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 a...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1995-08, Vol.92 (16), p.7292-7296
Main Authors: Phillips, C. M., Mizutani, Y., Hochstrasser, R. M.
Format: Article
Language:English
Subjects:
Amides
Animals
Biochemistry
Cattle
Dyes
Heat
Hot Temperature
Hydrogen Bonding
In Vitro Techniques
Infrared radiation
Kinetics
Lasers
Molecular Structure
Molecules
Pancreas - enzymology
Protein Folding
Protein Structure, Secondary
Ribonuclease, Pancreatic - chemistry
Solutions
Spectroscopy, Fourier Transform Infrared
Temperature
Thermodynamics
Time dependence
Water
Water temperature
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Description
Summary: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⚬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 β-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-jump. The disruption of β-sheet therefore appears to be subject to a delay of ≈1 ns. Prior to 1 ns after the T-jump, water might be accessing the intact hydrophobic regions.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.92.16.7292