文章基本信息

标题：TROSY-NMR reveals interaction between ERp57 and the tip of the calreticulin P-domain
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作者：Eva-Maria Frickel ; Roland Riek ; Ilian Jelesarov 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2002
卷号：99
期号：4
页码：1954-1959
DOI：10.1073/pnas.042699099
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：The lectin chaperone calreticulin (CRT) assists the folding and quality control of newly synthesized glycoproteins in the endoplasmic reticulum (ER). It interacts with ERp57, a thiol-disulfide oxidoreductase that promotes the formation of disulfide bonds in glycoproteins bound by CRT. Here, we investigated the interaction between CRT and ERp57 by using biochemical techniques and NMR spectroscopy. We found that ERp57 binds to the P-domain of calreticulin, an independently folding domain comprising residues 189-288. Isothermal titration calorimetry showed that the dissociation constant of the CRT(189-288)/ERp57 complex is (9.1 {+/-} 3.0) x 10-6 M at 8{degrees}C. Transverse relaxation-optimized NMR spectroscopy provided data on the thermodynamics and kinetics of the complex formation and on the structure of this 66.5-kDa complex. The NMR measurements yielded a value of (18 {+/-} 5) x 10-6 M at 20{degrees}C for the dissociation constant and a lower limit for the first-order exchange rate constant of koff > 1,000 s-1 at 20{degrees}C. Chemical shift mapping showed that interactions with ERp57 occur exclusively through amino acid residues in the polypeptide segment 225-251 of CRT(189-288), which forms the tip of the hairpin structure of this domain. These results are analyzed with regard to the functional mechanism of the CRT/ERp57 chaperone system.