Arabidopsis HY5 protein functions as a DNA-binding tag for purification and functional immobilization of proteins on agarose/DNA microplate

被引：60

作者：

Sawasaki, Tatsuya ^{[1
,2
]}

Kamura, Nami ^{[1
,2
,3
]}

Matsunaga, Satoko ^{[1
,2
]}

Saeki, Mihoro ^{[3
]}

Tsuchimochi, Masateru ^{[1
,2
]}

Morishita, Ryo ^{[3
]}

Endo, Yaeta ^{[1
,2
,4
]}

机构：

[1] Ehime Univ, Cell Free Sci & Technol Res Ctr, Matsuyama, Ehime 7908577, Japan

[2] Ehime Univ, Venture Business Lab, Matsuyama, Ehime 7908577, Japan

[3] CellFree Sci Co Ltd, Kanagawa 2300046, Japan

[4] RIKEN, Genom Sci Ctr, Yokohama, Kanagawa 2300045, Japan

来源：

FEBS LETTERS | 2008年 / 582卷 / 02期

关键词：

Arabidopsis HY5; DNA-binding tag; cell-free protein synthesis; protein microarray : substrate screening; transcription factor;

D O I：

10.1016/j.febslet.2007.12.004

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Protein microarray is considered to be one of the key analytical tools for high-throughput protein function analysis. Here, we report that the Arabidopsis HY5 functions as a novel DNA-binding tag (DBtag) for proteins. We also demonstrate that the DBtagged proteins could be immobilized and purified on a newly designed agarose/DNA microplate. Furthermore, we show three applications using the microarray: (1) detection of autophosphorylation activity of DBtagged human protein kinases and inhibition of their activity by staurosporine, (2) specific cleavage of DBtagged proteins by a virus protease and caspase 3, and (3) detection of a protein-protein interaction between the DBtagged UBE2N and UBE2v1. Thus, this method may facilitate rapid functional analysis of a wide range of proteins. (C) 2007 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

引用

页码：221 / 228

页数：8

共 31 条

[1] Current strategies for the use of affinity tags and tag removal for the purification of recombinant proteins
Arnau, Jos
Lauritzen, Conni
Petersen, Gitte E.
Pedersen, John
[J]. PROTEIN EXPRESSION AND PURIFICATION, 2006, 48 (01) : 1 - 13
[2] BRIZZARD BL, 1994, BIOTECHNIQUES, V16, P730
[3] Cell-free expression systems for eukaryotic protein production
Endo, Yaeta
Sawasaki, Tatsuya
[J]. CURRENT OPINION IN BIOTECHNOLOGY, 2006, 17 (04) : 373 - 380
[4] ISOLATION OF MONOCLONAL-ANTIBODIES SPECIFIC FOR HUMAN C-MYC PROTO-ONCOGENE PRODUCT
EVAN, GI
LEWIS, GK
RAMSAY, G
BISHOP, JM
[J]. MOLECULAR AND CELLULAR BIOLOGY, 1985, 5 (12) : 3610 - 3616
[5] Analogs of staurosporine: Potential anticancer drugs?
Gescher, A
[J]. GENERAL PHARMACOLOGY, 1998, 31 (05): : 721 - 728
[6] RAPID AND EFFICIENT PURIFICATION OF NATIVE HISTIDINE-TAGGED PROTEIN EXPRESSED BY RECOMBINANT VACCINIA VIRUS
JANKNECHT, R
DEMARTYNOFF, G
LOU, J
HIPSKIND, RA
NORDHEIM, A
STUNNENBERG, HG
[J]. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1991, 88 (20) : 8972 - 8976
[7] One-step purification of recombinant proteins using a nanomolar-affinity streptavidin-binding peptide, the SBP-tag
Keefe, AD
Wilson, DS
Seelig, B
Szostak, JW
[J]. PROTEIN EXPRESSION AND PURIFICATION, 2001, 23 (03) : 440 - 446
[8] KELLERMANN OK, 1982, METHOD ENZYMOL, V90, P459
[9] Activation of hPAK65 by caspase cleavage induces some of the morphological and biochemical changes of apoptosis
Lee, N
MacDonald, H
Reinhard, C
Halenbeck, R
Roulston, A
Shi, T
Williams, LT
[J]. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1997, 94 (25) : 13642 - 13647
[10] A highly efficient and robust cell-free protein synthesis system prepared from wheat embryos: Plants apparently contain a suicide system directed at ribosomes
Madin, K
Sawasaki, T
Ogasawara, T
Endo, Y
[J]. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2000, 97 (02) : 559 - 564

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