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 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.
Huang
Fang
F
Department of Biochemistry and Biophysics, Arrhenius Laboratories of Natural Sciences, Stockholm University, SE-10691 Stockholm, Sweden.
Hedman
Erik
E
Funk
Christiane
C
Kieselbach
Thomas
T
Schröder
Wolfgang P
WP
Norling
Birgitta
B
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis*; D001426:Bacterial Proteins / Q000032:analysis* / Q000302:isolation & purification; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D008969:Molecular Sequence Data; D020543:Proteome / Q000032:analysis*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D013347:Subcellular Fractions / Q000378:metabolism; D046939:Synechocystis / Q000737:chemistry*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D020543:Proteome