Ultrafast thermally induced unfolding of RNase A.
A temperature jump (T-jump) method capable of initiating thermally induced processes on the picosecond time scale in aqueous solutions is introduced. Protein solutions are heated by energy from a laser pulse that is absorbed by homogeneously dispersed molecules of the dye crystal violet. These act as transducers by releasing the energy as heat to cause a T-jump of up to 10 K with a time resolution of 70 ps. The method was applied to the unfolding of RNase A. At pH 5.7 and 59 degrees C, a T-jump of 3-6 K induced unfolding which was detected by picosecond transient infrared spectroscopy of the amide I region between 1600 and 1700 cm-1. The difference spectral profile at 3.5 ns closely resembled that found for the equilibrium (native-unfolded) states. The signal at 1633 cm-1, corresponding to the beta-sheet structure, achieved 15 +/- 2% of the decrease found at equilibrium, within 5.5 ns. However, no decrease in absorbance was detected until 1 ns after the T-ump. The disruption of beta-sheet therefore appears to be subject to a delay of approximately 1 ns. Prior to 1 ns after the T-jump, water might be accessing the intact hydrophobic regions.
Phillips
C M
CM
Department of Chemistry, University of Pennsylvania, Philadelphia 19104, USA.
Mizutani
Y
Y
Hochstrasser
R M
RM
D000818:Animals; D002417:Cattle; D006358:Hot Temperature; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D007700:Kinetics; D015394:Molecular Structure; D010179:Pancreas / Q000201:enzymology; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012259:Ribonuclease, Pancreatic / Q000737:chemistry*; D012996:Solutions; D017550:Spectroscopy, Fourier Transform Infrared; D013816:Thermodynamics; D014867:Water
D012996:Solutions; D014867:Water; D012259:Ribonuclease, Pancreatic