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Second harmonic generation polarization microscopy as a tool for protein structure analysis.

Junichi Kaneshiro1, Yasushi Okada2,3, Tomohiro Shima2

  • 1Laboratory for Comprehensive Bioimaging, RIKEN Center for Biosystems Dynamics Research (BDR), Suita, Osaka 565-0874, Japan.

Biophysics and Physicobiology
|October 30, 2019
PubMed
Summary

Optical second-harmonic generation (SHG) is a new method for analyzing protein structures and dynamics in physiological conditions. This technique successfully detected conformational changes in microtubules and photo-switchable proteins without labeling.

Keywords:
SHG microscopylabel-freeprotein structurestructural variationstructure dynamics

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Area of Science:

  • Biophysics
  • Structural Biology
  • Nonlinear Optics

Background:

  • Established techniques like cryo-electron microscopy, X-ray crystallography, and nuclear magnetic resonance are crucial for protein structure and dynamics analysis.
  • There is a need for methods that can probe protein structure and dynamics under physiological conditions without labeling.

Purpose of the Study:

  • To introduce optical second-harmonic generation (SHG) as a novel tool for protein structure analysis.
  • To demonstrate SHG's capability in detecting conformational changes and dynamics in protein macromolecular assemblies.
  • To link protein structure analysis with biological function under physiological conditions.

Main Methods:

  • Utilized optical second-harmonic generation (SHG), a nonlinear optical scattering process.
  • Developed and employed an SHG polarization microscope for analyzing protein structures in solution.
  • Measured SHG polarization dependence to detect structural differences and temporal variations.

Main Results:

  • Successfully detected conformational differences between microtubules with varying binding molecules using SHG polarization.
  • Captured temporal structural variations in a photo-switchable protein crystal during activation and inactivation.
  • Demonstrated SHG's sensitivity to conformational changes and dynamics in protein macromolecular assemblies.

Conclusions:

  • Optical second-harmonic generation (SHG) shows significant potential as a label-free method for protein structure analysis.
  • SHG enables the study of protein structure and dynamics in physiological solutions at room temperature.
  • This technique can bridge the gap between protein structure and biological function by probing structural changes in situ.