Membrane proteome analysis of the green-sulfur bacterium Chlorobium tepidum.
An extensive proteomic approach relies on the possibility to visualize and analyze various types of proteins, including membrane proteins, which are rarely detectable on two-dimensional electrophoresis gels. In this study, different methods were employed for the enrichment of membrane proteins from Chlorobium tepidum prior to analysis with two-dimensional electrophoresis (2-DE). Isolated membranes were solubilized with Triton X-100 and from the supernatant we identified 58 unique proteins. The use of ionic sodium dodecyl sulfate (SDS) for protein solubilization, combined with acetone precipitation, resulted in an improved 2-DE pattern and the total number of the identified proteins was increased to 117. The use of acetone for protein precipitation improved the results by extracting compounds potentially deleterious to the resolution of 2-DE. However, the additional proteins detected by the use of SDS are in the majority more difficult to solubilize than less hydrophobic proteins. Further our attempts for selective extraction of the outer membrane proteins using the acid glycine method allowed the identification of 37 proteins of which 14 were predicted to have a signal sequence indicating their localization in the periplasmic space or in the outer membrane.
Aivaliotis
Michalis
M
Division of Biochemistry, Department of Chemistry, University of Crete, Heraklion, Greece.
Corvey
Carsten
C
Tsirogianni
Irene
I
Karas
Michael
M
Tsiotis
Georgios
G
D000096:Acetone / Q000737:chemistry; D001426:Bacterial Proteins / Q000032:analysis*; D011232:Chemical Precipitation; D041883:Chlorobium / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D008565:Membrane Proteins / Q000032:analysis*; D020543:Proteome / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry
D001426:Bacterial Proteins; D008565:Membrane Proteins; D020543:Proteome; D000096:Acetone; D012967:Sodium Dodecyl Sulfate