Photoaffmity cross-linking of F1ATPase from the thermophilic bacterium PS 3 by 3'-arylazido-β-alanyl-8-azido ATP

AbstractTo study the localization of the nucleotide binding sites of coupling factor 1 (TF1) from the thermophilic bacterium PS3 we used the bifunctional (cross-linking) 3'-arylazido-β-alanyl-8-azido ATP (DiN3ATP) for photoaffinity labeling. DiN3ATP is hydrolyzed by TF1 in the absence of ultraviolet light. Irradiation (UV light) of TF1 in the presence of DiN3ATP results in a nucleotide-specific reduction of ATPase activity and in a nucleotide-specific formation of different cross-linked proteins (dimers, trimers, oligomers) formed by the major subunits α and/or β. The results suggest that nucleotide binding sites (one, two, possibly all) are located at the interfaces between these subunits