Measurement of the forced rupture of biotin from streptavidin, using the fo
rce microscope, revealed a logarithmic dependence on the rate of loading. T
ransition state theory predicts that the rupture force is dependent on the
rate of force loading and the dissociation rate constant of the interaction
. Analysis of the dependence of the rupture force with loading rate reveals
that the barrier to unbinding under the loading rates employed here is sit
uated between 0.12 and 0.18 nm away from the bound state. The position of t
he barrier determined is similar in value to the results obtained using com
plementary force techniques, and is matched by calculations from computatio
nal simulation. Thus, force rupture measurements may be used to profile the
internal energy pathway of molecular dissociation events. The results sugg
est, however, that the loading rates employed here, using cantilever retrac
t velocities as low as 1 nm s(-1), are still too high to explore the whole
of the unbinding energy landscape, and suggests further avenues for instrum
ental and experimental development.