Experimental investigation on the folding dynamics of proteins

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Proteins can fold from extended conformations of unfolded polypeptides to tightly packed and functional structures. Therefore, the mechanism of protein folding is important for the understanding of the principles of protein structures and functions. In this study, we investigated the structural events involved in the dynamics of protein folding based on the time-resolved technique combined with various spectroscopic techniques. We utilized small angle X-ray scattering for the characterization of protein compactness and circular dichroism, infrared absorption, fluorescence and resonance Raman spectroscopies for the detection of secondary and tertiary structure contents. The rapid mixing method was developed to increase the time resolution of the dynamics. We investigated the folding process of apomyogrobin, single chain monnelin, cytochrome c, heme oxygenase and ribonuclease A, and observed that the folding transitions of proteins with more than 100 residues generally follow the collapse and search mechanism, in which the rapid collapse and the subsequent search in the collapsed conformations are the characteristic events. Furthermore, we presented evidence suggesting that the initial collapse is caused by the coil-globule transition. Interestingly, it was observed that proteins of less than 100 residues usually do not accumulate kinetic intermediates, and possess the expanded R_g after the refolding jump. We propose that T_θ for small proteins is lower than the experimental temperatures causing small proteins to remain expanded until the moment of folding. Thus, the coil-globule transition offers a general explanation for the initial folding mechanisms of small and larger proteins. A further understanding on the molecular mechanism of the collapse will likely give an important insight that might help improve the structure prediction.