Mapping the Membrane Proteome of Corynebacterium Glutamicum

Proteomics. 2005 Apr;5(5):1317-30. doi: 10.1002/pmic.200400993.

Abstract

In order to avoid the specific problems with intrinsic membrane proteins in proteome analysis, a new procedure was developed which is superior to the classical two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) method in terms of intrinsic membrane proteins. For analysis of the membrane proteome from Corynebacterium glutamicum, we replaced the first separation dimension, i.e., the isoelectric focusing step, by anion-exchange chromatography, followed by sodium dodecyl sulfate (SDS)-PAGE in the second separation dimension. Enrichment of the membrane intrinsic subproteome was achieved by washing with 2.5 M NaBr which removed more than 35% of the membrane-associated soluble proteins. For the extraction and solubilization of membrane proteins, the detergent amidosulfobetaine 14 (ASB-14) was most efficient in a detailed screening procedure and proved also suitable for chromatography. 356 gel bands were spotted, and out of 170 different identified proteins, 50 were membrane-integral. Membrane proteins with one up to 13 transmembrane helices were found. Careful analysis revealed that this new procedure covers proteins from a wide pI range (3.7-10.6) and a wide mass range of 10-120 kDa. About 50% of the identified membrane proteins belong to various functional categories like energy metabolism, transport, signal transduction, protein translocation, and proteolysis while for the others a function is not yet known, indicating the potential of the developed method for elucidation of membrane proteomes in general.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / analysis*
  • Cell Fractionation
  • Cell Membrane / chemistry*
  • Chromatography, Ion Exchange
  • Corynebacterium glutamicum / chemistry*
  • Detergents / chemistry
  • Electrophoresis, Gel, Two-Dimensional / methods
  • Membrane Proteins / analysis*
  • Proteome / analysis*
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Substances

  • Bacterial Proteins
  • Detergents
  • Membrane Proteins
  • Proteome