Efficient suppression of the amber codon in E-coli in vitro translation system

被引：26

作者：

Agafonov, DE ^{[1
]}

Huang, YW ^{[1
]}

Grote, M ^{[1
]}

Sprinzl, M ^{[1
]}

机构：

[1] Univ Bayreuth, Biochem Lab, D-95440 Bayreuth, Germany

来源：

FEBS LETTERS | 2005年 / 579卷 / 10期

关键词：

in vitro translation; suppression; tRNA; release factor 1; esterase; termination;

D O I：

10.1016/j.febslet.2005.03.004

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：

An mRNA encoding the esterase from Alicyclobacillus acidocaldarius with catalytically essential serine codon (ACG) replaced by an amber (UAG) codon was used to study the suppression in in vitro translation system. Suppression of UAG by tRNA (Ser(CUA)) was monitored by determination of the full-length and active esterase. It was shown that commonly used increase of suppressor tRNA concentration inhibits protein production and therefore limits suppression. In situ deactivation of release factor by specific antibodies leads to efficient suppression already at low suppressor tRNA concentration and allows an in vitro synthesis of fully active enzyme in high yield undistinguishable from wild-type protein. (c) 2005 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

引用

页码：2156 / 2160

页数：5

共 22 条

[1] THE CONCENTRATION OF POLYPEPTIDE-CHAIN RELEASE FACTOR-1 AND FACTOR-2 AT DIFFERENT GROWTH-RATES OF ESCHERICHIA-COLI [J].

ADAMSKI, FM ;

MCCAUGHAN, KK ;

JORGENSEN, F ;

KURLAND, CG ;

TATE, WP .

JOURNAL OF MOLECULAR BIOLOGY, 1994, 238 (03) :302-308

[2] Suppression of eukaryotic translation termination by selected RNAs [J].

Carnes, J ;

Frolova, L ;

Zinnen, S ;

Drugeon, G ;

Phillippe, M ;

Justesen, J ;

Haenni, AL ;

Leinwand, L ;

Kisselev, LL ;

Yarus, M .

RNA, 2000, 6 (10) :1468-1479

[3] A snapshot of a transition state analogue of a novel thermophilic esterase belonging to the subfamily of mammalian hormone-sensitive lipase [J].

De Simone, G ;

Galdiero, S ;

Manco, G ;

Lang, D ;

Rossi, M ;

Pedone, C .

JOURNAL OF MOLECULAR BIOLOGY, 2000, 303 (05) :761-771

[4] Co-variation of tRNA abundance and codon usage in Escherichia coli at different growth rates [J].

Dong, HJ ;

Nilsson, L ;

Kurland, CG .

JOURNAL OF MOLECULAR BIOLOGY, 1996, 260 (05) :649-663

[5] Peptide conjugates as tools for the study of biological signal transduction [J].

Eisele, F ;

Owen, DJ ;

Waldmann, H .

BIOORGANIC & MEDICINAL CHEMISTRY, 1999, 7 (02) :193-224

[6]

FORCHHAMMER K, 1990, J BIOL CHEM, V265, P9346

[7] The accuracy of codon recognition by polypeptide release factors [J].

Freistroffer, DV ;

Kwiatkowski, M ;

Buckingham, RH ;

Ehrenberg, M .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2000, 97 (05) :2046-2051

[8]

Gilmore MA, 1999, TOP CURR CHEM, V202, P77

[9] A new UAG-encoded residue in the structure of a methanogen methyltransferase [J].

Hao, B ;

Gong, WM ;

Ferguson, TK ;

James, CM ;

Krzycki, JA ;

Chan, MK .

SCIENCE, 2002, 296 (5572) :1462-1466

[10] CODING FROM A DISTANCE - DISSECTION OF THE MESSENGER-RNA DETERMINANTS REQUIRED FOR THE INCORPORATION OF SELENOCYSTEINE INTO PROTEIN [J].

HEIDER, J ;

BARON, C ;

BOCK, A .

EMBO JOURNAL, 1992, 11 (10) :3759-3766

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