Analysis of Bacterial Membrane Proteins Produced During Mammalian Infection Using Hydrophobic Antigen Tissue Triton Extraction (HATTREX)

Timothy R. Crother1, Jarlath E. Nally2

1 Pediatrics Infectious Diseases, Cedars‐Sinai Medical Center, University of California, Los Angeles, California, 2 Veterinary Sciences Centre, School of Agriculture, Food Science and Veterinary Medicine, University College Dublin, Dublin, Ireland
Publication Name:  Current Protocols in Microbiology
Unit Number:  Unit 12.1
DOI:  10.1002/9780471729259.mc1201s9
Online Posting Date:  May, 2008
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Abstract

Hydrophobic antigen tissue Triton extraction (HATTREX) provides a method to extract and identify hydrophobic bacterial membrane proteins from host‐infected tissues. The non‐ionic detergent Triton X‐114 is used to solubilize host‐infected tissues at 4°C. Subsequent phase partitioning of Triton X‐114 extracted material at 37°C results in a “detergent poor” aqueous phase and a “detergent rich” detergent phase. While soluble proteins partition to the aqueous phase, hydrophobic proteins, such as proteins of the outer and inner membranes, can be found in the detergent phase. Characterization of the detergent phase sample provides insights into the proteome of bacterial membranes during infection. Curr. Protoc. Microbiol. 9:12.1.1‐12.1.5. © 2008 by John Wiley & Sons, Inc.

Keywords: Triton X‐114; in vivo; hydrophobic; membrane; antigen

     
 
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Table of Contents

  • Reagents and Solutions
  • Commentary
  • Literature Cited
     
 
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Materials

Basic Protocol 1:

  Materials
  • Tissue to be processed
  • Tissue suspension buffer (see recipe)
  • Triton X‐114 (protein grade, Calbiochem)
  • Trichloroacetic acid
  • Acetone
  • 1‐D SDS‐PAGE solubilization buffer (see recipe)
  • 2‐D SDS‐PAGE solubilization buffer (see recipe)
  • PowerGen 125 tissue grinder (Fisher Scientific) or similar device
  • Vortex
  • Shaking incubator
  • 37°C water bath
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Figures

Videos

Literature Cited

Literature Cited
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   Brusca, J.S. and Radolf, J.D. 1994. Isolation of integral membrane proteins by phase partitioning with Triton X‐114. Methods Enzymol. 228: 182‐193.
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