Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Single-particle selection and alignment with heavy atom cluster-antibody conjugates

G J Jensen1, R D Kornberg

  • 1Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.

Proceedings of the National Academy of Sciences of the United States of America
|August 5, 1998
PubMed
Summary
This summary is machine-generated.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Type VI secretion requires a dynamic contractile phage tail-like structure.

Nature·2012
Same author

The molecular basis of eucaryotic transcription.

Cell death and differentiation·2007
Same author

The eukaryotic gene transcription machinery.

Biological chemistry·2001
Same author

Quantitation of the RNA polymerase II transcription machinery in yeast.

The Journal of biological chemistry·2001
Same author

Alignment error envelopes for single particle analysis.

Journal of structural biology·2001
Same author

A multiprotein complex that interacts with RNA polymerase II elongator.

The Journal of biological chemistry·2001

This study introduces a novel cryoelectron microscopy method using heavy atom clusters to precisely locate and orient biological particles. This technique enhances high-resolution structural determination by improving image selection and reducing defects.

Area of Science:

  • Structural biology
  • Biophysics
  • Biochemistry

Background:

  • Cryoelectron microscopy (cryo-EM) is crucial for determining the high-resolution structures of biological macromolecules.
  • Current cryo-EM methods face challenges in precisely orienting and selecting images of single particles, especially under low-dose conditions optimal for preserving structural integrity.
  • Defects such as specimen movement and particle heterogeneity can further complicate structural analysis.

Purpose of the Study:

  • To propose and evaluate a novel method for selecting and aligning images of single biological particles for high-resolution cryo-EM.
  • To enable precise determination of particle locations and orientations using heavy atom clusters, even with low electron doses and near-focus imaging.
  • To facilitate the selection of high-quality images free from common artifacts.

Related Experiment Videos

Main Methods:

  • Labeling biological particles with multiple heavy atom clusters attached to engineered single-chain Fv antibody fragments.
  • Utilizing the precise locations of heavy atom clusters to determine particle positions and orientations.
  • Developing and implementing computational algorithms for image analysis and particle selection based on cluster projections.
  • Simulating the method's feasibility to assess critical parameters like cluster mobility and localization precision.

Main Results:

  • Demonstrated that heavy atom clusters can precisely determine particle locations and relative orientations.
  • Showed that this labeling strategy allows for image selection free from specimen movement and particle inhomogeneity.
  • Validated the method's potential for high-resolution structural determination under optimal low-dose imaging conditions through simulations.
  • Confirmed the feasibility of using cluster projections for particle orientation and selection.

Conclusions:

  • The proposed heavy atom cluster labeling method offers a robust approach for enhancing single-particle cryo-EM.
  • This technique is expected to improve the accuracy and efficiency of high-resolution structural determination of biological molecules.
  • Further development and experimental validation are warranted to fully realize the potential of this method in structural biology.