Isolation and mass spectrometry of transcription factor complexes

被引:68
作者
Winkler, GS
Lacomis, L
Philip, J
Erdjument-Bromage, H
Svejstrup, JQ
Tempst, P
机构
[1] Mem Sloan Kettering Canc Ctr, Program Mol Biol, New York, NY 10021 USA
[2] Imperial Canc Res Fund, Clare Hall Labs, Canc Res UK London Res Inst, S Mimms EN6 3LD, Herts, England
关键词
D O I
10.1016/S1046-2023(02)00030-0
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Protocols are described that enable the isolation of novel proteins associated with a known protein and the subsequent identification of these proteins by mass spectrometry. We review the basics of nanosample handling and of two complementary approaches to mass analysis, and provide protocols for the entire process. The protein isolation procedure is rapid and based on two high-affinity chromatography steps. The method does not require previous knowledge of complex composition or activity and permits subsequent biochemical characterization of the isolated factor. As an example, we provide the procedures used to isolate and analyze yeast Elongator, a histone acetyltransferase complex important for transcript elongation, which led to the identification of three novel subunits. (C) 2002 Elsevier Science (USA). All rights reserved.
引用
收藏
页码:260 / 269
页数:10
相关论文
共 34 条
[1]   RSC, an essential, abundant chromatin-remodeling complex [J].
Cairns, BR ;
Lorch, Y ;
Li, Y ;
Zhang, MC ;
Lacomis, L ;
ErdjumentBromage, H ;
Tempst, P ;
Du, J ;
Laurent, B ;
Kornberg, RD .
CELL, 1996, 87 (07) :1249-1260
[2]   A protein complex containing Tho2, Hpr1, Mft1 and a novel protein, Thp2, connects transcription elongation with mitotic recombination in Saccharomyces cerevisiae [J].
Chávez, S ;
Beilharz, T ;
Rondón, AG ;
Erdjument-Bromage, H ;
Tempst, P ;
Svejstrup, JQ ;
Lithgow, T ;
Aguilera, A .
EMBO JOURNAL, 2000, 19 (21) :5824-5834
[3]   AN APPROACH TO CORRELATE TANDEM MASS-SPECTRAL DATA OF PEPTIDES WITH AMINO-ACID-SEQUENCES IN A PROTEIN DATABASE [J].
ENG, JK ;
MCCORMACK, AL ;
YATES, JR .
JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY, 1994, 5 (11) :976-989
[4]   Examination of micro-tip reversed-phase liquid chromatographic extraction of peptide pools for mass spectrometric analysis [J].
Erdjument-Bromage, H ;
Lui, M ;
Lacomis, L ;
Grewal, A ;
Annan, RS ;
McNulty, DE ;
Carr, SA ;
Tempst, P .
JOURNAL OF CHROMATOGRAPHY A, 1998, 826 (02) :167-181
[5]   HIGH-SENSITIVITY SEQUENCING OF LARGE PROTEINS - PARTIAL STRUCTURE OF THE RAPAMYCIN-FKBP12 TARGET [J].
ERDJUMENTBROMAGE, H ;
LUI, M ;
SABATINI, DM ;
SNYDER, SH ;
TEMPST, P .
PROTEIN SCIENCE, 1994, 3 (12) :2435-2446
[6]   The Elp2 subunit of elongator and elongating RNA polymerase II holoenzyme is a WD40 repeat protein [J].
Fellows, J ;
Erdjument-Bromage, H ;
Tempst, P ;
Svejstrup, JQ .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2000, 275 (17) :12896-12899
[7]   PURIFICATION OF A RAS-RESPONSIVE ADENYLYL CYCLASE COMPLEX FROM SACCHAROMYCES-CEREVISIAE BY USE OF AN EPITOPE ADDITION METHOD [J].
FIELD, J ;
NIKAWA, J ;
BROEK, D ;
MACDONALD, B ;
RODGERS, L ;
WILSON, IA ;
LERNER, RA ;
WIGLER, M .
MOLECULAR AND CELLULAR BIOLOGY, 1988, 8 (05) :2159-2165
[8]   Tuning of an electrospray ionization source for maximum peptide-ion transmission into a mass spectrometer [J].
Geromanos, S ;
Freckleton, G ;
Tempst, P .
ANALYTICAL CHEMISTRY, 2000, 72 (04) :777-790
[9]  
Gharahdaghi F, 1999, ELECTROPHORESIS, V20, P601, DOI 10.1002/(SICI)1522-2683(19990301)20:3<601::AID-ELPS601>3.0.CO
[10]  
2-6