Asymmetry of catalytic but not of noncatalytic sites on Escherichia coli F1-ATPase in solution as observed using electron spin resonance spectroscopy.

We have employed electron spin resonance (ESR) spectroscopy using different spin-labeled nucleotides to probe the environment of nucleotides bound at catalytic and noncatalytic nucleotide binding sites of the Escherichia coli F1-ATPase. We found that nucleotides bound in the noncatalytic binding sites were strongly immobilized and resulted in ESR spectra with one single corresponding spectral component. Nucleotide bound at the catalytic binding sites gave rise to two different signals in the ESR spectra indicative of two distinct conformations of the catalytic sites of the protein. One conformation of the catalytic sites is very tight, resulting in signals identical to those of the noncatalytic sites, while the second type of catalytic sites permitted an unusually high mobility of the bound spin-labeled nucleotide. The findings are compared to the requirements of the binding change mechanism and to the features of the nucleotide binding sites as elucidated from the X-ray structural model of the beef heart mitochondrial enzyme.