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Isolation of protein complexes involved in mitosis and cytokinesis from Drosophila cultured cells

Isolation of protein complexes involved in mitosis and cytokinesis from Drosophila cultured cells
Isolation of protein complexes involved in mitosis and cytokinesis from Drosophila cultured cells
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.
99-112
Humana Press
D'Avino, Pier Paolo
0e659e0c-c3a7-4344-8850-3364810b9445
Archambault, Vincent
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Przewloka, Marcin
9b25e73c-ec15-43df-a5a4-ac9574bb20ab
Zhang, Wei
fa6d7061-bfce-48c0-879c-925302213c1a
Laue, Ernest D.
9bfea318-d6a1-493a-9055-41a9f9e06f17
Glover, David M.
cca9cd19-3e1e-4906-b418-41876c1f9c61
McAinsh, Andrew D.
D'Avino, Pier Paolo
0e659e0c-c3a7-4344-8850-3364810b9445
Archambault, Vincent
3654cc6d-d3f9-4404-8ae5-9d8af8b008dc
Przewloka, Marcin
9b25e73c-ec15-43df-a5a4-ac9574bb20ab
Zhang, Wei
fa6d7061-bfce-48c0-879c-925302213c1a
Laue, Ernest D.
9bfea318-d6a1-493a-9055-41a9f9e06f17
Glover, David M.
cca9cd19-3e1e-4906-b418-41876c1f9c61
McAinsh, Andrew D.

D'Avino, Pier Paolo, Archambault, Vincent, Przewloka, Marcin, Zhang, Wei, Laue, Ernest D. and Glover, David M. (2009) Isolation of protein complexes involved in mitosis and cytokinesis from Drosophila cultured cells. In, McAinsh, Andrew D. (ed.) Mitosis. (Methods in Molecular Biology, 545) New York, US. Humana Press, pp. 99-112. (doi:10.1007/978-1-60327-993-2_6).

Record type: Book Section

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.

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More information

Published date: 2009
Organisations: Molecular and Cellular, Centre for Biological Sciences

Identifiers

Local EPrints ID: 393865
URI: http://eprints.soton.ac.uk/id/eprint/393865
PURE UUID: 4118cf2f-45b5-4ebe-b2c5-60c658dec62d
ORCID for Marcin Przewloka: ORCID iD orcid.org/0000-0002-0329-9162

Catalogue record

Date deposited: 13 May 2016 15:50
Last modified: 15 Mar 2024 03:54

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Contributors

Author: Pier Paolo D'Avino
Author: Vincent Archambault
Author: Wei Zhang
Author: Ernest D. Laue
Author: David M. Glover
Editor: Andrew D. McAinsh

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