Proteomic study of the Arabidopsis thaliana chloroplastic envelope membrane utilizing alternatives to traditional two-dimensional electrophoresis

J Proteome Res. Jul-Aug 2003;2(4):413-25. doi: 10.1021/pr034025j.

Abstract

With the completion of the sequencing of the Arabidopsis genome and with the significant increase in the amount of other plant genome and expressed sequence tags (ESTs) data, plant proteomics is rapidly becoming a very active field. We have pursued a high-throughput mass spectrometry-based proteomics approach to identify and characterize membrane proteins localized to the Arabidopsis thaliana chloroplastic envelope membrane. In this study, chloroplasts were prepared from plate- or soil-grown Arabidopsis plants using a novel isolation procedure, and "mixed" envelopes were subsequently isolated using sucrose step gradients. We applied two alternative methodologies, off-line multidimensional protein identification technology (Off-line MUDPIT) and one-dimensional (1D) gel electrophoresis followed by proteolytic digestion and liquid chromatography coupled with tandem mass spectrometry (Gel-C-MS/MS), to identify envelope membrane proteins. This proteomic study enabled us to identify 392 nonredundant proteins.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis / chemistry*
  • Arabidopsis / genetics
  • Arabidopsis / physiology
  • Arabidopsis Proteins / analysis*
  • Arabidopsis Proteins / classification
  • Arabidopsis Proteins / isolation & purification
  • Blotting, Western
  • Chloroplasts / chemistry*
  • Chloroplasts / physiology
  • Computational Biology
  • Databases, Protein
  • Electronic Data Processing
  • Electrophoresis, Polyacrylamide Gel
  • Gas Chromatography-Mass Spectrometry
  • Membrane Proteins / analysis*
  • Membrane Proteins / classification
  • Membrane Proteins / isolation & purification
  • Molecular Sequence Data
  • Proteomics*

Substances

  • Arabidopsis Proteins
  • Membrane Proteins