Fast Helix Formation in the B Domain of Protein A Revealed by Site-Specific Infrared Probes

Comparison of experimental and computational protein folding studies can be difficult because of differences in structural resolution. Isotope-edited infrared spectroscopy offers a direct measure of structural changes involved in protein folding at the single-residue level. Here we demonstrate the i...

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Bibliographic Details
Published in:Biochemistry (Easton) 2015-03, Vol.54 (9), p.1758-1766
Main Authors: Davis, Caitlin M, Cooper, A. Kat, Dyer, R. Brian
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Circular Dichroism
computer simulation
Infrared Rays
infrared spectroscopy
Kinetics
methionine
Methionine - chemistry
Molecular Dynamics Simulation
Molecular Probes - chemistry
Molecular Sequence Data
Peptides - chemistry
Protein Folding
Protein Structure, Secondary
Protein Structure, Tertiary
protein synthesis
recombinant proteins
Spectrophotometry, Infrared - methods
Spectroscopy, Fourier Transform Infrared
Staphylococcal Protein A - chemistry
temperature
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Summary:Comparison of experimental and computational protein folding studies can be difficult because of differences in structural resolution. Isotope-edited infrared spectroscopy offers a direct measure of structural changes involved in protein folding at the single-residue level. Here we demonstrate the increased resolution of site-specific infrared probes to the peptide backbone in the B domain of staphylococcal protein A (BdpA). 13C18O-labeled methionine was incorporated into each of the helices using recombinant protein expression. Laser-induced temperature jumps coupled with infrared spectroscopy were used to probe changes in the peptide backbone on the submillisecond time scale. The relaxation kinetics of the buried helices, solvated helices, and labeled positions were measured independently by probing the corresponding bands assigned in the amide I region. Using these wavelength-dependent measurements, we observe a fast nanosecond phase and slower microsecond phase at each position. We find at least partial formation of helices 1–3 in the fast intermediate state that precedes the transition state. These measurements provide direct, time-resolved experimental evidence of the early formation of partial helical structure in helices 1 and 3, supporting folding models proposed by computer simulations.
ISSN:0006-2960
1520-4995
1520-4995
DOI:10.1021/acs.biochem.5b00037