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.
Department of Biochemistry and Biophysics, Arrhenius Laboratories of Natural Sciences, Stockholm University, SE-10691 Stockholm, Sweden.
Fang
F
Huang
Erik
E
Hedman
Christiane
C
Funk
Thomas
T
Kieselbach
Wolfgang P
WP
Schröder
Birgitta
B
Norling
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