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Related Concept Videos

Introduction to Membrane Proteins01:16

Introduction to Membrane Proteins

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The cell membrane, or plasma membrane, is an ever-changing landscape. It is described as a fluid mosaic where various macromolecules are embedded in the phospholipid bilayer. Among the macromolecules are proteins. The protein content varies across cell types. For example, mitochondrial inner membranes contain ~76% protein content, while myelin contains ~18% protein content. Individual cells contain many types of membrane proteins—red blood cells contain over 50—and different cell...
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Membrane Proteins01:30

Membrane Proteins

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Plasma membranes have integral transmembrane proteins involved in facilitated transport. These proteins are collectively referred to as transport proteins, and they function as either channels for the material or as carriers themselves. Channel proteins have hydrophilic domains exposed to the intracellular and extracellular fluids and a hydrophilic channel through their core that provides a hydrated opening for solutes to pass through the membrane layers. Passage through the channel allows...
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Protein Diffusion in the Membrane01:24

Protein Diffusion in the Membrane

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Proteins show rotational as well as lateral diffusion across the membrane. The lateral diffusion of proteins was confirmed through the cell fusion experiment where mouse and human cells were fused, resulting in hybrid cells. When the human and mouse cells fused, the specific membrane proteins on human and mouse cells were marked with the red and green-fluorescent markers, respectively. Initially, the red and green fluorescence was located on the respective hemisphere of the cell. As time...
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Tail-anchoring of Proteins in the ER Membrane01:45

Tail-anchoring of Proteins in the ER Membrane

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Tail-anchored, or TA, proteins are estimated to make up to 3-5% of membrane proteins found in the eukaryotic cell. Such proteins have a single transmembrane domain located approximately 30 amino acid residues upstream from the C-terminal end. As a result, the signal recognition particle (SRP) cannot guide a TA protein to the ER membrane for cotranslational insertion. Hence, they are integrated into the ER membrane post-translationally using their C-terminal end as the anchor. TA proteins...
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Protein Translocation Machinery on the ER Membrane01:28

Protein Translocation Machinery on the ER Membrane

6.9K
The translocon complex situated on the ER membrane is the main gateway for the protein secretory pathway. It facilitates the transport of nascent peptides into the ER lumen and their insertion into the ER membrane.
Sec61 protein conducting channel
In eukaryotes, the translocon complex comprises a core heterotrimeric translocator channel called the Sec61 complex. This channel includes three transmembrane proteins, Sec61α, Sec61β, and Sec61γ, and is the largest subunit of the...
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Protein Transport to the Inner Chloroplast Membrane01:18

Protein Transport to the Inner Chloroplast Membrane

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Proteins targeted to the inner chloroplast membrane, or plastid proteins, are transported by two general pathways: the stop-transfer and the re-insertion or post-import pathways. Most plastid proteins carry N-terminal transit sequences and internal import sequences targeting it to the specific chloroplast subcompartment. Proteins targeted by the stop-transfer pathway have internal hydrophobic sequences that inhibit their translocation into the stroma. As a result, these precursors are arrested...
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Related Experiment Video

Updated: Feb 9, 2026

Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli
08:46

Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli

Published on: January 6, 2015

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Single molecule fluorescence for membrane proteins.

Oliver K Castell1, Patricia M Dijkman2, Daniel N Wiseman3

  • 1School of Pharmacy and Pharmaceutical Sciences, College of Biomedical and Life Sciences, Cardiff University, King Edward VII Avenue, Cardiff CF10 3NB, UK.

Methods (San Diego, Calif.)
|June 2, 2018
PubMed
Summary
This summary is machine-generated.

Researchers developed droplet interface bilayers (DIBs) for studying individual membrane proteins. This method, combined with TIRF microscopy, provides detailed insights into protein behavior and dynamics in vitro.

Keywords:
Droplet interface bilayerFRETMembrane proteinPhotobleachingSingle molecule fluorescenceTIRF

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Single Molecule Fluorescence Microscopy on Planar Supported Bilayers
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Area of Science:

  • Biochemistry
  • Biophysics
  • Cell Biology

Background:

  • Cell membranes are complex structures crucial for cellular function.
  • Understanding individual membrane protein behavior requires in vitro analysis.
  • Existing methods may have limitations in resolving molecular dynamics.

Purpose of the Study:

  • To detail the creation and application of droplet interface bilayers (DIBs) for single-molecule membrane protein studies.
  • To demonstrate how DIBs coupled with TIRF microscopy can yield spatiotemporal and kinetic data.
  • To provide guidance for implementing this technique in research.

Main Methods:

  • Modification of membrane proteins for fluorescent labeling.
  • Expression and purification of labeled membrane proteins.
  • Formation of DIBs and spontaneous protein incorporation.
  • Single-molecule Total Internal Reflection Fluorescence (TIRF) microscopy.
  • Techniques including step-wise photobleaching, Förster Resonance Energy Transfer (FRET), and single particle tracking.

Main Results:

  • Successful incorporation of membrane proteins into DIBs.
  • Acquisition of spatiotemporal and kinetic information for individual proteins.
  • Determination of parameters like oligomerization state and protein dynamics.
  • Demonstration of the utility of DIBs for advanced single-molecule analysis.

Conclusions:

  • DIBs offer a powerful in vitro system for studying membrane protein behavior.
  • The DIB-TIRF microscopy approach provides valuable insights into protein dynamics and interactions.
  • This method enhances our understanding of cell membrane molecular mechanisms.