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

Subunit counting in membrane-bound proteins.

Maximilian H Ulbrich1, Ehud Y Isacoff

  • 1Department of Molecular and Cell Biology, University of California, Berkeley Material Science & Physical Bioscience Divisions, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA.

Nature Methods
|March 21, 2007
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

The small bacterial membrane protein YohP induces nucleoid condensation in <i>E. coli</i> and inhibits oligomerization of antimicrobial peptides.

microLife·2025
Same author

Reversible Antagonism of Dopamine D1 Receptor Using a Photoswitchable Remotely Tethered Ligand.

ACS chemical biology·2025
Same author

Deficiency in transmitter release triggers homeostatic transcriptional changes that increase presynaptic excitability.

Proceedings of the National Academy of Sciences of the United States of America·2025
Same author

Reversible Antagonism of Dopamine D1 Receptor using a Photoswitchable Remotely Tethered Ligand.

bioRxiv : the preprint server for biology·2025
Same author

Balanced synapse-to-synapse short-term plasticity ensures constant transmitter release.

Current biology : CB·2025
Same author

Transgenic human nephrin in Drosophila nephrocytes facilitates variant analysis.

Kidney international·2025
Same journal

ClairS: a deep-learning method for long-read tumor-normal pair somatic small variant calling.

Nature methods·2026
Same journal

RNAbpFlow: base pair-augmented SE(3) flow matching for conditional RNA 3D structure generation.

Nature methods·2026
Same journal

Spatio-DARLIN enables robust and efficient in situ lineage tracing in mice at single-cell resolution.

Nature methods·2026
Same journal

EasyGrid: a versatile platform for automated cryo-EM sample preparation and quality control.

Nature methods·2026
Same journal

Cloud-based microscope enables live neuroimaging for 24 h and beyond with worldwide access.

Nature methods·2026
Same journal

Deep molecular profiling in three dimensions.

Nature methods·2026
See all related articles

Determining membrane protein subunit number is challenging. A new single-molecule method using GFP bleaching in live cells accurately counts subunits, revealing NMDA receptor composition.

Area of Science:

  • Biochemistry and Molecular Biology
  • Cell Biology
  • Neuroscience

Background:

  • Understanding the stoichiometry of membrane-bound proteins is crucial for elucidating their function.
  • Existing methods often require disrupting the native membrane environment, limiting their applicability.
  • Accurate determination of protein subunit composition in situ remains a significant challenge.

Purpose of the Study:

  • To develop and validate a novel single-molecule technique for quantifying subunit numbers of membrane proteins in live cells.
  • To overcome the limitations of current methods that disrupt the cellular membrane environment.
  • To apply the technique to determine the subunit composition of specific protein complexes, such as NMDA receptors.

Main Methods:

  • A single-molecule imaging approach was employed, utilizing the photobleaching steps of Green Fluorescent Protein (GFP) fused to the protein of interest.

Related Experiment Videos

  • The method was tested and validated using proteins with known stoichiometry expressed in Xenopus laevis oocytes.
  • Live cell membrane environments were maintained throughout the observation period.

    • Main Results:

    • The developed single-molecule technique successfully counted subunits of membrane-bound proteins without disrupting their native environment.
    • Validation in Xenopus oocytes confirmed the accuracy of the subunit counting method.
    • The composition of NMDA receptors, specifically involving NR1 and NR3 subunits, was resolved using this technique.

    Conclusions:

    • This study presents a robust single-molecule method for determining the subunit number and stoichiometry of membrane proteins in live cells.
    • The technique offers a valuable tool for studying protein complex assembly and function in their native cellular context.
    • The resolved composition of NMDA receptors highlights the utility of this method in neuroscience research.