High throughput construction and small scale expression screening of multi-tag vectors in Escherichia coli

被引：68

作者：

Bird, Louise E. ^{[1
,2
]}

机构：

[1] Univ Oxford, Div Struct Biol, Oxford OX3 7BN, England

[2] Rutherford Appleton Lab, OPPF UK, Didcot OX11 0FA, Oxon, England

来源：

METHODS | 2011年 / 55卷 / 01期

基金：

英国生物技术与生命科学研究理事会; 英国惠康基金;

关键词：

Hexa-histidine tag; Solubilising protein; N-terminal fusion; Escherichia coli; Protein expression; InFusion cloning; SOLUBLE RECOMBINANT PROTEINS; MALTOSE-BINDING-PROTEIN; FUSION PARTNERS; SOLUBILITY; PURIFICATION; CLONING; CRYSTALLIZATION; AUTOINDUCTION; POLYPEPTIDES; TECHNOLOGY;

D O I：

10.1016/j.ymeth.2011.08.002

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

A suite of protein fusion vectors is presented that has been designed so that nine separate fusion vectors can be constructed from one PCR product using lnFusion (TM) cloning. These vectors in combination with a small scale Escherichia coli expression screen can be used to assess in parallel the effect of fusion tags on solubility. The vectors were tested with 20 target proteins and the results suggest that the vectors are useful both as a rescue strategy if the N-terminal hexa-histidine tagged construct does not express and also as part of a primary expression experiment. (C) 2011 Elsevier Inc. All rights reserved.

引用

页码：29 / 37

页数：9

共 42 条

[1] Implementation of semi-automated cloning and prokaryotic expression screening: the impact of SPINE [J].

Alzari, Pedro M. ;

Berglund, H. ;

Berrow, N. S. ;

Blagova, E. ;

Busso, D. ;

Cambillau, C. ;

Campanacci, V. ;

Christodoulou, E. ;

Eiler, S. ;

Fogg, M. J. ;

Folkers, G. ;

Geerlof, A. ;

Hart, D. ;

Haouz, A. ;

Herman, M. D. ;

Macieira, S. ;

Nordlund, P. ;

Perrakis, A. ;

Quevillon-Cheruel, S. ;

Tarandeau, F. ;

van Tilbeurgh, H. ;

Unger, T. ;

Luna-Vargas, M. P. A. ;

Velarde, M. ;

Willmanns, M. ;

Owens, Raymond J. .

ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY, 2006, 62 :1103-1113

[2] Expression, purification and crystallization of a lyssavirus matrix (M) protein [J].

Assenberg, Rene ;

Delmas, Olivier ;

Graham, Stephen C. ;

Verma, Anil ;

Berrow, Nick ;

Stuart, David I. ;

Owens, Raymond J. ;

Bourhy, Herve ;

Grimes, Jonathan M. .

ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS, 2008, 64 :258-262

[3] The Protein Data Bank [J].

Berman, HM ;

Westbrook, J ;

Feng, Z ;

Gilliland, G ;

Bhat, TN ;

Weissig, H ;

Shindyalov, IN ;

Bourne, PE .

NUCLEIC ACIDS RESEARCH, 2000, 28 (01) :235-242

[4] A versatile ligation-independent cloning method suitable for high-throughput expression screening applications [J].

Berrow, Nick S. ;

Alderton, David ;

Sainsbury, Sarah ;

Nettleship, Joanne ;

Assenberg, Rene ;

Rahman, Nahid ;

Stuart, David I. ;

Owens, Raymond J. .

NUCLEIC ACIDS RESEARCH, 2007, 35 (06)

[5] Recombinant protein expression and solubility screening in Escherichia coli:: a comparative study [J].

Berrow, Nick S. ;

Buessow, K. ;

Coutard, B. ;

Diprose, J. ;

Ekberg, M. ;

Folkers, G. E. ;

Levy, N. ;

Lieu, V. ;

Owens, R. J. ;

Peleg, Y. ;

Pinaglia, C. ;

Quevillon-Cheruel, S. ;

Salim, L. ;

Scheich, C. ;

Vincentelli, R. ;

Busso, Didier .

ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY, 2006, 62 :1218-1226

[6]

Berrow Nick S., 2009, V498, P75, DOI 10.1007/978-1-59745-196-3_5

[7] A family of E-coli expression vectors for laboratory scale and high throughput soluble protein production [J].

Cabrita, LD ;

Dai, WW ;

Bottomley, SP .

BMC BIOTECHNOLOGY, 2006, 6 (1)

[8]

Davis GD, 1999, BIOTECHNOL BIOENG, V65, P382, DOI 10.1002/(SICI)1097-0290(19991120)65:4<382::AID-BIT2>3.3.CO

[9]

2-9

[10] Bacteria co-transformed with recombinant proteins and chaperones cloned in independent plasmids are suitable for expression tuning [J].

de Marco, A ;

De Marco, V .

JOURNAL OF BIOTECHNOLOGY, 2004, 109 (1-2) :45-52

← 1 2 3 4 5 →