Peptide binding and photo‐crosslinking to detergent solubilized and to reconstituted transporter associated with antigen processing (TAP)

FEBS Letters

Tampé, Robert²  Weber, Peter J.A¹  Plantinga, Titia²  Beck-Sickinger, Annette G¹  Uebel, Stephan² 

[1]Eidgenössische Technische Hochschule Zürich, Department Pharmazie, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland

[2]Max-Planck-Institut für Biochemie, Am Klopferspitz 18a, D-82152 Martinsried, Germany

关键词: ABC transporter;    Solubilization;    Reconstitution;    Membrane protein;    Photo-crosslinking;    BIGCHAP;    N;    N-bis[3-d-gluconamidopropyl]-cholamide;    C12E9;    nonaethylene glycol monododecyl ether;    CHAPS;    3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate;    DTT;    1;    4-dithio-dl-threitol;    MEGA 9;    nonanoyl-N-methylglucamide;    MEGA 10;    decanoyl-N-methylglucamide;    NP40;    nonaethylene glycol octylphenyl ether;    PAGE;    polyacrylamide gel electrophoresis;    SDS;    sodium dodecyl sulfate;    sulfobetaine;    N;    N-dimethyl-3-ammino-1-propane-sulfonate;    Tween 80;    polyoxyethylene sorbitan monooleate;   

DOI  :  10.1016/S0014-5793(97)01222-2

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The transporter associated with antigen processing (TAP) is essential for peptide loading onto major histocompatibility (MHC) class I molecules by translocating peptides into the endoplasmic reticulum. We have explored the conditions for detergent solubilization of functionally active, heterologously expressed human TAP from microsomal membranes. The efficiency to solubilize TAP was tested for a variety of detergents as well as for different solubilization conditions. The activity of the solubilized TAP complex was analyzed over time, using a non-radioactive crosslinking assay with a photo-activateable peptide, in the presence or absence of external lipid. The detergent CHAPS was found optimally to retain activity and thus allowed us to reconstitute detergent-solubilized, active TAP into proteoliposomes.

【 授权许可】

Unknown