Two-dimensional gel electrophoresis-based proteomics of mycobacteria.
Two-dimensional gel electrophoresis (2-DE) in combination with mass spectrometry (MS) is the classic proteomics approach used to monitor the dynamics of protein abundance and posttranslational modifications in biological systems. In this chapter, we provide detailed protocols for 2-DE-based proteomics of mycobacteria. Adequate standard operating procedures for mycobacterial culture, subcellular fractionation, and selective enrichment of proteins are indispensable prerequisites for targeted proteome analyses. Therefore, we also provide approved protocols for selective and efficient extraction of cytosolic, secreted, and hydrophobic plasma membrane proteins of mycobacteria, as well as for isolation of mycobacteria from infected macrophages.
Mattow
Jens
J
Department of Immunology, Max Planck Institute for Infection Biology, Charitéplatz 1, D-10117 Berlin, Germany. mattow@mpiib-berlin.mpg.de
Siejak
Frank
F
Hagens
Kristine
K
Kreuzeder
Julia
J
Kaufmann
Stefan H E
SH
Schaible
Ulrich E
UE
D000818:Animals; D001426:Bacterial Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D008264:Macrophages / Q000382:microbiology; D009161:Mycobacterium / Q000302:isolation & purification / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods
D001426:Bacterial Proteins