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Automated nanoflow liquid chromatography/tandem mass spectrometric identification of proteins from Shewanella putrefaciens separated by two-dimensional polyacrylamide gel electrophoresis
Devreese, B.; Vanrobaeys, F.; Van Beeumen, J. (2001). Automated nanoflow liquid chromatography/tandem mass spectrometric identification of proteins from Shewanella putrefaciens separated by two-dimensional polyacrylamide gel electrophoresis. Rapid Comm. Mass Spectrom. 15(1): 50-56. dx.doi.org/10.1002/1097-0231(20010115)15:1<50::AID-RCM191>3.0.CO;2-V
In: Rapid Communications in Mass Spectrometry. Wiley: London. ISSN 0951-4198, more
Peer reviewed article  

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Keyword
    Marine

Authors  Top 
  • Devreese, B.
  • Vanrobaeys, F.
  • Van Beeumen, J.

Abstract
    The implementation of nanoflow liquid chromatography offers unique opportunities for automation of proteomics research. We demonstrate that automated nanoflow LC/MS/MS allowed the unambiguous identification of proteins from the omnipotent bacterium Shewanella putrefaciens, based on similarity searches against the completely determined genome of related microorganisms and against non-redundant databases. Total protein extracts were separated by 2-dimensional polyacrylamide electrophoresis. Only 1/20th of a tryptic digest mixture obtained from a single Coomassie Blue stained spot was loaded on the nanoflow LC column using a preconcentration/desalting step, and analyzed on-line on a hybrid quadrupole time-of-flight mass spectrometer with an automated MS-to-MS/MS switching protocol. This method allowed the de novo peptide sequence determination of several tryptic fragments and the identification of different proteins.

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