Probing protein multidimensional conformational fluctuations by single-molecule multiparameter photon stamping...

Area of Science:

• Biophysics
• Biochemistry
• Molecular Biology

Background:

• Protein conformational motions are complex and crucial for function.
• Traditional methods often simplify protein dynamics to one dimension.
• Understanding multi-dimensional dynamics is key to enzyme function.

Purpose of the Study:

• To probe the multi-dimensional conformational dynamics of proteins.
• To investigate T4 lysozyme's hinge-bending motions beyond one dimension.
• To correlate protein dynamics with enzymatic functions.

Main Methods:

• Developed single-molecule multiparameter photon stamping spectroscopy.
• Integrated fluorescence anisotropy, Förster Resonance Energy Transfer (FRET), and fluorescence lifetime.
• Simultaneously observed site-to-site dynamics and molecular rotational motions.

Main Results:

• Observed wide-distributed rotational flexibility and multiple intermediate conformational states.
• Presented a comprehensive view of T4 lysozyme's open-close hinge-bending motions.
• Identified dynamic and static inhomogeneity in protein conformational fluctuations.

Conclusions:

• Multi-dimensional protein dynamics significantly impact enzymatic functions.
• The developed spectroscopy provides fundamental understanding of enzyme turnover dynamics.
• Revealed active-site conformational fluctuations not detectable by conventional methods.