Tandem affinity purification of functional TAP-tagged proteins from human cells

被引：44

作者：

Gregan, Juraj ^{[1
,2
]}

Riedel, Christian G. ^{[2
]}

Petronczki, Mark ^{[2
]}

Cipak, Lubos ^{[1
]}

Rumpf, Cornelia ^{[1
]}

Poser, Ina ^{[3
]}

Buchholz, Frank ^{[3
]}

Mechtler, Karl ^{[2
]}

Nasmyth, Kim ^{[2
,4
]}

机构：

[1] Univ Vienna, Dept Chrosome Biol, Max F Perutz Labs, A-1030 Vienna, Austria

[2] Res Inst Mol Pathol, A-1030 Vienna, Austria

[3] Max Planck Inst Mol Cell Biol & Genet, D-01307 Dresden, Germany

[4] Univ Oxford, Dept Biochem, Oxford OX1 3QU, England

来源：

NATURE PROTOCOLS | 2007年 / 2卷 / 05期

基金：

奥地利科学基金会;

关键词：

D O I：

10.1038/nprot.2007.172

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Tandem affinity purification ( TAP) is a generic two-step affinity purification protocol for isolation of TAP-tagged proteins together with associated proteins. We used bacterial artificial chromosome to heterologously express TAP-tagged murine Sgo1 protein in human HeLa cells. This allowed us to test the functionality of the Sgo1-TAP protein by RNA interference-mediated depletion of the endogenous human Sgo1. Here, we present an optimized protocol for purification of TAP-tagged Sgo1 protein as well as KIAA1387 from HeLa cells with detailed instructions. The purification protocol can be completed in 1 day and it should be applicable to other proteins.

引用

页码：1145 / 1151

页数：7

共 23 条

[1] A physical and functional map of the human TNF-α NF-κB signal transduction pathway [J].

Bouwmeester, T ;

Bauch, A ;

Ruffner, H ;

Angrand, PO ;

Bergamini, G ;

Croughton, K ;

Cruciat, C ;

Eberhard, D ;

Gagneur, J ;

Ghidelli, S ;

Hopf, C ;

Huhse, B ;

Mangano, R ;

Michon, AM ;

Schirle, M ;

Schlegl, J ;

Schwab, M ;

Stein, MA ;

Bauer, A ;

Casari, G ;

Drewes, G ;

Gavin, AC ;

Jackson, DB ;

Joberty, G ;

Neubauer, G ;

Rick, J ;

Kuster, B ;

Superti-Furga, G .

NATURE CELL BIOLOGY, 2004, 6 (02) :97-+

[2] An efficient tandem affinity purification procedure for interaction proteomics in mammalian cells [J].

Buerckstuemmer, Tilmann ;

Bennett, Keiryn L. ;

Preradovic, Adrijana ;

Schutze, Gregor ;

Hantschel, Oliver ;

Superti-Furga, Giulio ;

Bauch, Angela .

NATURE METHODS, 2006, 3 (12) :1013-1019

[3] Interaction network containing conserved and essential protein complexes in Escherichia coli [J].

Butland, G ;

Peregrín-Alvarez, JM ;

Li, J ;

Yang, WH ;

Yang, XC ;

Canadien, V ;

Starostine, A ;

Richards, D ;

Beattie, B ;

Krogan, N ;

Davey, M ;

Parkinson, J ;

Greenblatt, J ;

Emili, A .

NATURE, 2005, 433 (7025) :531-537

[4]

CHEESEMAN IM, 2005, SCI STKE, P1

[5] AU binding proteins recruit the exosome to degrade ARE-containing mRNAs [J].

Chen, CY ;

Gherzi, R ;

Ong, SE ;

Chan, EKL ;

Raijmakers, R ;

Pruijn, GJM ;

Stoecklin, G ;

Moroni, C ;

Mann, M ;

Karin, M .

CELL, 2001, 107 (04) :451-464

[6] An efficient protein complex purification method for functional proteomics in higher eukaryotes [J].

Forler, D ;

Köcher, T ;

Rode, M ;

Gentzel, M ;

Izaurralde, E ;

Wilm, M .

NATURE BIOTECHNOLOGY, 2003, 21 (01) :89-92

[7] Proteome survey reveals modularity of the yeast cell machinery [J].

Gavin, AC ;

Aloy, P ;

Grandi, P ;

Krause, R ;

Boesche, M ;

Marzioch, M ;

Rau, C ;

Jensen, LJ ;

Bastuck, S ;

Dümpelfeld, B ;

Edelmann, A ;

Heurtier, MA ;

Hoffman, V ;

Hoefert, C ;

Klein, K ;

Hudak, M ;

Michon, AM ;

Schelder, M ;

Schirle, M ;

Remor, M ;

Rudi, T ;

Hooper, S ;

Bauer, A ;

Bouwmeester, T ;

Casari, G ;

Drewes, G ;

Neubauer, G ;

Rick, JM ;

Kuster, B ;

Bork, P ;

Russell, RB ;

Superti-Furga, G .

NATURE, 2006, 440 (7084) :631-636

[8] Functional organization of the yeast proteome by systematic analysis of protein complexes [J].

Gavin, AC ;

Bösche, M ;

Krause, R ;

Grandi, P ;

Marzioch, M ;

Bauer, A ;

Schultz, J ;

Rick, JM ;

Michon, AM ;

Cruciat, CM ;

Remor, M ;

Höfert, C ;

Schelder, M ;

Brajenovic, M ;

Ruffner, H ;

Merino, A ;

Klein, K ;

Hudak, M ;

Dickson, D ;

Rudi, T ;

Gnau, V ;

Bauch, A ;

Bastuck, S ;

Huhse, B ;

Leutwein, C ;

Heurtier, MA ;

Copley, RR ;

Edelmann, A ;

Querfurth, E ;

Rybin, V ;

Drewes, G ;

Raida, M ;

Bouwmeester, T ;

Bork, P ;

Seraphin, B ;

Kuster, B ;

Neubauer, G ;

Superti-Furga, G .

NATURE, 2002, 415 (6868) :141-147

[9] Tandem affinity purification and identification of protein complex components [J].

Gould, KL ;

Ren, LP ;

Feoktistova, AS ;

Jennings, JL ;

Link, AJ .

METHODS, 2004, 33 (03) :239-244

[10] New partners of acyl carrier protein detected in Escherichia coli by tandem affinity purification [J].

Gully, D ;

Moinier, D ;

Loiseau, L ;

Bouveret, E .

FEBS LETTERS, 2003, 548 (1-3) :90-96

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