Abstract
We present a novel direct protocol for deep proteome characterization of microorganisms in soil. The
method employs thermally assisted detergent-based cellular lysis (SDS) of soil samples, followed by
TCA precipitation for proteome extraction/cleanup prior to liquid chromatography-mass spectrometric
characterization. This approach was developed and optimized using different soils inoculated with
genome-sequenced bacteria (Gram-negative Pseudomonas putida or Gram-positive Arthrobacter
chlorophenolicus). Direct soil protein extraction was compared to protein extraction from cells isolated
from the soil matrix prior to lysis (indirect method). Each approach resulted in identification of greater
than 500 unique proteins, with a wide range in molecular mass and functional categories. To our
knowledge, this SDS-TCA approach enables the deepest proteome characterizations of microbes in
soil to date, without significant biases in protein size, localization, or functional category compared to
pure cultures. This protocol should provide a powerful tool for ecological studies of soil microbial
communities.
