Specific interaction of ERp57 and calnexin determined by NMR spectroscopy and an ER two-hybrid system

被引:95
作者
Pollock, S
Kozlov, G
Pelletier, MF
Trempe, JF
Jansen, G
Sitnikov, D
Bergeron, JJM
Gehring, K
Ekiel, I
Thomas, DY
机构
[1] McGill Univ, Fac Med, Dept Biochem, Montreal, PQ H3G 1Y6, Canada
[2] Yale Sch Med, Dept Mol Biophys & Biochem, New Haven, CT USA
[3] McGill Univ, Fac Med, Dept Anat & Cell Biol, Montreal, PQ, Canada
[4] Biotechnol Res Inst, Hlth Sector, Montreal, PQ H4P 2R2, Canada
关键词
calnexin; ERp57; ER yeast two-hybrid; NMR;
D O I
10.1038/sj.emboj.7600119
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
Calnexin and ERp57 act cooperatively to ensure a proper folding of proteins in the endoplasmic reticulum (ER). Calnexin contains two domains: a lectin domain and an extended arm termed the P-domain. ERp57 is a protein disulfide isomerase composed of four thioredoxin-like repeats and a short basic C-terminal tail. Here we show direct interactions between the tip of the calnexin P-domain and the ERp57 basic C-terminus by using NMR and a novel membrane yeast two-hybrid system (MYTHS) for mapping protein interactions of ER proteins. Our results prove that a small peptide derived from the P-domain is active in binding ERp57, and we determine the structure of the bound conformation of the P-domain peptide. The experimental strategy of using the MYTHS two-hybrid system to map interaction sites between ER proteins, together with NMR, provides a powerful new strategy for establishing the function of ER complexes.
引用
收藏
页码:1020 / 1029
页数:10
相关论文
共 27 条
[1]   THE PROGRAM XEASY FOR COMPUTER-SUPPORTED NMR SPECTRAL-ANALYSIS OF BIOLOGICAL MACROMOLECULES [J].
BARTELS, C ;
XIA, TH ;
BILLETER, M ;
GUNTERT, P ;
WUTHRICH, K .
JOURNAL OF BIOMOLECULAR NMR, 1995, 6 (01) :1-10
[2]  
Brunger AT, 1998, ACTA CRYSTALLOGR D, V54, P905, DOI 10.1107/s0907444998003254
[3]   Translational attenuation mediated by an mRNA intron [J].
Chapman, RE ;
Walter, P .
CURRENT BIOLOGY, 1997, 7 (11) :850-859
[4]   N-linked glycans direct the cotranslational folding pathway of influenza hemagglutinin [J].
Daniels, R ;
Kurowski, B ;
Johnson, AE ;
Hebert, DN .
MOLECULAR CELL, 2003, 11 (01) :79-90
[5]   NMRPIPE - A MULTIDIMENSIONAL SPECTRAL PROCESSING SYSTEM BASED ON UNIX PIPES [J].
DELAGLIO, F ;
GRZESIEK, S ;
VUISTER, GW ;
ZHU, G ;
PFEIFER, J ;
BAX, A .
JOURNAL OF BIOMOLECULAR NMR, 1995, 6 (03) :277-293
[6]   Setting the standards: Quality control in the secretory pathway [J].
Ellgaard, L ;
Molinari, M ;
Helenius, A .
SCIENCE, 1999, 286 (5446) :1882-1888
[7]   NMR structures of 36 and 73-residue fragments of the calreticulin P-domain [J].
Ellgaard, L ;
Bettendorff, P ;
Braun, D ;
Herrmann, T ;
Fiorito, F ;
Jelesarov, M ;
Güntert, P ;
Helenius, A ;
Wüthrich, K .
JOURNAL OF MOLECULAR BIOLOGY, 2002, 322 (04) :773-784
[8]   NMR structure of the calreticulin P-domain [J].
Ellgaard, L ;
Riek, R ;
Herrmann, T ;
Güntert, P ;
Braun, D ;
Helenius, A ;
Wüthrich, K .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2001, 98 (06) :3133-3138
[9]   The protein disulphide-isomerase family:: unravelling a string of folds [J].
Ferrari, DM ;
Söling, HD .
BIOCHEMICAL JOURNAL, 1999, 339 :1-10
[10]   TROSY-NMR reveals interaction between ERp57 and the tip of the calreticulin P-domain [J].
Frickel, EM ;
Riek, R ;
Jelesarov, I ;
Helenius, A ;
Wüthrich, K ;
Ellgaard, L .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2002, 99 (04) :1954-1959