title: Isolation of protein complexes involved in mitosis and cytokinesis from Drosophila cultured cells.
abstract: The identification of all the individual components that constitute the plethora of complexes in each cell type represents perhaps the most exciting challenge of postgenomic biology. This is particularly important in the study of events such as mitosis and cytokinesis, in which rapid and precise protein-protein interactions regulate both the direction and accuracy of these intricate processes. Here we describe an experimental strategy to isolate protein complexes involved in mitosis and cytokinesis in cultured Drosophila cells. This method involves the tagging of the bait protein with two IgG binding domains of Protein A and the isolation of the tagged bait along with its interacting partners by a single affinity purification step. These isolated complexes can then be analysed by several methods including mass spectrometry and Western blotting. Although this method has proven very successful in isolating mitotic and cytokinetic complexes, it can also be used to characterise protein complexes involved in many other cellular processes.
authors.affiliation: Cancer Research UK Cell Cycle Genetics Research Group, Department of Genetics, University of Cambridge, Cambridge, UK.
authors.forename: Pier Paolo
authors.initials: PP
authors.lastname: D'Avino
authors.forename: Vincent
authors.initials: V
authors.lastname: Archambault
authors.forename: Marcin R
authors.initials: MR
authors.lastname: Przewloka
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.forename: Ernest D
authors.initials: ED
authors.lastname: Laue
authors.forename: David M
authors.initials: DM
authors.lastname: Glover
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins / Q000302:isolation & purification*; D002478:Cells, Cultured; D004330:Drosophila / Q000737:chemistry* / Q000166:cytology; D029721:Drosophila Proteins / Q000302:isolation & purification*; D008938:Mitosis; D046912:Multiprotein Complexes / Q000302:isolation & purification
chemical_list: D018797:Cell Cycle Proteins; D029721:Drosophila Proteins; D046912:Multiprotein Complexes