Fluorescence quenching: A tool for single-molecule protein-folding study

By using titin as a model system, we have demonstrated that fluorescence qu enching can be used to study protein folding at the single molecule level. The unfolded titin molecules with multiple dye molecules attached are able to fold to the native state. In the native folded state, the fluorescence f rom dye molecules is quenched due to the close proximity between the dye mo lecules. unfolding of the titin leads to a dramatic increase in the fluores cence intensity. Such a change makes the folded and unfolded states of a si ngle titin molecule clearly distinguishable and allows us to measure the fo lding dynamics of individual titin molecules in real time. We have also sho wn that fluorescence quenching can signal folding and unfolding of a small protein with only one immunoglobulin domain.