Related Experiment Video

Updated: Aug 9, 2026

Reconstitution of a Kv Channel into Lipid Membranes for Structural and Functional Studies
10:22

Reconstitution of a Kv Channel into Lipid Membranes for Structural and Functional Studies

Published on: July 13, 2013

Reconstitution of protein translocation activity from partially solubilized microsomal vesicles

D L Zimmerman1, P Walter

  • 1Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0448.

The Journal of Biological Chemistry
|March 5, 1990
PubMed

Insights

Protein translocation activity in rough endoplasmic reticulum vesicles can be restored after partial solubilization. Removing n-octyl-beta-glucopyranoside detergent allowed reconstitution of translocation-competent membranes.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Microsomal vesicles from the rough endoplasmic reticulum (RER) are crucial for protein translocation.
  • Detergent treatment can disrupt membrane integrity and affect protein translocation activity.

Purpose of the Study:

  • To investigate the recovery of protein translocation activity in RER-derived microsomal vesicles after partial solubilization.
  • To establish conditions for reconstituting translocation-competent membranes.

Main Methods:

  • Utilized a reconstitution assay to assess protein translocation.
  • Employed two independent methods to determine partial solubilization conditions for RER microsomes using n-octyl-beta-glucopyranoside.
  • Evaluated membrane integrity and translocation activity following detergent treatment and removal.

Main Results:

  • Partial solubilization of RER microsomal membranes with n-octyl-beta-glucopyranoside was achieved.
  • Membranes with perturbed lipid bilayer integrity due to detergent were inactive for translocation.
  • Successful reconstitution of translocation-competent membranes was demonstrated after detergent removal.

Conclusions:

  • Protein translocation activity can be recovered in RER-derived vesicles.
  • Detergent removal is essential for restoring the translocation competence of solubilized microsomal membranes.
  • This study provides a method for reconstituting functional RER vesicles.

More Related Videos

Reconstitution of a Transmembrane Protein, the Voltage-gated Ion Channel, KvAP, into Giant Unilamellar Vesicles for Microscopy and Patch Clamp Studies11:42

Reconstitution of a Transmembrane Protein, the Voltage-gated Ion Channel, KvAP, into Giant Unilamellar Vesicles for Microscopy and Patch Clamp Studies

Published on: January 22, 2015

Construction of Out-of-Equilibrium Metabolic Networks in Nano- and Micrometer-Sized Vesicles10:56

Construction of Out-of-Equilibrium Metabolic Networks in Nano- and Micrometer-Sized Vesicles

Published on: April 12, 2024

Related Concept Videos

Translocation of Proteins into the Mitochondria01:19

Translocation of Proteins into the Mitochondria

Mitochondrial precursors are translocated to the internal subcompartments via independent mechanisms involving distinct protein machineries called translocases.
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...
Cotranslational Protein Translocation01:20

Cotranslational Protein Translocation

Translocation of proteins across membranes is an ancient process that occurs even in bacteria and archaebacteria. In fact, the components of the translocation machinery are still conserved between prokaryotes and eukaryotes.
Sec61 channel partners for cotranslational translocation
During cotranslational translocation, the Sec61 channel partners with the signal recognition particle (SRP), the signal recognition particle receptor (SR), and the ribosomes to transport the nascent polypeptide chain...