Fluorescence resonance energy transfer mapping of the fourth of six nucleotide-binding sites of chloroplast coupling factor 1.

Equilibrium dialysis measurements of adenine nucleotide binding to chloroplast coupling factor 1 suggest that the enzyme has six binding sites for ADP, adenylyl-beta,gamma-imidodiphosphate (AMP-PNP), and 2'(3')-O-2,4,6-trinitrophenyl-ATP (TNP-ATP). High affinity binding at all six sites requires the divalent cation, Mg2+. Three of the nucleotide-binding sites, sites 1, 2, and 3, have been mapped by fluorescence resonance energy transfer distance measurements (see McCarty, R. E., and Hammes, G. G. (1987) Trends Biochem. Sci. 12, 234-237). Using the same technique, we mapped the location of nucleotide-binding site 4, a tight, exchangeable site (Shapiro, A. B., Huber, A. H., and McCarty, R. E. (1991) J. Biol. Chem. 266, 4194-4200). Two arrangements of the energy transfer map of coupling factor 1 were found which are compatible with the available data. The two arrangements make different predictions about which sites interact in cooperative catalysis.