A rapid and efficient purification method for recombinant annexin V for biophysical studies

FEBS Letters

Berendes, R.¹  Burger, A.¹  Demange, P.¹  Huber, R.¹  Voges, D.¹ 

[1] Max-Planck-Institute für Biochemie, D-82152 Martinsried, Germany

关键词: Annexin;    Ion channel;    Protein purification;    Crystallization;    Patch clamp;    Electron microscopy;    DNase;    desoxyribonuclease;    EDTA;    ethylene diamine tetraacetic acid;    E. coli;    Escherichia coli;    HPLC;    high-pressure liquid chromatography;    IPTG;    isopropylthiogalactoside;    PAGE;    polyacrylamide gel-electrophoresis;    PMSF;    phenylmethylsulfonyl fluoride;    RNase;    ribonuclease;    SDS;    sodium dodecyl sulfate;    v/v;    volume per volume;    w/v;    weight per volume;   

DOI  :  10.1016/0014-5793(93)80185-W

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Annexin V binds in a calcium-dependent manner to acidic phospholipids and exhibits ion channel activity in vitro. We are investigating mutants of annexin V by single channel measurements, X-ray crystallography and electron microscopy in order to understand the structure-function relationships of the ion channel activity. We describe here a method to obtain very pure reeombinant annexin V required for such studies. The initial step is the mild opening of the bacterial cells by an osmotic shock. In the purification procedure, use is made of the reversible calcium-mediated binding of annexin V to liposomes. In the last purification step the protein is subjected to ion-exchange chromatography and elutes as a single peak free of any detectable contaminants.

【 授权许可】

Unknown