A new fluorescence‐based, hydrophobic photolabeling technique for analyzing membrane‐associated proteins

FEBS Letters

Hess, D¹  Isenberg, G¹ 

[1] Biophysics-Dept. E22, Technical University of Munich, James Franck Str., D-85747 Garching, Germany

关键词: Fluorescence;    Hydrophobic photolabelling;    Membrane-associated protein;    Talin;    RNase;    G-actin;    DMPC;    dimyristoyl-phosphatidyl-choline;    DMPG;    dimyristoyl-phospho-rac-glycerol;    DMTAP;    dimyristoyl-tetramethylammoniumpropane;    DSC;    differential scanning calorimetry;    RNase A;    ribonucleic acid hydrolase A;    SANU;    N 1-(N 2-(sulforhodamine B sulfonyl)aminoethyl;    11-(5-azido-1-naphtoxy))undecanylamide;    SDS-PAGE;    sodium dodecylsulfate polyacrylamide gel electrophoresis;    TLC;    thin layer chromatography;   

DOI  :  10.1016/S0014-5793(99)00155-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

We introduce a new, fluorescent and photoactivatable fatty acid derivative (SANU) for hydrophobic labelling of membrane-bound proteins. The technique allows fast and highly sensitive screening of hydrophobically inserting proteins analyzed by SDS-PAGE with a detection limit below 0.1 pmol. A reliable calculation of labelling efficiencies is achieved by simultaneous densitometry of fluorescence and protein staining. We have applied the new technique on the membrane inserting protein talin, G-actin, and, as a negative control, on RNase, which only binds electrostatically to negatively charged lipid interfaces. In several ways superior to radiolabelling, we can recommend this technique for all laboratories under any circumstances.

【 授权许可】

Unknown