Intermediate states of molecular self-assembly from liquid-cell electron microscopy

Area of Science:

• Macromolecular science
• Biophysics
• Materials science

Background:

• Traditional single-molecule techniques lack the ability to capture dynamic conformational changes during self-assembly.
• Understanding real-time molecular behavior is crucial for fields like nanotechnology and molecular biology.

Purpose of the Study:

• To develop and demonstrate a novel method for observing time-dependent conformational adaptations of macromolecules.
• To investigate the kinetics of DNA hybridization and self-assembly at the single-molecule level.

Main Methods:

• Utilized graphene liquid-cell electron microscopy (GLCEM) with low-energy, low-dose electrons.
• Observed single-stranded DNA (ssDNA) hybridization into double-stranded DNA (dsDNA) in real-time.
• Compared DNA sequences with varying microstructures (random, blocky, palindromic hairpin).

Main Results:

• Resolved conformational adaptations of DNA over minutes, limited by motion blurring.
• Observed hybridization coupled with enhanced translational mobility and torsion-induced rotation.
• Identified prevalent transient loops in error-correction, transient melting, and failed binding events.
• Discovered previously unobserved metastable intermediate states for certain DNA sequences.

Conclusions:

• GLCEM provides unprecedented temporal resolution for macromolecular dynamics, surpassing traditional methods.
• The study revealed surprising molecular behaviors including enhanced mobility and extensive looping during DNA hybridization.
• The methodology is generalizable for visualizing other organic macromolecules and their interactions.