Crystallization of a mammalian membrane protein overexpressed in Saccharomyces cerevisiae

被引：75

作者：

Jidenko, M

Nielsen, RC

Sorensen, TLM

Moller, JV

le Maire, M

Nissen, P

Jaxel, C ^{[1
]}

机构：

[1] CEA Saclay, URA 2096, CNRS, F-91191 Gif Sur Yvette, France

[2] CEA Saclay, Serv Biophys Fonct Membranaires, Dept Biol Joliot Curie, F-91191 Gif Sur Yvette, France

[3] Univ Paris Sud, Lab Rech Associe 17V, Paris, France

[4] Univ Paris Sud, Inst Federat Rech 46, Paris, France

[5] Aarhus Univ, Dept Mol Biol, DK-8000 Aarhus, Denmark

[6] Aarhus Univ, Inst Physiol & Biophys, Dept Biophys, DK-8000 Aarhus, Denmark

来源：

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA | 2005年 / 102卷 / 33期

关键词：

overexpression; Ca2+-ATPase; SERCA1a;

D O I：

10.1073/pnas.0503986102

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

The Ca2+-ATPase SERCA1a (sarcoplasmic-endoplasmic reticulum Ca2+-ATPase isoform 1a) from rabbit has been overexpressed in Saccharomyces cerevisiae. This membrane protein was purified by avidin agarose affinity chromatography based on natural biotinylation in the expression host, followed by HPLC gel filtration. Both the functional and structural properties of the overexpressed protein validate the method. Thus, calcium-dependent ATPase activity and calcium transport are essentially intact after reconstitution in proteoliposomes. Moreover, the recombinant protein crystallizes in a form that is isomorphous to the native SERCA1a protein from rabbit, and the diffraction properties are similar. This represents a successful crystallization of a mammalian membrane protein derived from a heterologous expression system, and it opens the way for the study of mutant forms of SERCA1a.

引用

页码：11687 / 11691

页数：5