Identification of Effectors: Precipitation of Supernatant Material

Methods Mol Biol. 2017;1615:459-464. doi: 10.1007/978-1-4939-7033-9_31.

Abstract

Bacterial secretion systems allow the transport of proteins, called effectors, as well as external machine components in the extracellular medium or directly into target cells. Comparison of the secretome, i.e. the proteins released in the culture medium, of wild-type and mutant cells provides information on the secretion profile. In addition, mass spectrometry analyses of the culture supernatant of bacteria grown in liquid culture under secreting conditions allows the identification of secretion system substrates. Upon identification of the substrates, the secretion profile serves as a tool to test the functionality of secretion systems. Here we present a classical method used to concentrate the culture supernatant, based on trichloroacetic acid precipitation.

Keywords: Secretome; Supernatant; TCA precipitation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacteria / metabolism*
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / isolation & purification*
  • Bacterial Proteins / metabolism*
  • Bacterial Secretion Systems*
  • Blotting, Western
  • Mass Spectrometry

Substances

  • Bacterial Proteins
  • Bacterial Secretion Systems