Suppressing sidechain modes and improving structural resolution for 2D IR spectroscopy via vibrational lifetimes

Area of Science:

• Biophysical Chemistry
• Spectroscopy
• Structural Biology

Background:

• Protein structure analysis often employs vibrational spectroscopy with 13C18O-labeling for enhanced resolution.
• Spectral overlap between isotope-labeled backbone carbonyls and protein sidechains complicates data interpretation.
• Distinguishing between backbone and sidechain vibrational modes is crucial for accurate structural determination.

Purpose of the Study:

• To investigate the use of pulse waiting times in 2D IR spectroscopy for suppressing interfering sidechain signals.
• To determine if vibrational lifetimes can differentiate between backbone amide I' modes and sidechain modes.
• To assess the potential for assigning secondary structures based on lifetime differences of 13C18O-amide I' modes.

Main Methods:

• Utilized two-dimensional infrared (2D IR) spectroscopy with varying pump-probe pulse waiting times.
• Studied model disordered and β-sheet peptides incorporating 13C18O-labeling.
• Analyzed vibrational lifetimes of backbone amide I' modes and sidechain modes.

Main Results:

• Delayed waiting times in 2D IR spectroscopy effectively suppressed protein sidechain vibrational modes.
• Backbone amide I' modes exhibited longer vibrational lifetimes than sidechain modes in both disordered and β-sheet peptides.
• β-sheet structures showed longer amide I' mode lifetimes compared to disordered structures.

Conclusions:

• Pulse waiting time in 2D IR spectroscopy is a viable method for suppressing unwanted sidechain signals.
• Vibrational lifetime differences can be leveraged to assign secondary structures of labeled protein residues.
• This approach enhances the analysis of protein structures using vibrational spectroscopy.