Nucleotide binding to chloroplast F1‐ATPase (CF1) cannot be explained by a two‐site model

Abstract

AbstractThe kinetics of nucleotide binding to the chloroplast coupling factor, CF1, have been analysed by stopped‐flow measurements using the fluorescent trinitrophenyl analog TNP‐ADP. Experimental determined time courses were compared with several different theoretical time courses. The following observations were obtained: (a) in stopped‐flow experiments the half time of the reaction was at highest with a ligand‐to‐enzyme ratio of 2:1. The half time of the binding reaction decreased, if the ratio was lower than 2:1. (b) This characteristic cannot be explained by a bimolecular reaction mechanism of several independent (classes of) binding sites, (c) Theoretical time courses according to a “two‐site” model with the inclusion of a conformational change of each site after binding of ligand showed the observed characteristic (see (a)), (d) However, this “two‐site” model including a conformational change of each site after binding of ligand failed to simulate the measured time courses of nucleotide binding.