Using stable isotope tagging and mass spectrometry to characterize protein complexes and to detect changes in their composition.
One of the primary goals of proteomics is the description of the composition, dynamics, and connections of the multiprotein modules that catalyze a wide range of biological functions in cells. Mass spectrometry (MS) has proven to be an extremely powerful tool for characterizing the composition of purified complexes. However, because MS is not a quantitative technique, the usefulness of the data is limited. For example, without quantitative measurements, it is difficult to detect dynamic changes in complex composition, and it can be difficult to distinguish bona fide complex components from nonspecifically copurifying proteins. In this chapter, we describe a strategy for characterizing the composition of protein complexes and their dynamic changes in composition by combining affinity purification approaches with stable isotope tagging and MS. The use of software tools for statistical analysis of the data is also described.
Ranish
Jeffrey A
JA
Institute for Systems Biology, Seattle, WA, USA.
Brand
Marjorie
M
Aebersold
Ruedi
R
D000818:Animals; D045744:Cell Line, Tumor; D002846:Chromatography, Affinity; D007553:Isotope Labeling / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D046912:Multiprotein Complexes / Q000378:metabolism*; D010103:Oxygen Isotopes / Q000737:chemistry / Q000378:metabolism; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics; D012441:Saccharomyces cerevisiae / Q000378:metabolism
D046912:Multiprotein Complexes; D010103:Oxygen Isotopes; D011506:Proteins