Fast Events in Protein Folding Initiated by Nanosecond Laser Photolysis

Initiation of protein folding by light can dramatically improve the time resolution of kinetic studies. Here we present an example of an optically triggered folding reaction by using nanosecond photodissociation of the heme-carbon monoxide complex of reduced cytochrome c. The optical trigger is base...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1993-12, Vol.90 (24), p.11860-11864
Main Authors: Jones, Colleen M., Henry, Eric R., Hu, Yi, Chan, Chi-Kin, Luck, Stan D., Bhuyan, Abani, Roder, Heinrich, Hofrichter, James, Eaton, William A.
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Binding Sites
Biological and medical sciences
Biology
Carbon Monoxide - metabolism
Conformational dynamics in molecular biology
Cytochrome c Group - chemistry
Cytochrome c Group - metabolism
Cytochromes
Datasets
Dichroism
Fluorescence
Fundamental and applied biological sciences. Psychology
Guanidine
Guanidines - pharmacology
Heme - metabolism
Horses
Kinetics
Lasers
Ligands
Methionine
Molecular biophysics
Optics
Photolysis
Physics
Protein Conformation
Protein Folding
Protein Structure, Secondary
Proteins
Spectrophotometry
Time Factors
Wavelengths
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Summary:Initiation of protein folding by light can dramatically improve the time resolution of kinetic studies. Here we present an example of an optically triggered folding reaction by using nanosecond photodissociation of the heme-carbon monoxide complex of reduced cytochrome c. The optical trigger is based on the observation that under destabilizing conditions cytochrome c can be unfolded by preferential binding of carbon monoxide to the covalently attached heme group in the unfolded state. Photodissociation of the carbon monoxide thus triggers the folding reaction. We used time-resolved absorption spectroscopy to monitor binding at the heme. Before folding begins we observe transient binding of both nonnative and native ligands from the unfolded polypeptide on a microsecond time scale. Kinetic modeling suggests that the intramolecular binding of methionine-65 and -80 is faster than that of histidine-26 and -33, even though the histidines are closer to the heme. This optical trigger should provide a powerful method for studying chain collapse and secondary structure formation in cytochrome c without any limitations in time resolution.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.90.24.11860