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Peptide binding and photo‐crosslinking to detergent solubilized and to reconstituted transporter associated with antigen processing (TAP)
Author(s) -
Uebel Stephan,
Plantinga Titia,
Weber Peter J.A,
Beck-Sickinger Annette G,
Tampé Robert
Publication year - 1997
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(97)01222-2
Subject(s) - solubilization , chemistry , peptide , transporter associated with antigen processing , transporter , antigen , biochemistry , oligopeptide , microbiology and biotechnology , antigen presentation , biology , in vitro , immunology , gene , cytotoxic t cell
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.