Watching proteins fold one at a time

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One of the greatest challenges in life sciences is to understand the complex relationship between the amino acid sequence of a protein and its three-dimensional shape. Protein folding is so complex because of countless ways an unfolded peptide can reach the functional folded structure. By avoiding averaging, single-molecule methods are uniquely suited to follow individual folding pathways, test and improve the current models. My research program will make use of an innovative fluorescence microscope that records simultaneously the wavelength, the emission time and the polarization of each photon. The evolution of several molecular parameters can be tracked simultaneously and the accuracy of distance measurements through resonance energy transfer (FRET) will be improved. I will study proteins of different size, origin and function under conditions that favor frequent transitions between the folded and the unfolded state(s). The time course of folding will be detected using a combination of FRET and new environment-sensitive probes. In the cell, protein folding may start at very early stages of peptide synthesis along a narrow tunnel spanning the ribosome, but the characteristic dynamics and conformational changes remain elusive. It is expected that multiparameter single-molecule studies will provide a unique insight into the folding dynamics of nascent peptides, the interaction with the ribosome and with helper biomolecules (chaperones).