membrane using two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry followed by database identification. Forty-nine proteins were identified corresponding to 29 different gene products. All of the identified proteins have a putative N-terminal signal peptide. About 40% of the proteins identified represent hypothetical proteins with unknown function. Among the proteins identified are a Toc75 homologue, a protein that was initially found in the outer envelope of chloroplasts in pea, as well as TolC, putative porins, and a pilus protein. Other proteins identified include ABC transporters and GumB, which has a suggested function in carbohydrate export. A number of proteases such as HtrA were also found in the outer membrane of Synechocystis sp. PCC 6803.
ISSN:1535-9476
1535-9484
1535-9484
DOI:10.1074/mcp.M300137-MCP200