A new fluorescence-based, hydrophobic photolabeling technique for analyzing membrane-associated proteins.
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.
Hess
D
D
Biophysics-Dept. E22, Technical University of Munich, Garching, Germany.
Isenberg
G
G
D000818:Animals; D003720:Densitometry; D005456:Fluorescent Dyes; D005470:Fluorometry / Q000379:methods; D008565:Membrane Proteins / Q000032:analysis*; D015394:Molecular Structure; D011817:Rabbits; D012235:Rhodamines
D005456:Fluorescent Dyes; D008565:Membrane Proteins; D012235:Rhodamines; C022027:lissamine rhodamine B