Temperature Induced Protein Unfolding and Folding of RNase a Studied by Time-Resolved Infrared Spectroscopy

When a protein finds its native three-dimensional structure from the unstructuredamino-acid chain various processes spanning a large time range are relevant. Tounderstand the mechanism of protein folding one needs to cover the entire folding/refolding (U↔N) reaction on a structural level. In the case of RNase A, the mainstructural changes occur in the ms time range, that can be monitored with rapid-scan-FTIR spectroscopy combined with rapid mixing techniques. To induce unfolding weinject aqueous protein solution into a hot IR cuvette and record the time course of thespectral changes. A lag phase is found when the unfolding conditions are relatively weak,suggesting an unfolding intermediate.