Investigative proteomics: identification of an unknown plant virus from infected plants using mass spectrometry

J Am Soc Mass Spectrom. 2003 Jul;14(7):736-41. doi: 10.1016/S1044-0305(03)00125-9.

Abstract

We describe the identification of a previously uncharacterized plant virus that is capable of infecting Nicotiana spp. and Arabidopsis thaliana. Protein extracts were first prepared from leaf tissue of uninfected tobacco plants, and the proteins were visualized with two-dimensional electrophoresis (2-DE). Matching gels were then run using protein extracts of a tobacco plant infected with tobacco mosaic virus (TMV). After visual comparison, the proteins spots that were differentially expressed in infected plant tissues were cut from the gels and analyzed by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Tandem mass spectrometry data of individual peptides was searched with SEQUEST. Using this approach we demonstrated a successful proof-of-concept experiment by identifying TMV proteins present in the total protein extract. The same procedure was then applied to tobacco plants infected with a laboratory viral isolate of unknown identity. Several of the differentially expressed protein spots were identified as proteins of potato virus X (PVX), thus successfully identifying the causative agent of the uncharacterized viral infection. We believe this demonstrates that HPLC-MS/MS can be used to successfully characterize unknown viruses in infected plants.

MeSH terms

  • Amino Acid Sequence
  • Electrophoresis, Gel, Two-Dimensional
  • Mass Spectrometry
  • Molecular Sequence Data
  • Plant Leaves / chemistry
  • Plant Leaves / virology
  • Plant Viruses / chemistry*
  • Plant Viruses / classification*
  • Plant Viruses / isolation & purification
  • Potexvirus / chemistry
  • Potexvirus / classification
  • Potexvirus / isolation & purification
  • Proteome / analysis*
  • Proteome / chemistry
  • Proteomics / methods*
  • Tobacco / virology*
  • Viral Proteins / analysis*
  • Viral Proteins / chemistry

Substances

  • Proteome
  • Viral Proteins