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Conventional electron microscopy (EM) involves dehydration, fixation, and staining of biological samples, which distorts the native state of biological molecules and results in several artifacts. Also, the high-energy electron beam damages the sample and makes it difficult to obtain high-resolution images. These issues can be addressed using cryo-EM, which uses frozen samples and gentler electron beams. The technique was developed by Jacques Dubochet, Joachim Frank, and Richard Henderson, for...
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DoubleHelix: nucleic acid sequence identification, assignment and validation tool for cryo-EM and crystal structure

Grzegorz Chojnowski1

  • 1European Molecular Biology Laboratory, Hamburg Unit, Notkestraße 85, 22607 Hamburg, Germany.

Nucleic Acids Research
|July 3, 2023
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Summary

DoubleHelix is a new method for assigning and validating nucleic acid sequences in cryo-electron microscopy (cryo-EM) and macromolecular crystallography (MX) structures. It helps identify errors, especially in low-resolution data where interpretation is challenging.

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Area of Science:

  • Structural biology
  • Biochemistry
  • Computational biology

Background:

  • Accurate sequence assignment is crucial for interpreting macromolecular structures from cryo-EM and MX.
  • Existing model validation tools are insufficient for nucleic acid structures, leading to potential errors.
  • Challenges in sequence assignment are amplified at lower resolutions.

Purpose of the Study:

  • To introduce doubleHelix, a comprehensive method for nucleic acid sequence assignment, identification, and validation.
  • To provide a tool that assists in building accurate nucleic acid models from cryo-EM and MX data.
  • To improve the reliability of structural models, particularly for nucleic acids.

Main Methods:

  • Development of a neural network classifier for nucleobase identification.
  • Implementation of a sequence-independent secondary structure assignment approach.
  • Integration of these components into the doubleHelix software.

Main Results:

  • DoubleHelix successfully assists in nucleic acid sequence assignment, especially at lower resolutions.
  • The method identified sequence assignment errors in ribosome structures from the Protein Data Bank that were missed by other validation tools.
  • Demonstrated utility in both cryo-EM and MX structural data.

Conclusions:

  • DoubleHelix offers a robust solution for nucleic acid sequence assignment and validation in cryo-EM and MX.
  • The tool enhances model accuracy and reliability, addressing a gap in current validation strategies.
  • It is particularly valuable for low-resolution structures and complex systems like ribosomes.