Energy landscape of streptavidin-biotin complexes measured by atomic forcemicroscopy

The dissociation of ligand and receptor involves multiple transitions betwe en intermediate states formed during the unbinding process. In this paper, we explored the energy landscape of the streptavidin-biotin interaction by using the atomic force microscope (AFM) to measure the unbinding dynamics o f individual ligand-receptor complexes. The rupture force of the streptavid in-biotin bond increased more than 2-fold over a range of loading rates bet ween 100 and 5000 pN/s. Moreover, the force measurements showed two regimes of loading in the streptavidin-biotin force spectrum, revealing the presen ce of two activation barriers in the unbinding process. Parallel experiment s carried out with a streptavidin mutant (W120F) were used to investigate t he molecular determinants of the activation barriers. From these experiment s, we attributed the outer activation barrier in the energy landscape to th e molecular interaction of the '3-4' loop of streptavidin that closes behin d biotin.