Insights into ATP synthase structure and function using affinity and site-specific spin labeling

A variety of different approaches has been used during the last couple of decades to investigate structure and function relationships within the catalytic portion of the FoF1-ATP synthase and of its interactions with the proton-translocator F-o. In our group, we employ ESR spectroscopy with the use of stable organic radicals, so-called spin labels, as reporter groups. The radicals are either attached to substrates/ligands or specifically inserted into the protein structure by "site-specific spin labeling." Both approaches bear intrinsic advantages for their special uses and result in the specific information that is available through ESR, e.g., structural changes due to binding of effector molecules (e.g., Mg2+ ions), conformational transitions during catalytic turnover, distance information on radicals bound at 20 Angstrom or less, and information on the binding characteristics of labeled substrates. This review summarizes the results of a variety of different approaches we have used during the last years to study, with the help of ESR spectroscopy, the structure of the nucleotide binding sites of F-1-ATPases of different origins as well as interactions with F-o subunits.